Inflamación

ANRV281-BE08-04 ARI 12 June 2006 17:8
Analysis of Inflammation
Geert W. Schmid-Schönbein
Department of Bioengineering, The Whitaker Institute for Biomedical Engineering, University of
Annu. Rev. Biomed. Eng. 2006.8:93-151. Downloaded from www.annualreviews.org
California San Diego, La Jolla, California 92093-0412; email: gwss@bioeng.ucsd.edu

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Annu. Rev. Biomed. Eng. Key Words

2006. 8:93–151
microcirculation, cell activation, signaling cascades, cardiovascular
First published online as a
Review in Advance on disease, auto-digestion,
April 17, 2006
Abstract
The Annual Review of
Biomedical Engineering is In the past, inflammation has been associated with infections and with the immune
online at system. But more recent evidence suggests that a much broader range of diseases have
bioeng.annualreviews.org
telltale markers for inflammation. Inflammation is the basic mechanism available for
doi: 10.1146/ repair of tissue after an injury and consists of a cascade of cellular and microvascular
annurev.bioeng.8.061505.095708
reactions that serve to remove damaged and generate new tissue. The cascade includes
Copyright
c 2006 by elevated permeability in microvessels, attachment of circulating cells to the vessels
Annual Reviews. All rights
reserved in the vicinity of the injury site, migration of several cell types, cell apoptosis, and
growth of new tissue and blood vessels. This review provides a summary of the
1523-9829/06/0815-
0093$20.00 major microvascular, cellular, and molecular mechanisms that regulate elements of
the inflammatory cascade. The analysis is largely focused on the identification of the
major participants, notably signaling and adhesion molecules, and their mode of
action in the inflammatory cascade. We present a new hypothesis for the generation
of inflammatory mediators in plasma that are derived from the digestive pancreatic
enzymes responsible for digestion. The inflammatory cascade offers a large number
of opportunities for development of quantitative models that describe various aspects
of human diseases.
93
ANRV281-BE08-04 ARI 12 June 2006 17:8
INTRODUCTION
Inflammation or the inflammatory process is a medical term, not an engineering one.
Since the early nineteenth century, the inflammatory process has been one of the
most intensively investigated areas of experimental medicine. Starting with Adamin
of McGill University (1), medical textbooks devoted major sections to inflammation,
and today a number of textbooks and conference proceedings are entirely focused
on the topic (2). The microcirculation is the major playground where the events in
inflammation can be studied and analyzed. In fact, one of the founders of modern
bioengineering, Benjamin W. Zweifach of the University of California, San Diego
(UCSD), a physiologist and a pioneer in microvascular research (3, 4), set out to
develop new quantitative approaches for the study of inflammation when he joined
a group of engineers at the California Institute of Technology. His effort led to the
formation of a formal alliance between engineering and medicine and the start of
bioengineering as an organized academic activity at UCSD.
Although recognized for its classical medical signs of acute pain sensation, heat,
redness, swelling, and eventual healing of the tissue with scar formation, from an
engineering point of view, inflammation may be more appropriately referred to as an
inflammatory cascade. It consists of a long chain of reactions and cellular activities that
serve to repair a tissue in many circumstances of life, from a small skin cut or repair
of tissue after birth to healing of the most severe burn injuries. The inflammatory
cascade at the tissue and cellular level involves a sequence of events with dilation of the
arterioles and venules, as well as increased blood vessel permeability and blood flow,
followed in many cases by stasis and thrombosis, infiltration of leukocytes into the
tissue, escape of plasma into the tissue, a breakdown of tissue by proteolytic activity
and oxygen free radical formation, necrosis and apoptosis, removal by phagocytic
cells, generation of new humoral mediators for cell growth, and regeneration of new
functional and connective tissue. The latter stage of inflammation has been referred
to as resolution of inflammation (5, 6) (Figure 1). An inflammatory cascade that does
not reach resolution leads to organ dysfunction and eventually death. Just about every
tissue has the ability to execute an inflammatory cascade. The inflammatory cascade is
preprogrammed and stereotypic. It is the only known mechanism for tissue repair after
injury and as such it is at the heart of medical thinking. Some of the most interesting
cellular processes (e.g., chemotaxis, phagocytosis, mitosis, cell differentiation) are
part of the inflammatory cascade.
The literature on inflammation is very rich and far beyond the scope of a single
review. Many medical journals are devoted to the subject and many more list in-
flammation as one of their topics of interest. I focus here on selected microvascular,
cellular, and molecular events and otherwise will refer to reviews. The discussion will
be limited to an outline of some key events in the inflammatory cascade. There is a
large literature on how the immune system (antibodies and cellular immunity) trig-
gers an inflammatory reaction and there is another large clinical literature that deals
with interventional studies against individual steps in inflammation. Without doubt,
future articles in this series will need to address selected aspects of the inflammatory
cascade.
94 Schmid-Schönbein
ANRV281-BE08-04 ARI 12 June 2006 17:8
Trigger mechanism
Immunological
Humoral via positive and negative feedback
Biomechanical
Physical transients
Non-resolved chronic inflammation

Cell response
Calcium entry and ion exchange
Receptor activation
Oxygen free radical formation
Upregulation of pro-inflammatory genes
Downregulation of anti-inflammatory genes
Inflammatory mediator release

Resolution of inflammation
Expression of adhesion molecules

Phagocytosis / debris removal Growth factor synthesis
Cell degranulation Angiogenesis
Pseudopod formation Tissue (scar) formation / remodeling
Microvascular consequences
Enhancement of endothelial permeability; cell degranulation
Leukocyte entrapment in capillaries
Reduction of tissue perfusion rate
Leukocyte/platelet aggregation, rolling and attachment
Migration into the tissue; chemotaxis
Monocyte and T-lymphocyte infiltration
Thrombosis
Apoptosis
Organ dysfunction
Minutes Hours Days/weeks
Figure 1
A schematic timeline of selected steps in the inflammatory cascade that either leads to
resolution and final repair of the initial injury, or under the influence of continued stimulation
progresses into a chronic inflammatory cascade.
INFLAMMATION AND DISEASE

Inflammation is present in patients with bacterial, viral, fungal or parasitic infections;
in anaphylaxis; in environmental diseases (smoke inhalation, asbestos exposure, etc.);
in rheumatoid arthritis, gout, autoimmune diseases, and intestinal diseases; and in en-
docrinological or autoimmune diseases; as well as in chronic diseases such as diabetes
(see, for example, any textbook on internal medicine).
But in the past three decades, it has also become evident that a much larger vari-
ety of diseases have telltale cellular and molecular evidence for inflammation. These
include chronic arterial and venous disease (7, 8), myocardial ischemia (9–11), acute
cerebral stroke and Alzheimer’s chronic disease (12–17), and more recently arterial
www.annualreviews.org • Inflammation 95
ANRV281-BE08-04 ARI 12 June 2006 17:8
hypertension (18) and cancer (19–21). For several decades there have been clinical
observations and recently also molecular evidence for inflammation in osteoarthritis
(22–24). A severe form of inflammation is observed in shock and multiorgan fail-
ure (25, 26), a condition with one of the highest mortalities. There are signs of
inflammation in patients with depression (27, 28) even without overt symptoms for
inflammation or just exposure to environmental risks (29). The list of diseases that
are associated with molecular markers of inflammation is large and growing. Since
introduction of effective and inexpensive measurements to detect signs of inflam-
mation in plasma of larger groups of patients [one of them is the c-reactive protein
(cRP) synthesized during inflammation in the liver together with other proteins like
fibrinogen], a wave of large-scale clinical studies provide supporting evidence for
the smaller experimental studies in the past (30–33). Clinical studies statistically link
inflammation to obesity (34, 35) and a lack of regular (moderate but not excessive)
exercise (36, 37). Inflammation is also a key problem in biomaterials dealing with
the events at the interface between a living tissue and a nonliving implant or a living
graft (38, 39) and in the design of blood substitutes (40). Therefore inflammation is
a central issue in tissue engineering. Today, inflammation has become the holy grail
for studies of human disease. Antiinflammatory treatments that have been shown to
be effective in one disease may turn out to be effective in another, thereby opening a
wider range of opportunities for intervention.
In the meantime, from a bioengineering point of view, no quantitative (i.e., pre-
dictive) model of the inflammatory cascade is available. But selected aspects have been
studied, although only within a narrow scope considering the cascade as a whole. Most
aspects of the inflammatory cascade are still at the beginning of a quantitative analysis,
and yet we are at one of the most important steps in the analysis, i.e., identification
of the tissues, cells, proteins, and genes that participate in the cascade. Medicine has
been deeply engaged; engineering science has to become involved as well. Tradi-
tional thinking of what constitutes a valid engineering analysis of well-defined field
equations and boundary value analysis, their prediction and experimental verification,
must be expanded. Identification of the players (among different cell types, thousands
of genes, proteins, lipids, and carbohydrates) has to become an integral and early part
of the engineering analysis. Identification of the molecular players constitutes signif-
icant progress and can often lead to new ideas for possible interventions. There are
numerous opportunities for diagnostic and interventional designs. The complexity
of the inflammatory cascade is one of the great opportunities for biomedical engi-
neering.
EXAMPLES OF INFLAMMATION IN THE

MICROCIRCULATION
The various forms of inflammation differ in large part by their location in the tissue,
the organ where they occur, and the nature and the severity of the tissue injury (e.g.,
mechanical injury, infectious agent, chemical injury, burn injury, radiation, tissue
injury owing to lack of organ perfusion, and oxygen supply).
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A Localized Endothelial Injury

A particularly localized form of tissue injury that permits experimental study of
in vivo inflammation almost at the level of single cells is by means of a narrowly
focused laser burn. Generation of a laser injury on microvascular endothelium leads
to rapid growth of a platelet thrombus over the injury site by attachment of platelets
to the injured endothelium and to each other with little entrapment of red cells.
Depending on the size and intensity of the laser injury, the thrombus may grow to
occlude the microvessel, but may also stop growing and detach from the injury site.
Detachment may occur in form of fragmentation of the thrombus, for example, from
its tip, but may also detach at the base of the thrombus where it is attached to the
endothelium, and thus this is the region in the thrombus with the lowest shear stress.
This evidence suggests that there is an enzymatic process that serves to cleave the
thrombus over the endothelium. Upon release the thrombus is carried by the blood
stream in the circulation, smaller thrombi are broken up as they pass through the cap-
illary network, and large aggregates may obstruct microvessels in which they become
entrapped. The original burn site may be subject to renewed platelet adhesion and
repetition of the cycle that start with a platelet thrombus grown followed by release
in form of an embolus. In the early phase of this form of tissue injury a few cells are
observed to migrate into the tissue (41–43).
A Tissue Burn
In contrast, if the laser burn is applied to a local tissue region in which entire cell
groups are injured, a somewhat different cellular sequence of events is observed. If
injury in the tissue is generated, one sees an elevation of microvascular permeability
in the adjacent blood vessels even though they may not have been hit by the laser
burn per se. Circulating leukocytes carried from central arteries that are unaffected
by the laser burn attach to the endothelium and eventually migrate toward the site of
the laser injury in a chemotactic process. Once leukocytes arrive at the laser injury,
their migration is arrested and they accumulate along the periphery of the burn injury
site. They phagocytose cell debris generated by the laser burn. In this injury model
there is less evidence of platelet adhesion to the neighboring microvessels, unless
the endothelium is actually injured. The leukocyte population initially consists of
neutrophils but is eventually followed by monocytes that upon migration into the
tissue differentiate into macrophages. In general, during the initial reaction the tissue
injury generated by the laser tends to grow in size but is eventually replaced by
new cells and even by growth of new blood vessels. The tissue that replaces the
burned tissue in many—but not all—cases is less-functional connective tissue (e.g.,
extracellular matrix, fibroblast, mast cells). The role of stem cells in regeneration of
functional tissue is largely unexplored in inflammation (44).
Ischemia
A different form of inflammation is observed in ischemia followed by reperfusion.
Any prolonged reduction of the blood flow in the microcirculation is followed by a
ANRV281-BE08-04 ARI 12 June 2006 17:8
period of reperfusion with typical signs of inflammation. Although cell injury may
occur already during ischemia, reperfusion frequently enhances injury or even death
of previously intact cells, the extent of which depends upon the duration and degree
of blood flow reduction during ischemia. Arterioles dilate, the endothelium in capil-
laries and postcapillaries develops an elevated permeability, leukocytes adhere to the
endothelium, individual capillaries may stop being perfused, tissue mast cells degran-
ulate, blood flow may even reduce as reperfusion is continued, and organ and cell
functions deteriorate to the point of complete failure. The tissue injury is enhanced
if the reduction of blood flow is accompanied by elevation of the microvascular blood
pressure (i.e., by an occlusion on the venous side of the circulation) as compared
to an occlusion on the arterial side (with reduction of microvascular pressure dur-
ing the occlusion) (45). Leukocytes also adhere to the endothelium and migrate into
the adjacent tissue, but are less directed than in the case of a local laser injury. In
more severe forms of ischemia/reperfusion, red cells may escape into the interstitial
space, especially from microvessels with elevated blood pressure (such as a venous
occlusion).
A similar form of inflammation can be generated by a number of proinflamma-
tory mediators, such as generation of platelet-activating factor (PAF, a membrane
fragment derived from phospolipids), the presence of complement fragments (com-
plement plays an important role in antigen-antibody reactions and is proteolytically
fragmented from a large initial complex into smaller bioactive peptides), cytokines
(small signaling proteins without enzymatic activity), or nitric oxide blockade. The
inflammation generated by such biochemical mediators does not require a reduction
of blood flow but qualitatively leads to similar microvascular manifestation of the
inflammatory cascade, as observed in ischemia/reperfusion.
Physiological Shock
A somewhat different inflammatory cascade can be observed in physiological shock
(e.g., owing to a major blood loss, a major burn, or trauma). In shock, organs that are
not involved in the initial injury may become victims of the inflammatory cascade.
The intestine or the pancreas are especially effective in this respect. The intestine
can generate a form of inflammation that spreads globally and causes injury in re-
mote innocent bystander organs that are not involved in the initial events that led to
inflammation in the intestine (e.g., by occlusion of the superior mesenteric artery).
This particular form of inflammation is accompanied by entrapment of platelets and
leukocytes in the microcirculation of remote organs (especially the lung and liver),
generation of blood coagulation and widespread capillary occlusion, elevated per-
meability and tissue swelling, aggregation of red cells, reduction of central blood
pressure, and cell apoptosis and parenchymal tissue failure in different organs (pul-
monary, renal, hepatic, and central nervous organ dysfunctions). Consequently, the
intestine is a source of inflammation in which multiple organs fail that were unaffected
by the original injury. It is the way many people die. We discuss this extraordinarily
important case of inflammation below.
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Inflammation Generated by Arthus and Shwartzman Reaction

A specialized form of inflammation was observed by Arthus after repeated injections
of horse serum into the skin of a rabbit. He noted that at first there was redness and
swelling. With each subsequent injection the reaction became more intense and fi-
nally led to necrotic lesions. The reaction can be evoked by several different materials
(bacterial infiltrates) and is accompanied by accumulation of leukocytes, degranula-
tion, and apoptosis.
Shwartzman in 1927 described a related inflammatory skin reaction, which now
bears his name (46). Injection of a filtrate of Bacillus typhosus in the skin by itself
produced only a mild reaction. If the initial injection is followed 24 h later by an in-
travenous injection with the same bacterial filtrate, a hemorrhagic and often necrotic
skin lesion is produced at the original injection site. The reaction can be produced with
different bacterial filtrates and with more purified endotoxin. In fact, many inflam-
matory materials of larger molecular weight that do not diffuse significantly from the
injection site produce a Shwartzman reaction. It requires the presence of neutrophil
infiltration into the tissue, which is the consequence of the primary injection.
There are as many forms of inflammation as there are ways to injure living tis-
sues; each plays out in its specific way. But all forms of inflammation under normal
conditions end up with a repaired tissue, whereas inflammation that goes unchecked
leads eventually to loss of organ function and even to the point of tissue necrosis if
the affected tissue does not belong to the vital organs (e.g., as seen in skin ulcers).
Even newborns have the ability to mount an inflammatory reaction, although in
some cases with attenuated level of inflammatory indicators (47, 48). Inflammation
tends to become more extensive and aggravated with age (49).
CELLULAR MARKERS OF INFLAMMATION

Although there are many approaches to study inflammation in living tissues of acute or
chronically instrumental models, the study of inflammation in man is still limited un-
less the inflammation is superficial and accessible by current microscopic techniques.
There is an evolving technology using microbubbles that adhere to inflammatory sites
and can be detected with whole-body imaging techniques (e.g., ultrasound) (50–53).
These may facilitate future direct study of inflammatory markers in individual organs
of patients.
In the past, the majority of studies of inflammation have relied on a collection of
venous blood samples, or to a limited degree urine or lymph samples. Such samples
will only be useful if the inflammation has reached a more advanced stage so that
markers of inflammation become globally detectable. There are then two opportu-
nities, testing either circulating cells and/or plasma.
Cell Activation Observed In Vitro

Early signs of inflammation can be detected by study of individual cells in the circu-
lation, e.g., from a venous blood sample. A good candidate is circulating leukocytes
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because they not only play a central role in inflammation but they are also readily
available from patients and volunteers for experimental and patient studies. Two gen-
eral approaches are possible: a study of the level of activation of circulating cells or
collection of plasma from patients. This plasma can then be applied to naı̈ve cells from
nonsymptomatic volunteers without inflammation or to a cell culture previously not
exposed to overt inflammatory stimuli. The two approaches give different informa-
tion. A study of the level of activation of circulating cells has to depend on those cells
that can actually be collected, for example, from an arm vein. But the circulating cells
collected in this fashion are all cells that have the ability to circulate, and therefore
they have low levels of cell activation. Fully activated leukocytes or platelets quickly
become trapped in the next capillary network they enter and stop circulating. Thus a
venous blood sample contains a subpopulation of cells with relatively low levels of ac-
tivation and as such is not representative of the cells in general and is not participating
in the local inflammation (54). In contrast, if plasma samples are used for analysis of
cell activation, it is possible to determine whether there are inflammatory mediators
present in plasma. Such evidence raises questions about the biochemical nature and
source of humoral activators. Inflammatory mediators suspended in plasma have the
ability to reach every organ, and they may play an important role in spreading cell
activation and inflammatory reactions from one organ to the next.
Pseudopod Formation
One of the most visible signs of cell activation is cytoplasmic pseudopod formation
(Figure 2). Pseudopods are local cytoplasmic projections generated by reorganiza-
tion of actin and other cytoskeletal proteins that can reach several micrometers in
size, have many different shapes, and are membrane covered. A microcirculation with
low levels of inflammation has few leukocytes with pseudopods. Pseudopods are the
essential element to give a cell the ability to spread and move on an adhesive substrate
(55). Pseudopods are generated by actin polymerization (56), triggered typically by
G protein–coupled receptor activation (for example, the formyl peptide receptor, the
interleukin-8 receptor, and the platelet-activating receptor) (57) via a signaling path-
way that includes phosphatidylinositol 3-kinase (PI3 kinase), phosphatidylinositol-P3
(PIP3 ), small guanosine triphosphatases (GTPases), Wiskott-Aldrich syndrome pro-
tein (WASp), and actin-related protein 2/3 (Arp2/3) as an actin-binding protein that
determines the spreading directions of pseudopods (57, 58). Pseudopod projection
is not necessarily Ca2+ ion dependent (59), and there exist signaling pathways that
are PI3 kinase–independent if leukocytes are primed (exposed but without full stim-
ulation) with insulin (60). Growth of pseudopods proceeds from the tip by diffusion
of G-actin toward a thin actin polymerization layer underneath the cell membrane
where actin monomers are inserted into the actin filaments, thereby pushing the
membrane outward (61). Projection of pseudopods may occur whether a leukocyte
is in free suspension or whether it is attached to endothelium or other substrates.
Pseudopods are projected and retracted in a cyclic fashion and typically grow at a
rate of approximately 10 μm/min. When exposed to specific agonists, their rate of
projection depends on agonist concentration (62).
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Figure 2
Scanning electron micrograph of a neutrophil in a nonactivated state (with red cell) (top panel)
and a neutrophil after activation (bottom panel ). The nonactivated cell has an overall spherical
shape with small membrane folds. After activation, the cell projects characteristic pseudopods
owing to actin polymerization. The process is frequently accompanied by expression of
membrane adhesion molecules, degranulation, and oxygen free radical production. Bar
length = 10 μm.
Pseudopods are essential elements for many inflammatory activities of circulating

leukocytes, be it spreading on a substrate or over a bacterium during phagocyto-
sis or projection during migration across the endothelium into the tissue. Different
leukocyte types have different pseudopod shapes, some of them are sheet-like (lamel-
lipodia) and others are finger-like (filipodia), and membrane-covered tethers formed
at points of cell retraction. The mechanics of projecting pseudopods are dominated
by the polymerization of actin filaments either at their tips underneath the membrane
or along preexisting actin fibers at the point of the actin-related protein (ARP2/3),
a process that permits spreading of the actin polymers into veil-like shapes (63). In
contrast, the mechanics of tethers are determined largely by membrane tension (64).
Even though pseudopods are mobile due to cyclic actin assembly and disassembly,
at any instant of time pseudopods have cross-linked fibrous actin, with mechanical
properties that are elastic with a minimal viscoelastic creep (65).

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Figure 3
Transmission electron
micrograph of activated
capillary endothelial cells
in rat skeletal muscle after
pressure reduction with
extensive pseudopod
formation (P). The
individual pseudopods
grow preferentially into
the lumen with less
growth into the basement
membrane (BM), and they
contain actin in a fibrous
form, but few caveolae

(C). J: cell junction.

Adapted from Reference
66.
Pseudopod projections are rarely detected on endothelial cells in normal blood

vessels. In contrast, lamellar-shaped pseudopods are observed in capillaries after blood
pressure reduction (Figure 3). They frequently appear at the tight junctions between
neighboring endothelial cells or are formed when endothelial cells in capillaries attach
to themselves to form a tubular structure that makes up the capillary. The cytoplas-
mic projections have a thin sheet-like shape that may reach several micrometers into
the lumen of microvessels. In capillaries with narrow vessel lumen, such cytoplasmic
projections are sufficient to interrupt the flow of blood cells (66). Cytoplasmic pseu-
dopods formed by endothelial cells are also observed when leukocytes migrate across
the endothelium (see below), when endothelial cells phagocytose, or when they mi-
grate during angiogenesis. Dewey and colleagues have introduced a direct technique
with fluorescently labeled actin to visualize cytoplasmic actin fibers in endothelial
cells and to study the turnover of actin in pseudopods (67, 68).
Platelets also undergo dramatic shape changes after activation. Besides transfor-
mation from their normal discoid shape into spheres (69), in a process that is tempera-
ture sensitive (70), they form cytoplasmic extensions that are finger shaped (filopodia)
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and may reach several micrometers by both calcium-sensitive and Rho-associated ki-
nase p160ROCK pathways. Filopodia contain actin bundles of long actin filaments
that end with an actin-capping protein at their tip. They also form filipodia bun-
dles that are abnormal in that they do not end at the filopodial tips but form actin
loops that return to the cell body (71). The discoid shape of platelets in the passive
state is maintained by a marginal band that consists of a single microtubule wound
into a coil (72).
Membrane Adhesion Molecules

An important form of cell activation in the microcirculation is the expression of mem-
brane adhesion molecules that facilitate the attachment of circulating cells to the en-
dothelium and to tissue structures over which they migrate. Two general classes of
adhesion molecules have been described in leukocytes, integrins, and selectins and
their ligands. They are preexpressed in the plasma membrane and also in cytoplasmic
granules, or may be newly synthesized upon stimulation. The topic is discussed in
detail in several recent reviews (73, 74); therefore I highlight only a few selected topics
important for microvascular inflammation. Initial leukocyte attachment to endothe-
lial membranes is facilitated by selectins, a group of three mammalian lectins (P-, E-,
L-selectin) capable of generating rapid and strong carbohydrate-protein bonds with
their ligands (P-selectin glycoprotein ligand-1, PSGL-1; sialyl Lewis X; and many
others). Leukocytes have L-selectin located preferentially on the tip of leukocyte
microvilli regions of the membrane that make initial attachment with the endothe-
lium (75). P-selectin in endothelial cells preexpressed in membrane bounds vesicles,
the Weibel-Palade bodies, a set of membrane-covered granules in their cytoplasm
(76) from which they can be released onto the plasma membrane (77), and there-
fore rapidly facilitate rolling attachment. P-selectin can also be induced by de novo
protein synthesis, a much slower process. E-selectin is predominantly on endothe-
lium and needs to be induced under inflammatory conditions. The lifetime of the
bond between P-selectins and their counter-receptors increases with applied force
but without a tendency for firm adhesion (78, 79), and vice versa, as the force is re-
duced the two molecules reduce the bond lifetime to the point that at low forces they
detach altogether (80) (designated as catch bonds). Evans and his colleagues have
identified the PSGL-1/P-selectin attachment to depend on metal-ion (Ca2+ ) bonds
and weaker hydrogen bonds (81).
In contrast, firm membrane adhesion and spreading of cells on the endothelium
are facilitated by integrins, a family of heterodimers, and tyrosine kinase receptors
that attach extracellular matrix adhesion sites to the structural proteins in the cell
cytoplasm and also serve molecular signaling. Leukocyte adhesion associated with
migration across the endothelium requires the engagement of integrins. The inte-
grins used by leukocytes share a common β2 -chain (CD18) and four different α-
chains (αl , CD11a; αm , CD11b; αX , CD11c; αmD , CD11d). One of the important
integrins in the inflammatory reaction is CD11b/CD18 (also known as complement
receptor 3 or Mac-1) expressed on neutrophils, monocytes and macrophages, and
natural killer lymphocytes. Mac-1 is presynthesized (like P-selectin) and stored in

ANRV281-BE08-04 ARI 12 June 2006 17:8
cytoplasmic granules and transported to the cell surface upon stimulation or linking of
L-selectin.
An interesting feature in inflammation is the fact that the integrin can be found
in different stages: a passive stage, an activated stage, and even intermediate stages.
Activation can be achieved by stimulation of the cells with humoral inflammatory
mediators, but it can also be achieved by fluid mechanical stress (H. Shin & G.W.
Schmid-Schönbein, unpublished results). In the activated stage, Mac-1 undergoes a
conformational change in its molecular structure that exposes an active binding site
(82).
Mac-1 can bind to a variety of ligands, an important one being the intercellular
adhesion molecule 1 (ICAM-1) expressed on specific endothelial cells in the venules
of the microcirculation (83). Arterioles and capillaries have less ICAM-1 expression.
ICAM-1 is a member of the immunoglobulin superfamily. Its expression is regu-

lated by transcription through mechanisms in the microcirculation that are largely

unknown. Animals with genetic deletions of ICAM-1 exhibit significant reduction
of inflammatory signals after challenge with endotoxin, ischemia/reperfusion, and
others.
Platelets have, in part, similar and also their own membrane adhesion molecules,
some of which are stored in granules and released upon stimulation (84). Platelet
P-selectin in the plasma membrane surface serves as a cell adhesion receptor to inter-
act with other cell receptors, including PSGL-1 and glycoprotein (GP) Iβ, a subgroup
of the platelets membrane glycoprotein (GP) Ib-IX-V complex (85, 86). Platelets also
utilize the glycoprotein (GP) Ib-IX-V complex to roll and slow down on a surface
coated with von Willebrand factor (vWF), a protein that can be released from the
Weibel-Palade bodies of activated endothelial cells (87, 88). They also use the GP
Ib-IX-V complex to bind to subendothelial extracellular matrix proteins (88). Binding
to collagen is facilitated by the platelet collagen receptors α2 β1 and GPVI. These two
molecules also play an important role in the activation of platelets (43). The integrin
α2b β3 binds platelets firmly to extracellular matrix proteins (collagen, fibronectin,
laminin, among others) (89). Platelet adhesion to the endothelium in turn serves to
facilitate adhesion of other cells to the endothelium or adhesion of platelets to other
circulating cells (86, 90–93).
The pathways that lead to expression and activation of the adhesion molecules in
inflammation have mostly been studied in acute situations, less in chronic conditions.
The interactions are complex. They may be redundant, in part owing to the fact that
specific adhesion molecules may only play a vital role under specific inflammatory
stimulations.
Oxygen Free Radical Formation

A major hallmark of cell activation in the microcirculation is oxygen free radical
formation (superoxide O2 − , hydrogen peroxide H2 O2 , the hydroxyl radical, nitric
oxide-related oxidants) (94–96). A number of techniques have been introduced that
permit detection of free radical formation in vivo with fluorescent indicators, so that
their location and time course can be studied. One of the earliest of these in vivo
ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 4
Selected micrographs of the rat mesentery with visualization of oxyradical-dependent
photoemission with ultrasensitive video intensifier microscopy during
endothelium-granulocyte interaction in microvascular beds treated with platelet-activating
factor (100 nM) (97). The photonic burst were detected with a chemiluminescence probe
(luminol) and superimposed digitally on a bright field image of the same region of the
mesentery (335). A: arteriole, V: venule. Reproduced with permission, courtesy of Dr. Makoto
Suematsu.
techniques was introduced by M. Suematsu and his colleagues at Keio University

using an ultrasensitive photon counting technique in conjunction with fluorescent
indicators that are reactive to individual free radical species (97). Stimulation of a
microcirculation leads initially to superoxide formation that is closely colocalized
with individual neutrophils and occurs in bursts (Figure 4). Stimulation over several
minutes to hours leads to the formation of free radicals in more than the neutrophils.
Multiple free radical species are detectable in the vicinity of the endothelium in
venular locations where neutrophils migrate across the endothelium and into the in-
terstitial space. A lower level of free radicals is formed in the interstitial space adjacent
to the arterioles or capillaries where fewer or no leukocytes are present (98). One of
the most toxic free radicals formed in vivo is the hydroxyl radical, which requires
an iron-catalyzed reaction (Haber-Weiss reaction) for its formation. Therefore, the
source of iron is an important consideration in understanding the oxygen free rad-
ical dynamics. A major source of iron is derived from hemoglobin (99, 100). Thus,
while fresh hemorrhages into a tissue may in fact be protective against oxygen free

ANRV281-BE08-04 ARI 12 June 2006 17:8
radical–mediated parenchymal tissue injury—owing to the presence of scavenging

molecules (like superoxide dismutase and catalase)—older red cells become highly
cytotoxic to the microcirculation because they are depleted of such scavenging capa-
bility and at the same time may be a source of iron that catalyzes the Haber-Weiss
reaction. Escape of aged red cells into the tissue causes enhanced parenchymal cell
death (101).
Degranulation
The variety of granules and lysosomes carried by most cells in the microcircula-
tion gives degranulation during inflammation a special significance. Degranulation
of membrane-covered organelles requires cytoplasmic rearrangement and membrane
fusion. Membrane proteins may be incorporated into the plasma membrane, or the
presynthesized contents in the organelles may be released into the surrounding tissue.
The broad variety of granule contents that can be released in inflammation leads to
numerous signaling possibilities.
One of the most visible signs for cell degranulation during inflammation is ob-
served in tissue mast cells, carriers of a prominent set of granules with histamine,
bradykinin, tryptase, and arachidonic acid derivatives, each of which in turn serve
to activate other microvascular cells (102). Mast cells can be degranulated by ni-
tric oxide suppression via a superoxide-mediated mechanism that in turn generates
leukocyte adhesion to the endothelium and triggers an inflammatory cascade with
elevated permeability (103, 104). Mast cell degranulation also plays an important
role in the initiation of inflammation in ischemia/reperfusion in an organ-dependent
way that can generate microvascular dysfunction in the intestine but less in skele-
tal muscle (105). The process can be mediated by histamine (106) and PAF (107)
and involves expression of P-selectin, likely owing to release and degranulation of
Weibel-Palade bodies. The degranulation of Weibel-Palade bodies facilitates release
of VWF, a molecule that serves to initiate coagulation and platelet adhesion (108, 109)
and of interleukin-8 (IL-8), which in turn mediates inflammation (76). Consequently,
degranulation of Weibel-Palade bodies and P-selectin expression on the endothelial
surface facilitates neutrophil adhesion, which stimulates release of proteolytic gran-
ules, carriers of a wide variety of antibacterial agents and lysosomal enzymes (110,
111). Hypoxia (112) and activated platelets (90), in addition to other stimuli (throm-
bin, epinephrine, or histamine), will also stimulate secretion of Weibel-Palade bodies
(76). Thus, there is an intricate network of cells, adhesion molecules, and humoral
signaling molecules that serves to control the inflammatory cascade at this step. No
quantitative engineering analysis exists.
Endothelial Glycocalyx
Recent evidence supports the notion that the glycocalyx, which grows by biosynthesis
of glycoaminoglycans on endothelial cells, may form a layer, which if intact, has an
antiinflammatory effect (113). The glycocalyx may reach a length that in some studies
was reported to be several hundred nanometers (114) and clearly exceeds the size
ANRV281-BE08-04 ARI 12 June 2006 17:8
of the extracellular domain of adhesion molecules such as ICAM-1. Even though

ICAM-1 is constitutively present on postcapillary venules, it serves only as an active
binding site if the glycocalyx is enzymatically cleaved during inflammation or after
ischemia/reperfusion (115).
Plasma Markers for Inflammation

The most direct and conclusive way to demonstrate the presence of inflammatory me-
diators in plasma is to mix the plasma with naı̈ve cells (such as leukocytes, platelets, and
endothelial cells) and measure markers of inflammation on those cells (such as pseu-
dopod formation, free radical formation, etc., as discussed above). But this approach
is less practical in large-scale clinical studies in which a large number of samples are
under investigation. The ideal way is to detect directly the molecule responsible for
the cell activation. But we do not know presently the specific molecule(s) that cause(s)
cell activation during inflammation, and much of the current evidence suggests that
there may be more than one inflammatory mediator present. Instead, it is necessary
to rely on specific molecules produced during inflammation. There are a number
of possible candidates. One class of molecules has been designated as acute-phase
proteins (also designated acute-phase reactants). They are synthesized in the liver,
neurons, lymphocytes, and monocytes during inflammation and released into the
plasma. These molecules include fibrinogen, c-reactive protein (cRP), serum amy-
loid A, α1-acid glycoprotein, α-1 antichymotrypsin, α-1 antitrypsin, haptoglobins,
ferritin, and others. The protein synthesis can be induced by cytokines (IL-1, IL-6).
cRP and fibrinogen levels have been found to be useful clinical indicators that have
low daily fluctuations (116, 117). An alternate approach is to measure the production
of oxygen free radicals in fresh plasma (118).
MICROVASCULAR MANIFESTATIONS
OF CELL ACTIVATION
Endothelial Permeability and Microhemorrhage
One of the earliest signs of inflammation in the microcirculation is an elevation of
endothelial permeability. Continuous endothelium becomes leaky for just about all
components contained in plasma, water, and ions, up to the size of large protein
species. The mechanisms by which permeability is elevated involves transmembrane
signaling and cytoplasmic reactions in the endothelial cells (119–121). A large number
of biochemical inflammatory mediators cause an elevation of permeability, as does
adhesion of leukocytes to the endothelium (122), or a lack of fluid shear stress acting
on the endothelial membrane (123). The most prominent and earliest site of elevated
permeability is in the postcapillary venules of the microcirculation, although arterioles
also contribute to transport of plasma proteins into the tissue (124). Early forms of
elevated permeability are accompanied by separation of VE-cadherins at intercellular
junctions, a situation that leads to rapid pore formation at intercellular junctions (125)
(Figure 5). The pores can close again by actin repolymerization (126).

ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 5
Scanning electron
micrograph of a
postcapillary venule with
continuous endothelium
in mesentery after
stimulation with an
inflammatory tripeptide
(f-Met-Leu-Phe) (10 nM,
1 h). The cell junctions
are recognized by the
enhanced ridges (arrows;
N: cell nuclei, P:
endothelial pore).
Individual pores after such

acute stimulation are

visible (insert below) and
positioned at the
junctions. Courtesy of Dr.
David K. Tung.
Pore formation in endothelium can also be observed inside endothelial cells, es-
pecially if the cells are mechanically stretched (127, 128), as is the case when stasis is
caused by occlusions on the venous side (45). Pores in or between endothelial cells are
facilitated by actin depolymerization and can be shaped by membrane tension (129).
McDonald and his colleagues also reported interesting pores that have membrane
tethers (126).
In more advanced stages of inflammation, one frequently observes extravasation
of red cells. The escape occurs across pores with a size that permits even transfer of
red cells and build up of microvascular hemorrhages. The egress of the red cells is
driven by convective flow, and their passage through pores is largely determined by
ANRV281-BE08-04 ARI 12 June 2006 17:8
the red cell ratio of membrane surface and cell volume (130). In the skin, such cell
extravasation leads to characteristic discolorations, which serve as clinical indicators
for various diseases.
An important element in any consideration of elevated permeability in microves-
sels is the structure of the basement membrane. Increasing evidence suggests that the
basement membrane is a mechanically strong structure that is able to carry most of
the mechanical stress in the wall of microvessels in spite of the fact that it is thin com-
pared to the thickness of the endothelium. Transfer of leukocytes across the vascular
wall requires proteolytic breakdown of the basement membrane (131), but the exact
proteolytic mechanism is still uncertain (132). In the case of neutrophils, this may be
achieved by release of elastase and matrix metalloproteinases, especially by gelatinase,
from their secondary granules after attachment of the neutrophil membrane to the
basement membrane via integrins (133). Breaks in the basement membrane are likely
repaired by the endothelial cells.
Leukocyte Infiltration into the Microcirculation

The accumulation of circulating neutrophils is one of the most visible signs of in-
flammation. To understand the interaction between leukocyte or platelets and the
endothelium in the microcirculation, we need to recognize that this is a flow domain
with low Reynolds number and virtual absence of inertial effects. Other than a mild
radial migration of the highly flexible erythrocytes away from the endothelium and
formation of a thin stochastic plasma-free zone in larger venules, stiffer cells such as
platelets and leukocytes experience negligible radial dispersion unless they interact
hydrodynamically with red cells in a velocity shear field.
In acute local inflammation, neutrophils (which exhibit few signs of activation
while in the free circulation) accumulate in postcapillary venules (typically about 10
to 12 μm in diameter and the first site in the venular microcirculation with ICAM-1
expression) where they are forced by a specific hydrodynamic interaction with red
cells to make membrane contact with the endothelium (Figure 6). The interaction
is brought about because the larger and stiffer leukocytes move with a lower velocity
than the red cells, leading to their accumulation upstream of leukocytes (134). In the
absence of red cells, no leukocyte makes membrane attachment with the endothe-
lium in postcapillary venules (135). Leukocytes may also be forced to make membrane
attachment with the endothelium at confluent venular bifurcations (136). The neu-
trophils that are attached to the endothelium are subject to a mechanical moment
that forces the cell to roll on the endothelium. The shear force distorts the cell into
a teardrop shape during rolling (137). The centroid velocity of a typical neutrophil
rolling on the endothelium is of the order of 1%–3% of the centerline red cell ve-
locity. The rolling velocity varies nonlinearly with the applied shear rate (138, 139),
it depends on the distribution and tethering of selectin to the cytoskeleton (78), and
appears to reach a threshold that may be L-selectin-specific because it can be observed
in cell-free systems (140). Neutrophil rolling on the endothelium in the narrow post-
capillary venules is almost always present, with or without inflammation, owing to the
inevitable interaction with red cells. Without inflammation, however, the neutrophils,

ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 6
Schematic diagram displaying various stages of leukocyte interaction with the endothelium on
the venous side of the microcirculation. The initial point of leukocyte attachment is in
postcapillary venules ( position 1) and at small convergent bifurcations ( position 2) where the
leukocytes are displaced by red cells to the endothelium. Depending on the presence and
activity of endothelial membrane adhesion molecules, the leukocytes roll on the endothelium,
stop rolling and responding to the fluid shear stress applied by the blood flow in the venule
( position 3), spread on the endothelium, and eventually may actively squeeze between and/or
through the endothelium and enzymatically break through the basement membrane to
actively migrate into the adjacent connective tissue. Leukocyte attachment to the venules
critically depends on the presence of the red cells.
as they enter larger venules, detach from the endothelium by random shear force in-
teractions with the red cells. In contrast, in inflammation, the rolling interaction
persists into the largest venules owing to the enhanced expression of selectins on
the endothelium and the fact that in inflammation the red cells may exhibit enhanced
membrane aggregation, and red cell aggregates displace smaller leukocytes on average
ANRV281-BE08-04 ARI 12 June 2006 17:8
toward the endothelium (141–144). Neutrophils roll on P-selectin, E-selectin (145,

146), and on their own L-selectin, and seem to involve a ligand protein [P-selectin
glycoprotein ligand-1, (147)]. Monocyte rolling can also be mediated by selectins and
their ligands or α4 β1 integrin interacting with endothelial VCAM-1 (148).
In acute inflammation, the main adhesion molecule used for rolling is P-selectin
on the endothelium, with PSGL-1 as the counter-receptor on the neutrophil (149,
150). While the cell rolls along the endothelium it is subject to increased activation
with elevation of intracellular Ca2+ (151) through multiple mechanisms, including
G protein–coupled receptors (such as the IL-8 receptor), the L-selectin/PSLG-1
interaction, attachment to E-selectin, and activation of CD18 (152).
Leukocyte rolling on the endothelium has received an extraordinary amount of
attention, and a variety of biophysical models have been developed. Early models
described the membrane adhesion process in the form of adhesion or peeling energy,
and demonstrated that bond length and bond flexibility play an important role on
the rolling motion (153). Using an energy balance between applied fluid mechanical
energy, the energy dissipated in viscoelastic deformation in the cell cytoplasm to form
a rapidly changing contact region during rolling, and given the membrane adhesion
energy it is possible to make predictions of average rolling velocity (154–156). Dembo
and coworkers established a quantitative relationship between applied force, adhesion
molecule chemical rate constants, and transient and steady-state detachment veloci-
ties (157). Damiano and his colleagues proposed a model in which adhesion energy
density varies inversely with instantaneous rolling velocity and directly with instanta-
neous deformation (137). Zhao developed a stochastic model of the micromechanics
of cell rolling and provides an analytical method for treating experimental data and
separating the contributions of temporal fluctuations and population heterogene-
ity of measured rolling velocities (158). Hammer and his coinvestigators proposed
a sequence of models in which the adhesive dynamics were considered in the form
of individual bonds, with bond formation and breakage as stochastic events, while
maintaining a relationship between rolling, force, and kinetic constants (159, 160).
They developed models for the fluctuation in the rolling velocity about the average
velocity (161), rolling on E-selectin (162), models with microvilli (163), and mod-
els that take into consideration the hydrodynamic interaction between neighboring
rolling leukocytes (164), as well as a number of elegant in vitro systems to study the
bond interaction during rolling, a topic that is outside the scope of this review.
Leukocyte rolling is followed by more firm adhesion to the endothelium. The ad-
hesion is facilitated by integrins and their counter receptors (e.g., ICAM-1, VCAM-1)
(165–167). A large number of publications have addressed this issue because blockade
of the adhesion serves to interrupt one of the key steps in the inflammatory cascade
involving leukocytes, and therefore raises the hope to interrupt inflammation as a
therapeutic intervention (168). Neutrophil adhesion to the endothelium is mediated
by β2 integrins. They become activated and in turn activate an extensive intracellular
molecular signaling network. On the apical surface of the endothelial cell, bound
chemokines (e.g., MCP-1, MIP-1α/β) can activate leukocyte β2 integrins for tight
adhesion to ICAM-1 and ICAM-2. I refer the reader to a previous review by Simon
(73).

ANRV281-BE08-04 ARI 12 June 2006 17:8
The site of leukocyte adhesion in the microcirculation is predominantly in post-

capillaries and venules with less but still occasional involvement of the arterioles.
Sites of leukocyte rolling and firm adhesion may be physically separated in the mi-
crocirculation but may also overlap. Little is known about this topographical aspect
of leukocyte adhesion in a microvascular network. The firm adhesion of leukocytes
is followed by pseudopod projection over the endothelium.
The transition to pseudopod formation in a venule is facilitated by stasis with
absence of fluid shear stress (169). But adhesion and cytoplasmic spreading can also
proceed in the presence of fluid flow in a microvessel if the leukocyte has been pre-
stimulated with an inflammatory mediator or if production of the second messenger,
cGMP, is suppressed by inhibition of nitric oxide (170). Thus, biochemical mediators
not only control adhesion and migration on the endothelium but also the ability of a
leukocyte to response to fluid mechanical stress. It is a truly interdependent process.

Fluid shear stress can impede adhesion, as may be the case in noninflammatory con-
ditions (169), but in the presence of inflammatory mediators that activate leukocytes,
it can also promote adhesion to the endothelium (171). This diverse set of reactions
to fluid shear stress is in line with the variety of shear stress responses observed in
leukocytes that depend on cofactors (170, 172). Migration on the endothelium may
persist, especially if the source of the inflammatory mediators is intravascular. The
migration may be in the direction of the fluid flow or opposite to it.
Trapping of Leukocyte in Capillaries

In contrast to the peripheral microcirculation referred to above, where most leuko-
cytes roll in venules, leukocytes in the lung accumulate in the capillaries (173). In
the peripheral circulation, entrapment of leukocytes and plugging of capillaries oc-
cur predominantly during periods of inflammation and central leukocyte activation.
There are two forms of leukocyte plugging in capillaries, transient and permanent
plugging (Figure 7). Transient plugging occurs almost exclusively at the endings of
the terminal arterioles (the smallest in the circulation), right at the entry into the
capillaries (174). It is present with and without inflammation. It is caused by transient
compression of leukocytes because passage of leukocytes through capillaries requires
deformation of the usually spherical cells (with a diameter of between ∼6.5 μm for an
average lymphocyte to ∼8 μm for neutrophils) into a compressed shape that fits into
the cylindrical lumen of capillaries (∼5 to 6 μm). The process requires compression
of the viscoelastic cell cytoplasm by a mechanical stress that is provided by the squeeze
pressure generated in a vessel with convergent lumen dimensions (175). As the cell
approaches such a capillary entry region, typically at a bifurcation, its cell centroid
velocity is rapidly reduced, and then it slowly creeps into the capillary lumen. At the
instant that the leukocyte fits into the lumen dimension, it rapidly speeds up again
and passes through the capillary (176). The typical entry time under physiological
conditions is approximately 1 s (as compared to the more rapid passage of a red cell
through the same entry region of approximately 10 ms) but the process is sensi-
tive with regard to the cytoplasmic rheological properties of the leukocyte and the
pressure drop across the capillary. There is no evidence for membrane attachment
ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 7
Schematic diagram displaying two cases of leukocyte capillary plugging at the bifurcation of
terminal arterioles (with single smooth muscle coat) into a capillary (case A) and inside a
capillary (case B). Case A shows the temporary plugging during the relatively slow deformation
of a leukocyte into an elongated shape ready to fit into the capillary lumen. The leukocyte
entry speed and deformation are determined by the pressure drop across the leukocyte, the
viscoelastic properties of its cytoplasm, and the exact dimensions of the capillary lumen and
the leukocyte. Activated leukocytes with stiffer cytoplasm tend to become more readily
trapped in capillaries, both at the entry and inside the capillaries. Case B shows permanent
plugging of leukocytes inside a capillary. Such permanent plugging leads to complete cessation
of cell motion, accumulation of red cells, and eventually apoptosis of the endothelial cells.
between leukocyte and endothelium during transient plugging of leukocytes. The

relatively low-pressure drop across the dense pulmonary capillary network makes
initial leukocyte attachment to the endothelium sensitive with regard to cytoplasmic
stiffness of leukocytes (e.g., pseudopod formation) and less dependent on adhesion
molecules (173, 177).
In contrast, permanent plugging of capillaries is caused by entrapment of leuko-
cytes in the lumen of a capillary and requires attachment of the leukocyte and capil-
lary endothelial membrane. The specific adhesion molecules involved in permanent
capillary plugging have not been determined. Permanent capillary plugging causes
occlusion of the capillary lumen (9, 17), a phenomenon that is responsible for the
capillary no-reflow phenomenon during periods of reperfusion following ischemia.

ANRV281-BE08-04 ARI 12 June 2006 17:8
The leukocytes in such obstructed capillaries are deformed into cylindrical cell shapes
that make broad membrane contact with the endothelial membrane. Just like transient
occlusion, permanent occlusion by leukocytes is also sensitive with respect to their cy-
toplasmic properties (178). Depending on the capillary network density, obstruction
of individual capillaries causes an increase in hemodynamic resistance in the micro-
circulation (179, 180) and an increase in capillary plugging enhances tissue apoptosis
(181). Depletion of leukocytes from the circulation attenuates the no-reflow phe-
nomenon in acute inflammation, reduces organ injury (182), and serves to maintain
tissue function in most organs tested (182–184).
Transendothelial Migration
In most inflammatory conditions, the leukocytes migrate within minutes after at-
tachment across and under the endothelium (185, 186) (Figure 8). The passageway
across the endothelium has been uncertain ever since the first electron microscopic
investigations of the phenomenon (187, 188). Much of the uncertainty has been fo-
cused around the issue of whether the leukocytes migrate through junctions between
endothelial cells or through new pores inside individual endothelial cells. While a
definitive analysis is not available today, much of the evidence points to both pos-
sibilities. The pathway selected by a neutrophil depends on the precondition of the
endothelial cells. In the case of continuous endothelial cells preexposed to low lev-
els of inflammatory mediators and to continuous fluid shear, the cell cytoplasm is
relatively thick and openings in the endothelium have been observed predominantly
between endothelial cells (paracellular) utilizing a PECAM-1 (a transmembrane pro-
tein and member of the immunoglobulin superfamily) (189, 190) interaction between
the monocyte and the endothelial cells, followed by similar homophilic adhesion
via CD99. Openings appear to involve lateral diffusion of interendothelial adhesion
molecules (VE-cadherins) (Figure 9) and opening of pores with actin depolymeriza-
tion. In contrast, endothelial cells that have a thin cytoplasm or endothelial cells with
fenestrations can have open pores inside (transcellular) the cells, especially if they are
subject to lateral stretch. In either case, leukocytes readily project pseudopods inside
either such pores. Neutrophils have the ability to migrate across pores, whether
made of cells or of artificial materials. In the pulmonary microcirculation, neu-
trophils frequently migrate across the endothelium at tricellular endothelial junctions
(191).
−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−−→
Figure 8
Transmission electron micrograph of venule in rat mesentery after 1 h stimulation with
f-Met-Leu-Phe. Different stages of the transendothelial migration by neutrophils (N) are
visible, from initial attachment with minimal membrane contact, to spreading and projection
of pseudopods through the endothelium (case B), to spreading of the endothelial cell over the
body of the transmigrating neutrophil (case A), as well as spreading of neutrophils on the
endothelial basement membrane and underlying connective tissue. There are some platelets
(P) in the vicinity of the site of transendothelial migration but this is not a requirement for
transendothelial migration. No extravasation of erythrocytes (E) is detectable.
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www.annualreviews.org • Inflammation
115
ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 9
Bright field and fluorescent images of the time course of a monocyte (red label ) migrating
across interendothelial junctions displayed by vascular endothelial (VE)-cadherin ( green
fluorescence). The VE-cadherin separates at the points where the monocyte crosses the
endothelium and reseals the interendothelial VE-cadherin after passage of the monocyte
(201). Courtesy of Dr. Francis W Luscinskas.
In acute inflammatory conditions, neutrophils may persist on the basement mem-

brane, form a contact area, and spread their cytoplasm (Figure 8). This evidence
clearly supports the hypothesis that the basement membrane is a mechanical barrier
for leukocyte migration. Breakage through the endothelial basement membrane is a
proteolytic process (192–196). It is interesting to note that most studies of the acute
inflammatory response have observed leukocyte migration (neutrophils, monocytes,
eosinophils, T-lymphcytes), whereas there are few studies that have reported platelet
migration across the endothelium in inflammation.
Neutrophils spread their pseudopods on the basement membrane underneath the
endothelium and slip across the pore, often aided by a contraction ring (a local con-
traction around the cell body that travels over the cell body) (65), while at the same
time the endothelial cell may project its own cytoplasm over the part of the leuko-
cyte body that is still inside the vessel lumen. This gives the appearance of a cup
shape by the endothelial cell (197). The neutrophil activates the myosin light chain
kinase activity and cAMP-protein kinase A activity, possibly by sliding ICAM-1 past
CD18 attachment sites (198). During passage the neutrophil and endothelial cell
membranes remain attached to each other. Studies with plasma protein labels point
to the fact that in the process of transendothelial migration, the seal between the
two cell membranes may remain tight without leakage of plasma or there may be
ANRV281-BE08-04 ARI 12 June 2006 17:8
transient plasma leakage depending on the particular choice of chemotactic agent

(199). The elevation of microvascular permeability during neutrophil transmigration
involves the kinetics of the focal adhesion kinase in the endothelial cells, suggesting
that contact with neutrophils leads to a cytoplasmic motion of the endothelial cells
(200). During transendothelial migration between endothelial cells under physiologi-
cal flow conditions, monocytes induce focal loss in the staining of the interendothelial
adhesion molecules VE-cadherin, alpha-catenin, beta-catenin, and plakoglobin (201),
suggesting that these adhesion molecules move laterally after dislodgement from the
underlying actin matrix and thereby facilitate interendothelial pore formation during
local actin depolymerization.
Upon breakage through the basement membrane, the leukocytes migrate in a
random pattern into the interstitial space with a direction that leads, on average,
away from the endothelium (202, 203) (Figure 10). Migration in the interstitium
requires attachment of neutrophils to extracellular collagen fibers via a mechanism

that involves β1 integrins (204). It has been proposed from studies on reconstituted
extracellular matrix proteins (fibrin) that β1 integrins also serve as regulators for con-
trol of neutrophil migration in a fashion that depends on the particular choice of the
chemotactic agent (205, 206). After penetration of the endothelial basement mem-
brane, monocytes migrate through the extracellular matrix of the tissues where they
may differentiate into tissue macrophages and/or migrate to sites of inflammation.
The migrating leukocytes displace perivascular cells, such as pericytes or fibroblasts
(207).
Additionally, monocytes in the tissues may migrate in the direction of the lym-
phatics and thereby return back into the bloodstream, both of which involve basal
to apical (reverse) transendothelial migration, possibly mediated by tissue factor and
p-glycoprotein (148).
It is also interesting to note that even though the neutrophils are one of the first
cell types to respond to an inflammatory signal, they may not necessarily participate
in large numbers in a local process of attachment and transendothelial migration, and
instead the majority of neutrophils continues to circulate (208).
Monocyte and Lymphocyte Infiltration

In inflammation the first wave of neutrophil migration into the tissue is followed by a
second wave of monocytes (cells that are much less frequent in the circulation). This
wave follows typically several hours after the initial inflammatory insult, but it persists
in chronic forms of inflammation when neutrophils may not be present anymore. The
monocyte infiltration is thus frequently observed in chronic inflammatory conditions,
be it in large vessels during atherosclerosis [an extensive literature exists on this topic
(7, 209, 210)] or in capillaries during chronic retinopathy (211). Monocytes also
utilize the β1 and β2 integrin to make firm membrane attachment. Upon migration
into the tissue, they differentiate into larger macrophages and phagocytose cell debris.
They carry a set of scavenger receptors (212) that permit binding of tissue degradation
products, such as oxidized low-density lipoprotein. They also synthesize and release a
number of growth factors that in the microcirculation lead to formation of a new set of

ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 10
Time-lapse record of a neutrophil (arrow) migrating across a venule (bottom of each panel ) into
the interstitial tissue of the mesentery after topical application of the f-Met-Leu-Phe. The cell
migrates, on average, away from the vessel wall, but is unaffected by the adjacent tissue mast
cell. M: mast cell, E: endothelial cell, L: venular lumen. From Reference 185.
ANRV281-BE08-04 ARI 12 June 2006 17:8
capillary blood vessels that can be observed to grow around clusters of macrophages
and give a coiled capillary appearance (211). The monocytes may accumulate cell
debris- or lipid-filled intracellular organelles in their cell cytoplasm, giving them a
morphological appearance of foam cells (213).
The mechanisms for attraction of monocytes to the lesion in arterial disease have
attracted intense interest. A family (with several subfamilies) of small, nonenzymatic
proteins, lymphokines, were identified that have characteristic cysteine inserts/repeats
(C, CC, CXC, and CXXXC) (214). One of these is the monocyte-chemoattractant
protein 1 (MCP-1), which can be synthesized by a number of cells. Deletion of
this cytokine or its receptors leads to reduction of atherosclerotic lesions in mice
that are otherwise susceptible to atherosclerosis (215). But in addition to MCP-1,
other chemokines have been implicated to serve as chemoattractant for monocytes
(reviewed in 209), but no conclusive analysis exists that would implicate any sin-
gle signaling molecule as the cause for the progression of monocyte infiltration in
atherosclerosis. Chemokines also influence other cell activities and may be one of
the sets of signaling molecules involved in the resolution of inflammation rather than
generation of inflammation.
Inflammatory sites are infiltrated by several other cell types. The thymus-derived
lymphocytes (T-lymphocytes) and some of their subtypes may accumulate in a chron-
ically inflamed site. These cells usually circulate and use a pathway through the lym-
phatics, which includes migration across a specialized endothelium in lymph nodes
and in Peyer’s patches (a specialized lymphoid tissue on the intestinal wall), and they
are regularly observed in the spleen. T-lymphocytes infiltrate tissue without overt
signs of inflammation but also migrate into almost any site of inflammation, also
utilizing selectins to initially adhere to the endothelium. The use of animals with
specific gene deletions has opened a new level of analysis into this extensive topic
closely associated with specific immunity [see, e.g., reviews on selectins and integrins
in T-lymphocyte adhesion (216, 217), the specific antigen mediated T-lymphocyte
activation (218), T-lymphocyte/monocyte interaction in chronic rheumatoid arthri-
tis (219) and in chronic pulmonary inflammation (220)]. Other cell types that may
infiltrate inflammatory sites are fibroblasts, smooth muscle cells, and mast cells.
Microvascular Cell Death

Inflammation is accompanied by both programmed (apoptotic) and necrotic cell
death, with and without attraction of inflammatory cells, such as neutrophils. A variety
of useful indicators have been introduced to detect cell death during inflammation in
a living microcirculation. The extent of cell death depends on the biochemical nature
and the level of the inflammatory stimulus. In acute ischemia and reperfusion, most
cell death of parenchymal tissue cells occurs during the period of reperfusion (Figure
11).
The presence of neutrophils per se does not necessarily lead to cell death. For
example, in the mesentery, after stimulation with a single inflammatory stimulus at
concentrations that lead to extensive neutrophil attachment to postcapillary venules
and migration into the mesentery tissue, no enhancement of parenchymal cell death

ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 11
Ischemia Reperfusion
Typical time course of cell
*
Number of PI(+) nuclei / area (n/mm2)

death in skeletal muscle (as 1000
detected by the life-death
indicator propidium iodine) *
(top panel), plugging of
capillaries by leukocytes 800
(middle panel), and adhesion *
of leukocytes to
postcapillary venules (bottom
600
panel) during inflammation
generated by ischemia and
followed by reperfusion. *
Ischemia (I group) by itself 400 Control

causes cell death, but there
is a dramatic enhancement I/R
of cell death if the ischemic I without R
period is followed by 200
reperfusion (I/R group).
Entrapment of leukocytes in
capillaries and venules is
enhanced during the course 0
of the reperfusion. Adapted
plugging WBCs (n/mm 2)
from Reference 222.

50
40
Number of
30 *
*
20
10
0
per 100 μm-length CV segment
*
12
Number sticking WBC
10
*
8 *
*
6
0
0 15 30 45 60 75 90 10 120 135 150
Time (min)
ANRV281-BE08-04 ARI 12 June 2006 17:8
can be detected (202, 221). The lack of parenchymal cell death is present over several
hours irrespective of the particular choice of inflammatory mediator that is used to
stimulate the inflammatory reaction (the synthetic tripeptide f-Met-Leu-Phe; PAF;
complement fragment C3a; tumor necrosis factor-α, TNFα). In contrast, if a com-
bination of two inflammatory mediators is applied at the same time irrespective of
their particular choice, then besides infiltration of leukocytes into the tissue, extensive
parenchymal cell death occurs, spatially correlated with the neutrophil accumulation.
The cell death observed under acute inflammatory conditions depends also on
physiological and metabolic activities. In skeletal muscle with a mixture of oxidative
and glycolytic muscle fibers, the first cells that are subject to cell death are fibers with
low oxidative metabolism, and therefore low levels of oxygen free radical scavengers.
Muscle fibers with high oxidative metabolism have a number of enzymes that scavenge
oxygen free radicals and therefore experience lower levels of cell death. In rat skeletal
muscle, the endothelial cells xanthine oxidase is a major source of oxygen free radicals
that kills the glycolytic muscle fibers (222).
Even though a number of mechanisms are well recognized to produce cell apop-
tosis in vitro (via death receptors: Fas and TNFα receptor 1, reactive oxygen free
radicals, a lack of integrin binding, and others), the molecular mechanisms that actu-
ally cause cell death in a living tissue in inflammation are less explored. The ability to
produce or inhibit apoptosis depends on cofactors. For example in endothelial cells,
application of fluid shear stress prior to exposure to hydrogen peroxide or TNFα
serves to prevent apoptosis (223, 224).
Besides parenchymal cells, cells that infiltrate into the tissue during inflammation
are also subject to apoptosis. Neutrophils have a natural average lifespan of less than
half a day, whereas mature peripheral T-lymphocytes have an estimated lifetime of 1–2
months. Interestingly, neutrophils at inflammatory sites have a delayed apoptosis (225)
that seems to depend on the presence of a growth factor (granulocyte/macrophage
colony-stimulating factor) released from activated endothelial cells. In the presence
of proapoptotic signals, such as tumor necrosis factor, Mac-1 engagement accelerates
apoptosis (226). Apoptotic neutrophils in the circulation are removed by the spleen,
or after migration into the peripheral microcirculation, by macrophage and fibroblast
phagocytosis (227, 228).
Capillary Restructuring Under Inflammation

Capillary and microvascular restructuring may occur at any stage of inflammation.
Many different shape and microvascular network changes can be observed. A typical
event, depending on the nature and duration of the inflammatory stimulus, is perma-
nent restructuring of the endothelium to the point that capillaries are converted and
enlarged into venules (229). Capillaries may disappear altogether, a process referred
to as rarefaction (230) and largely due to endothelial apoptosis (231). Resolution
of inflammation is associated with angiogenesis that may also go through several
phases. For example, inside a blood clot, new vessels will grow into the blood clot
from preexisting channels that are formed only inside the blood clot. The initial chan-
nels are highly irregular in shape and may have high network density but have no

ANRV281-BE08-04 ARI 12 June 2006 17:8
endothelial cells. Once endothelial cells start to grow into the newly formed channels,
these channels assume a morphology more typical for microvessels with narrow diam-
eters. The in-growth of the initial channels takes days; formation of a more complete
tissue with intact arterioles, nerves, and lymphatics requires several weeks (232). This
is an area important for further investigation because clinical interventions against
inflammation need to be evaluated in light of this repair process.
TRIGGER MECHANISMS FOR INFLAMMATION

An important consideration to control the level of inflammation is to identify molec-
ular mechanisms that can trigger an inflammatory reaction. To achieve this, it is
necessary to detect inflammation at an early stage. There are a large number of
mechanisms to activate cells and stimulate an inflammatory cascade. It is convenient

to classify mechanisms for cell activation into several general categories (233):
(a) Positive feedback mechanisms: There exists a class of inflammatory reactions that
are mediated by direct action of plasma inflammatory stimulators. A list of me-
diators proposed in the past include endotoxins, oxygen free radicals (234), PAF
(235), cytokines (a family of small molecular signaling proteins, e.g., TNF-α,
IL-1, IL-8) (236, 237), polypeptides [such as bradykinin, complement frag-
ments (238)], coagulation and fibrinolytic factors (239, 240), thrombin (241,
242), leukotrienes (243), histamine (106), and oxidized LDL (244, 245). The
list of inflammatory mediators is long indeed, and may in part be triggered by
trauma or by bacterial, viral, or fungal sources.
(b) Negative feedback mechanisms: An alternative pathway for cell upregulation in
the microcirculation is by depletion of antiinflammatory factors. This list is
shorter and includes nitric oxide (246), adenosine (247), adrenal glucocorticoids,
and selected cytokines (e.g., IL-10) (248). The reaction of these mediators is
nonlinear and depends on cofactors and concentrations.
(c) Contact activation: A specific form of cell activation by membrane contact has
been proposed in the form of juxtacrine activation (249). A nonactivated en-
dothelial cell may be stimulated during membrane contact by an activated leuko-
cyte and vice versa, e.g., by oxygen free radical production in the membrane
contact region between the cells and by formation of PAF and other bioactive
lipids.
(d) Activation by mechanotransduction: Alternative forms of cell activation owing to
either a shift to unphysiologically low or high fluid shear stresses acting on
the endothelium or a shift in the oxygen supply to the tissue (169, 250). Fluid
shear serves as a control mechanism for various forms of cell activation and
the expression of antiinflammatory and proinflammatory genes. Inflammatory
stimulators of the sort listed above can influence the fluid shear response via a
cGMP mediated mechanism (170) (see below).
(e) Activation by hyperglycemic/hyperosmolar conditions: A form of inflammation that
may be relevant to diabetes (251).
(f) Activation by physical transients: Transients of gas (like oxygen, carbon dioxide,
etc.) concentrations and temperature transients have the ability to stimulate
ANRV281-BE08-04 ARI 12 June 2006 17:8
cell activation irrespective of the direction of the transient (up or down) but
dependent on the magnitude of the transient (252–256).
Over the lifetime of a person, it is likely that several—if not all—of these mech-
anisms may at one time or the other stimulate inflammation. The challenge is to
identify dominating mechanisms. In the following, I discuss a source of inflammation
in physiological shock, one of the conditions with the highest level of inflammation
and also one of the conditions with highest mortality.
THE SOURCE OF INFLAMMATORY

MEDIATORS IN SHOCK
Inflammatory Mediators in Plasma

Plasma of animals (and patients), after intestinal ischemia and reperfusion as well
as in shock conditions, contains factors that exhibit a variety of biological activities,
including activation of circulating leukocytes, depression of T-lymphocyte prolifera-
tion, depression of cardiac muscle activity (the myocardial depression factor), carcino-
genic activity (clastogenic activity), leukocyte activating factor, and others (257–262).
There are many reports in the literature of pancreatic enzymes releasing active fac-
tors affecting heart tissue (257), with the hypothesis that the myocardial depression
factor is released from ischemic pancreas during shock. Recently, it was observed that
a chemotactic factor is released from self-digested spleen (263). Thus, what we regard
to be an inflammatory factor in the microcirculation during ischemia, for a number
of parenchymal cells may be a depressing factor. But none of these terms are entirely
adequate to describe the pancreatic protease-derived factor(s) encountered, consid-
ering the fact that one or more of the factors lead to direct cell death irrespective of
the presence or absence of leukocytes (264).
Traditionally, the most studied candidates for inflammatory mediators were bac-
teria in the intestine. Bacterial products in the form of lipopolysaccharide (endotoxin,
from the cell wall of bacteria) derived from gram-negative pathogenic bacteria (E.
coli, Salmonella, Shigella, Pseudomonas, Neisseria, Haemophilus, and other pathogens, in
contrast to Lactobacillus, Bifidobacteria, and Clostridium difficile, probiotics that support
digestion) can produce a strong inflammatory reaction (265) that can be as lethal as
hemorrhagic shock (266). There is an extensive literature on this topic (267). One
can find strong inflammatory mediators in the plasma, but their biochemical nature
and source have remained uncertain. The endotoxin and bacterial translocation hy-
pothesis states that endotoxin or whole intestinal bacteria may be transported from
the lumen of the intestine across the epithelial mucosal (brush border) barrier into
the wall of the intestine, where they are absorbed by the lymphatics or blood vessels
and carried back into the central circulation and produce inflammation (268). But
this hypothesis is not supported by conclusive evidence (269, 270).
We have recently obtained evidence that inflammatory and even cytotoxic me-
diators can also be derived from digested foods in the intestine (271). Fully char-
acterized bioactive peptides from milk and casein have been described as having a
wide range of functions (e.g., antimicrobial, immunostimulatory, immunosuppressive,

ANRV281-BE08-04 ARI 12 June 2006 17:8
antihypertensive, opioid-like, and mineral binding) (272–275). Cytotoxic peptides

have been identified in trypsin digests of casein (276, 277). Consequently, the lumi-
nal content of the intestine is a complex toxic mixture of mediators and factors that
are derived from digested foodstuff and the autolytic breakdown of the pancreatic
enzymes themselves.
Pancreatic Enzymes and the Production

of Inflammatory Mediators
It has been known for a long time that the intestine plays a central role in shock. The
collective set of pancreatic enzymes has the ability to digest virtually all biological
molecules except cellulose. This ability is one of the main requirements for normal
digestion. Digestive enzymes are discharged from the pancreas in the form of zy-
mogens and are activated by enterokinases in the duodenal segments of the upper
ileum (278). From there on, the digestive enzymes in the ileum are fully activated as
part of normal digestion until they are degraded and/or auto-degrade as they pass
through the lumen of the ileum. Under most circumstances, the digestive enzymes
are present in high concentrations in the entire ileum, cecum, and large intestine
(279). Even though food proteins, lipids, and complex carbohydrates are digested as
part of normal nutrition, digestion of the intestinal tissue itself appears to be largely
prevented. The mucosal barrier formed by the villi in the intestine minimizes self-
digestion by the activated pancreatic enzymes in the intestine. Two major mechanisms
are known to prevent transport of digestive enzymes into the wall of the intestine: (a)
the tight barrier formed by mucosal epithelium and (b) mucus secretion from goblet
cells that lead to a net outward transport away from the brush border barrier. The
epithelial cell lining is usually impermeable to digestive pancreatic enzymes (with a
barrier permeability range below the size of digestive enzymes) (280–282). However,
under ischemic conditions, the usually tight epithelial barrier is compromised and
higher-molecular-weight materials may penetrate the interepithelial gaps and enter
into the interstitial space of the intestinal wall (270, 283–288).
The New Autodigestion Hypothesis

If pancreatic digestive enzymes should penetrate through the mucosal epithelial brush
border, could this lead to production of inflammatory mediators? We explored this
question in Reference 289. To identify a possible source of inflammatory mediators,
tissues from different organs were homogenized and incubated for 2 h at 37◦ C and
centrifuged. The ability of the supernatant from these organs to activate was tested on
naı̈ve leukocytes. The experiments yielded a surprising result. All organ homogenates
produced only low levels of cell activation factors, except for a dramatically elevated
level of cell activation induced by homogenate from the pancreas (Figure 12). Similar
high levels of cell activation were detected independent of the species used (pig, rat,
mouse pancreas) (290). A mixture of organ homogenates that by themselves produce
only low levels of inflammation with selected pancreatic enzymes (serine proteases,
lipases) produces high levels of inflammatory mediators. This evidence suggests that
ANRV281-BE08-04 ARI 12 June 2006 17:8
100
Neutrophil mortality (%)

# *
75
50
25
0
l rt r n n s e y g
n tro ea ive rai lee rea stin agm ne un
H L B Sp nc te hr Kid L
Co p
Pa In Dia
Organ digests
Figure 12
Neutrophil death (detected by propidium iodide labeling) after exposure to homogenates of
fresh rat tissues from different organs. While several organs generate low levels of cell death
(and activation), the pancreas homogenate generates high levels of cell death. The cell death is
due to generation of inflammatory mediators by pancreatic digestive enzymes (serine
proteases, lipases) and can be generated in a similar fashion if digestive enzymes are added to
other organs. These results indicated a unique role for digestive enzymes as sources for
generation of humoral inflammatory and cytotoxic mediators (291). Courtesy of Steve Waldo.
the production of inflammatory mediators by pancreatic digestive enzyme is not

unique to the pancreas but instead depends on where in the tissue and in the circulation
pancreatic enzymes are transported and activated. A key organ to consider in this
respect is the intestine, where all digestive enzymes are fully activated and present in
high concentrations.
The supernatant from pancreatic homogenate not only produces upregulation of
leukocytes but is highly cytotoxic and causes high mortality (289, 291). Infusion of
a pancreatic homogenate derived from one third of a rat pancreas into an adult rat
causes death within 6 min, on average. Superfusion of whole pancreatic extract on
the mesentery microcirculation of normal rats, without exposure to shock, produces
signs of inflammatory reaction. This includes rolling of leukocytes on postcapillary
endothelium, adhesion, migration into the mesentery interstitial tissue, production
of peroxide by microvascular endothelium, mast cell degranulation, and cell death,
as detected by the life/death indicator propidium iodide (105, 222, 292–294). Serine
protease inhibitors block the production of cytotoxic mediators produced by pancre-
atic homogenate.
As indicated, the pancreas may not only be a major source for inflammatory medi-
ators but all organs may become a source for inflammatory mediators in the presence
of pancreatic or other tissue enzymes, especially serine proteases and lipases (289).

ANRV281-BE08-04 ARI 12 June 2006 17:8
This observation is especially relevant with respect to the intestine. If the intestine is
carefully rinsed of digestive enzyme, it produces relatively low levels of inflammatory
mediators after homogenization. In contrast, if the intestine (or any other organ)
is incubated with pancreatic enzymes, such as trypsin, chymotrypsin, elastase, or a
combination thereof, its homogenate causes inflammatory cell activation at a level
that is comparable to that of the pancreas (291). In light of the fact that the intestine
has most of the activated digestive enzymes, the critical issue in the production of in-
flammatory mediators is the distribution of these enzymes within the interstitial space
of the intestinal wall. Thus an important aspect in understanding the production of
inflammatory mediators produced by self-digestion of autologous tissue structures is
to study in the future the transport and activation of the digestive enzymes across the
intestinal barrier.
Pancreatic Enzyme Blockade in the Lumen of the Intestine

One of the experiments that serve to examine the role of pancreatic digestive enzymes
in acute ischemia is to rinse the initial intestinal contents and block pancreatic en-
zymes in the lumen of the intestine with an inhibitor [e.g., with the synthetic serine
protease/lipase inhibitor 6-amidino-2-naphtyl p-guanidinobenzoate dimethanesul-
fate, ANGD (295)] before and during ischemia of the small intestine (296). Such an
intervention almost completely inhibits activation of circulating leukocytes during
the ischemia and reperfusion, and prevents the appearance of inflammatory media-
tors in portal venous and systemic artery plasma. All early symptoms of organ injury
are attenuated. While the mean arterial blood pressure in the controls on average
falls below 40% of a normal mean arterial pressure after two hours of reperfusion,
mean arterial blood pressure on average remains at 84% of preischemic values in the
ANGD group. Digestive enzyme blockade in the intestinal lumen preserves pulse
pressures close to control values; attenuates the morphological damage to the in-
testinal wall and the accumulation of leukocytes in the intestine, the liver, and lung;
and it prevents formation of lung edema as an important early sign of organ failure.
Several protease inhibitors, if applied into the lumen of the intestine, also block the
production of inflammatory mediators in the intestine (297, 298). After blockade of
the production of inflammatory mediators in the intestine, peripheral organs are also
protected against inflammatory reactions (299). The intestinal tissue produces only
low levels of activators in the absence of pancreatic enzymes, while in the presence of
digestive enzymes, powerful inflammatory mediators are produced in a concentrated
and time-dependent fashion. This protection against inflammation has been observed
in several species (279) and confirmed in a shock model that consists of a combina-
tion of trauma and hemorrhagic shock (300). Protection against inflammation is seen
even with partial blockade of digestive enzymes without complete lavage of the small
intestine (279).
Furthermore, we also have been able to demonstrate protection against microvas-
cular inflammation after blockade of digestive enzymes and in the presence of en-
dotoxin (301). These results indicate that even though endotoxin can serve as an
inflammatory mediator, its effect is significantly amplified by the action of digestive
ANRV281-BE08-04 ARI 12 June 2006 17:8
enzymes because endotoxin per se serves to elevate the permeability of the intestinal
barrier, so that self-digestion is initiated.
TISSUE MECHANISMS TO REDUCE INFLAMMATION
Preconditioning
In contrast to the Arthus and Shwartzmann reactions that enhance inflammation (see
above), there are mechanisms to reduce the level of inflammation in a fashion that
depends on tissue pretreatment. These are some of the most interesting phenomena
associated with inflammation but are not well understood.
An interesting phenomenon described by Jennings and his associates at Duke
University is designated ischemic preconditioning. Multiple interruption of the blood

flow to an organ for short periods of time (on the order of 1 min) serves to reduce
the level of inflammation and organ dysfunction compared with control ischemia
without such temporary blood flow interruption (302, 303). Such preconditioning
reduces the development of inflammation after ischemia. Reduction of inflammation
by preconditioning can be generated in different organs and can also be simulated by
short exposure to other inflammatory challenges (304–306).
Tolerance
A strong level of protection against inflammation can be achieved by chronic pre-
treatment with an inflammatory mediator for a period of several days, such as mild
trauma or sublethal endotoxin administration (307, 308). Pretreatment dramatically
reduces signs of inflammation in ischemia or in shock and is referred to as tolerance
(309, 310). Tolerance can be achieved by a variety of different stimuli, but the pre-
treatment needs to be maintained. If it is stopped, the tolerance vanishes after several
days. The protection associated with tolerance has been attributed to production of
glucocorticoids and suppression of the inducible form of NOS (311).
FLUID SHEAR STRESS CONTROL OF INFLAMMATION
Endothelial Cells
There is an increasing body of evidence to suggest that inflammation is under the
control of fluid shear stress in the absence of biochemical agonists. By now, all cells
in the microcirculation have been shown to be controlled by fluid shear. In addition
to the elongation of the endothelial cells in the direction of flow, the endothelial cells
under steady shear increase the thickness of their cytoplasm and develop a set of actin
bundles (actin stress fibers) with firm adhesion to their substrate via focal adhesion
sites. But in addition to these morphological responses, endothelial cells also produce
a variety of other responses when exposed to shear stress or a tensile stretch. These
responses depend on the details of the stress history that is imposed on the cells and
on the level of attachment to surrounding cells and to the substrate. The topic is large
and has been previously reviewed (312–316).

ANRV281-BE08-04 ARI 12 June 2006 17:8
Physiological levels of fluid shear stress (e.g., steady shear at 10 dyn/cm2 for several
hours) applied to endothelial cells transforms the cells into a state in which expres-
sion of antiinflammatory genes (such as eNOS, COX-2, and Mn-SOD) and signs
of inflammation or thrombosis are reduced (317, 318). Gene-chip analysis confirms
that a number of proinflammatory genes are upregulated by laminar shear stress over
one day (319) and also when the endothelial cells are located close to smooth mus-
cle cells (320). Laminar shear stress reduces cell attachment to the endothelium and
the susceptibility of the endothelial cells to proinflammatory mediators (321, 322).
In contrast, unsteady shear stress, especially if accompanied by reversal of the shear
direction or by cyclic strain, induces proinflammatory genes (323).
Shear stress controls a wide variety of genes, and therefore it is of interest to identify
the transcription factors and DNA binding sites that are sensitive to mechanical
stress. Shear stress regulation appears to occur by binding of certain transcription

factors, such as NFκB and Egr-1, to shear stress response elements (SSREs) (324)
that are present in the promoters of biomechanically inducible genes. The Kruppel-
like factor 2, a member of the zinc finger family transcription factors, which regulate
cellular differentiation and tissue development, is induced by laminar shear stress in
endothelial cell cultures and has a distinct antiinflammatory effect (325).
Leukocytes
Leukocytes also respond to fluid shear stress, but in a different fashion. The leukocyte
membrane is subject to a different fluid stress field than the membrane of an endothe-
lial cell, and in fact the shear stress leukocytes experience in the circulation depends
on whether they are in free suspension or attached to the endothelium (326). The
response depends on the prior activity of the cell. When neutrophils migrate on a
substrate with cyclic projection of pseudopods, fluid shear stress leads to retraction of
these pseudopods (169) (Figure 13). Pseudopod retraction is also observed in mono-
cytes. In contrast, when the neutrophils are maintained in a deactivated state prior to
shear, they exhibit a reversed response and project pseudopods (172). Pseudopod re-
traction is associated with downregulation of small GTPase Rac1 and -2 (327). This
behavior is in line with the enhanced migration of stimulated lymphocytes under
fluid shear (171). Yap & Kamm observed transient neutrophil activation upon entry
into a narrow fluid channel (328, 329). Zhelev and his colleagues showed pseudopod
formation after mechanical extension of a membrane tether on a neutrophil (56).
Thus, clearly, the activity of a major class of inflammatory cells is under the control
of biomechanical stresses.
Pseudopod retraction occurs at levels of fluid shear that are typical for the micro-
circulation, but the stresses are too small to evoke a passive viscoelastic response. In
addition to retraction of pseudopods, the leukocytes also show other responses to fluid
shear. Neutrophil adhesion to each other (homotypic interaction) is under the control
of fluid shear (73). Neutrophils shed CD18 on their membrane by proteolytic cleavage
with cathepsin B derived from their granules (330), and they produce oxygen free rad-
icals. Pseudopod projection in free suspension is sensitive with respect to fluid shear
stress (rather than shear rate) but depends on the presence of red cells (331). When
ANRV281-BE08-04 ARI 12 June 2006 17:8
Figure 13
Time course of neutrophil spreading and pseudopod formation immediately before ( panel a ),
during ( panels b to h ), and after ( panels i, j ) application of a fluid shear stress by fluid discharge
from an adjacent micropipette positioned about one cell diameter from the surface of the
migrating neutrophil. The pipette generates a nonuniform fluid shear stress over the surface of
the cell, with peak value of the order of 1 dyn/cm2 (169). The migrating neutrophil retracts is
pseudopods during the exposure to the fluid shear stress. Sporadic platelets are present in these
experiments because separation between blood cells by centrifugation was minimized since it
interferes with the fluid shear response in leukocytes (336). Courtesy of Peter Marschel.
neutrophils adhere via β2 integrins to their substrate they respond readily to fluid
shear, whereas during attachment to β1 integrins the response is attenuated (332).
The question arises, How do leukocytes respond to fluid shear stress in the pres-
ence of inflammatory mediators? If neutrophils are subjected to increasing concen-
trations of inflammatory mediators (e.g., PAF or tripeptide f-Met-Leu-Phe), a pro-
gressive reduction of the fluid shear response is observed (170). The reduction of the
response can be reversed by the second messenger, cGMP, controlled by nitric oxide.
This mechanism facilitates an enhanced leukocyte response to fluid shear stress if the
cell is adhered to an endothelial cell that is still synthesizing nitric oxide as compared
to an endothelial cell in inflammation that has enhanced levels of superoxide and less
nitric oxide.
In light of the fact that the fluid shear stress response is quite specific for individual
cell types, we recently explored the question, What particular mechanisms may be
involved in the mechanosensing of fluid shear stress on the cell membrane? A number
of structures have been proposed (ion channels, the glycocalyx, the bilipid layer, and
others), but no proposal has been advanced that provides a hypothesis that is in line
with the high specificity actually observed among different cell types after exposure
to fluid shear stress. Chien and his collaborators have observed, in the case of the
endothelial cell, that the vascular endothelial growth factor receptor 2 is activated
by fluid shear stress together with the same signaling pathways that are activated by
an agonist to this receptor (VEGFA) (333). Thus, this preliminary evidence suggests
that one of the actual sensors for mechanical shear stress may be the G protein—
coupled receptor or tyrosine receptors in the membrane. Such receptors are unique

ANRV281-BE08-04 ARI 12 June 2006 17:8
to each cell and may provide the high specificity associated with the fluid shear stress
response. We tested this hypothesis directly in the case of the neutrophil, focusing on
the formyl peptide receptor (FPR), a receptor with the highest constitutive activity
in this particular cell. Deletion of the FPR receptor with silencing RNA (siRNA)
and generation of a cell without this receptor lead to abolishment of the fluid shear
response. In contrast, transfection and expression of the receptor in a cell line with-
out other receptors that respond to fluid shear serve to generate a cell with a single
FPR population. Such cells respond to fluid shear (334). Thus, this initial evidence
suggests that the humoral signaling pathways and the biomechanical signaling path-
ways overlap in inflammation, so that the state of receptor activation depends on
biomechanical stress and on the mediators synthesized and released.
Even though shear stress serves to control inflammatory signals in cell cultures
and in several in vivo models, and even though there is a nonuniform velocity field
around the risk sites for inflammation at arterial bifurcations, there is no conclusive
evidence that atherosclerosis is caused by fluid shear stress abnormalities at those sites.
The same anatomic risk for shear stress–induced inflammation at arterial bifurcations
exists at younger ages or during periods of time when arteries have no overt signs
of inflammation. Instead it is necessary to look at cofactors in patients. Patients may
have other trigger mechanisms for inflammation, such as infections, diabetes, or an
intestinal source for inflammatory mediators, to name just a few. These risk factors
generate their own proinflammatory state on which fluid shear stress is acting as an
antiinflammatory stimulus. It is a different situation compared to fluid shear stress
acting on noninflamed endothelium.
CONCLUSION
The study of inflammation is a key to understanding the origin and progression of
disease. While the inflammatory cascade is used during a lifetime as a tissue repair
mechanism, detection of signs of inflammation is now starting to be recognized as a
warning sign, indicating that the tissue is engaged in a repair that may or may not
come to a resolution. If it does not come to a resolution, cell death inevitably follows.
Some forms of interventions against inflammation have now been attempted at
several stages of the cascade, but with mixed results. This is not surprising if we con-
sider the facts that the inflammatory cascade serves primarily as a repair function and
that we do not know in many cases what caused the inflammation in the first place. To
achieve more control over the inflammatory cascade, it is essential to better under-
stand the actual trigger mechanisms that cause inflammation instead of interfering
only with subsequent events in the cascade.
In addition to traditional sources of inflammation, we have identified a funda-
mental mechanism for inflammation that is associated with the digestive system. The
digestive enzymes, an integral part of nutrient supply in the intestine, are present
over a lifetime and have the ability to digest any tissue with generation of power-
ful inflammatory mediators. Escape of the digestive enzymes from the lumen into
the wall of the intestine is prevented by the mucosal barrier in the intestine. Any
breach of this barrier leads to self-digestion and generation of powerful inflammatory
ANRV281-BE08-04 ARI 12 June 2006 17:8
mediators. This process has been shown to play a central role in experimental forms
of shock with high levels of inflammation.
Study of inflammation is one of the great problems of modern medicine and
it is most suitable to bioengineering analysis. Every new insight derived from the
engineering analysis will open the door to new and improved interventions.
ACKNOWLEDGMENT
Supported in part by NIH grants HL 10881, HL 43026, and HL 67825. I thank Dr.
Lars Bjursten for critical comments and many suggestions on the manuscript; Dr.
David K. Tung for providing Figures 5 and 8; and Peter J. Schmid-Schönbein for
preparation of Figures 2, 6, 7, and 8.
LITERATURE CITED
1. Adami J. 1909. Inflammation. An Introduction to the Study of Pathology. London:
McMillan
2. Gallin J, Goldstein I, Snyderman R. 1992. Inflammation. Basic Principles and
Clinical Correlates. New York: Raven Press
3. Zweifach B, Grant L, McCluskey R. 1965. The Inflammatory Process. London:
Academic
4. Zweifach B, Grant L, McCluskey R. 1974. The Inflammatory Process. I, II, III.
London: Academic
5. Henson PM, Larsen GL, Henson JE, Newman SL, Musson RA, Leslie CC.
1984. Resolution of pulmonary inflammation. Fed. Proc. 43:2799–806
6. Teder P, Vandivier RW, Jiang D, Liang J, Cohn L, et al. 2002. Resolution of
lung inflammation by cd44. Science 296:155–58
7. Ross R. 1999. Atherosclerosis—an inflammatory disease. N. Engl. J. Med.
340:115–26
8. Schmid-Schönbein GW, Takase S, Bergan JJ. 2001. New advances in the un-
derstanding of the pathophysiology of chronic venous insufficiency. Angiology
52(Suppl. 1):S27–34
9. Engler RL, Schmid-Schönbein GW, Pavelec RS. 1983. Leukocyte capillary
plugging in myocardial ischemia and reperfusion in the dog. Am. J. Pathol.
111:98–111
10. Entman ML, Michael L, Rossen RD, Dreyer WJ, Anderson DC, et al. 1991.
Inflammation in the course of early myocardial ischemia. FASEB J. 5:2529–37
11. Anselmi A, Abbate A, Girola F, Nasso G, Biondi-Zoccai GG, et al. 2004.
Myocardial ischemia, stunning, inflammation, and apoptosis during cardiac
surgery: a review of evidence. Eur. J. Cardiothorac. Surg. 25:304–11
12. Koistinaho M, Koistinaho J. 2005. Interactions between Alzheimer’s disease and
cerebral ischemia—focus on inflammation. Brain Res. Brain Res. Rev. 48:240–50
13. Iadecola C, Alexander M. 2001. Cerebral ischemia and inflammation. Curr.
Opin. Neurol. 14:89–94

ANRV281-BE08-04 ARI 12 June 2006 17:8
14. Jean WC, Spellman SR, Nussbaum ES, Low WC. 1998. Reperfusion injury after
focal cerebral ischemia: the role of inflammation and the therapeutic horizon.
Neurosurgery 43:1382–96; discussion 1396–87
15. del Zoppo GJ. 1997. Microvascular responses to cerebral is-
chemia/inflammation. Ann. N.Y. Acad. Sci. 823:132–47
16. Kontos CD, Wei EP, Williams JI, Kontos HA, Povlishock JT. 1992. Cytochem-
ical detection of superoxide in cerebral inflammation and ischemia in vivo. Am.
J. Physiol. Heart Circ. Physiol. 263:H1234–42
17. Del Zoppo GJ, Schmid-Schönbein GW, Mori E, Copeland BR, Chang C-M.
1991. Polymorphonuclear leukocytes occlude capillaries following middle cere-
bral artery occlusion and reperfusion. Stroke 22:1276–83
18. Suematsu M, Suzuki H, Delano FA, Schmid-Schönbein GW. 2002. The inflam-
matory aspect of the microcirculation in hypertension: oxidative stress, leuko-

cytes/endothelial interaction, apoptosis. Microcirculation 9:259–76

19. Li Q, Withoff S, Verma IM. 2005. Inflammation-associated cancer: Nf-κβ is
the lynchpin. Trends Immunol. 26:318–25
20. Karin M, Greten FR. 2005. Nf-κβ: linking inflammation and immunity to
cancer development and progression. Nat. Rev. Immunol. 5:749–59
21. Philip M, Rowley DA, Schreiber H. 2004. Inflammation as a tumor promoter
in cancer induction. Semin. Cancer Biol. 14:433–39
22. Benito MJ, Veale DJ, FitzGerald O, van den Berg WB, Bresnihan B. 2005.
Synovial tissue inflammation in early and late osteoarthritis. Ann. Rheum. Dis.
64:1263–67
23. Sturmer T, Brenner H, Koenig W, Gunther KP. 2004. Severity and extent of
osteoarthritis and low grade systemic inflammation as assessed by high sensi-
tivity c reactive protein. Ann. Rheum. Dis. 63:200–5
24. Hedbom E, Hauselmann HJ. 2002. Molecular aspects of pathogenesis in os-
teoarthritis: the role of inflammation. Cell. Mol. Life Sci. 59:45–53
25. Waxman K. 1996. Shock: ischemia, reperfusion, and inflammation. New Horiz.
4:153–60
26. Schmid-Schönbein GW, Hugli TE. 2005. A new hypothesis for microvascu-
lar inflammation in shock and multiorgan failure: self-digestion by pancreatic
enzymes. Microcirculation 12:71–82
27. Kop WJ, Gottdiener JS, Tangen CM, Fried LP, McBurnie MA, et al. 2002.
Inflammation and coagulation factors in persons >65 years of age with symp-
toms of depression but without evidence of myocardial ischemia. Am. J. Cardiol.
89:419–24
28. Toker S, Shirom A, Shapira I, Berliner S, Melamed S. 2005. The association
between burnout, depression, anxiety, and inflammation biomarkers: c-reactive
protein and fibrinogen in men and women. J. Occup. Health Psychol. 10:344–62
29. Pitzer JE, Del Zoppo GJ, Schmid-Schönbein GW. 1996. Neutrophil activation
in smokers. Biorheology 33:45–58
30. Ridker PM. 2004. High-sensitivity c-reactive protein, inflammation, and car-
diovascular risk: from concept to clinical practice to clinical benefit. Am. Heart
J. 148:S19–26
ANRV281-BE08-04 ARI 12 June 2006 17:8
31. Libby P, Ridker PM. 2004. Inflammation and atherosclerosis: Role of c-reactive
protein in risk assessment. Am. J. Med. 116(Suppl. 6A):9S–16
32. Kuo HK, Yen CJ, Chang CH, Kuo CK, Chen JH, Sorond F. 2005. Relation of
c-reactive protein to stroke, cognitive disorders, and depression in the general
population: systematic review and meta-analysis. Lancet Neurol. 4:371–80
33. Solem CA, Loftus EV Jr, Tremaine WJ, Harmsen WS, Zinsmeister AR,
Sandborn WJ. 2005. Correlation of c-reactive protein with clinical, endoscopic,
histologic, and radiographic activity in inflammatory bowel disease. Inflamm.
Bowel Dis. 11:707–12
34. Calabro P, Chang DW, Willerson JT, Yeh ET. 2005. Release of c-reactive
protein in response to inflammatory cytokines by human adipocytes: linking
obesity to vascular inflammation. J. Am. Coll. Cardiol. 46:1112–13
35. Nicklas BJ, Ambrosius W, Messier SP, Miller GD, Penninx BW, et al. 2004.
Diet-induced weight loss, exercise, and chronic inflammation in older, obese

adults: a randomized controlled clinical trial. Am. J. Clin. Nutr. 79:544–51
36. Lakka TA, Lakka HM, Rankinen T, Leon AS, Rao DC, et al. 2005. Effect of
exercise training on plasma levels of c-reactive protein in healthy adults: the
heritage family study. Eur. Heart J. 26:2018–25
37. Colbert LH, Visser M, Simonsick EM, Tracy RP, Newman AB, et al. 2004.
Physical activity, exercise, and inflammatory markers in older adults: findings
from the health, aging and body composition study. J. Am. Geriatr. Soc. 52:1098–
104
38. Anderson JM. 1988. Inflammatory response to implants. ASAIO Trans. 34:101–
7
39. Liu SQ, Moore MM, Glucksberg MR, Mockros LF, Grotberg JB, Mok AP.
1999. Partial prevention of monocyte and granulocyte activation in experimental
vein grafts by using a biomechanical engineering approach. J. Biomech. 32:1165–
75
40. Intaglietta M. 1999. Microcirculatory basis for the design of artificial blood.
Microcirculation 6:247–58
41. Grant L, Becker F. 1966. Mechanisms of inflammation. II. Laser-induced
thrombosis, histochemical considerations. Arch. Pathol. 81:36–41
42. Grant L, Becker FF. 1965. Mechanisms of inflammation. I. Laser-induced
thrombosis, a morphologic analysis. Proc. Soc. Exp. Biol. Med. 119:1123–29
43. Chen J, Lopez JA. 2005. Interactions of platelets with subendothelium and
endothelium. Microcirculation 12:235–46
44. Ramsey S, Grant L. 1974. Chemotaxis. See Ref. 4, pp. 287–362
45. Takase S, Lerond L, Bergan JJ, Schmid-Schönbein GW. 2002. Enhancement of
reperfusion injury by elevation of microvascular pressures. Am. J. Physiol. Heart
Circ. Physiol. 282:H1387–94
46. Shwartzman G. 1937. Phenomenon of Local Tissue Reactivity. New York: Harper
(Hoeber)
47. Wolach B, Sonnenschein D, Gavrieli R, Chomsky O, Pomeranz A, Yuli I. 1998.
Neonatal neutrophil inflammatory responses: parallel studies of light scattering,
cell polarization, chemotaxis, superoxide release, and bactericidal activity. Am.
J. Hematol. 58:8–15

ANRV281-BE08-04 ARI 12 June 2006 17:8
48. Johnston CJ, Oberdorster G, Gelein R, Finkelstein JN. 2000. Newborn mice
differ from adult mice in chemokine and cytokine expression to ozone, but not
to endotoxin. Inhal. Toxicol. 12:205–24
49. Harris NR, Rumbaut RE. 2001. Age-related responses of the microcirculation
to ischemia-reperfusion and inflammation. Pathophysiology 8:1–10
50. Yasu T, Greener Y, Jablonski E, Killam AL, Fukuda S, et al. 2005. Activated
leukocytes and endothelial cells enhance retention of ultrasound contrast mi-
crospheres containing perfluoropropane in inflamed venules. Int. J. Cardiol.
98:245–52
51. Weller GE, Villanueva FS, Tom EM, Wagner WR. 2005. Targeted ultra-
sound contrast agents: in vitro assessment of endothelial dysfunction and multi-
targeting to ICAM-1 and sialyl Lewis X. Biotechnol. Bioeng. 92:780–88
52. Bloch SH, Dayton PA, Ferrara KW. 2004. Targeted imaging using ultrasound
contrast agents. Progess and opportunities for clinical and research applications.
IEEE Eng. Med. Biol. Mag. 23:18–29
53. Lindner JR, Dayton PA, Coggins MP, Ley K, Song J, et al. 2000. Noninvasive
imaging of inflammation by ultrasound detection of phagocytosed microbub-
bles. Circulation 102:531–38
54. Wautier JL, Schmid-Schönbein GW, Nash GB. 1999. Measurement of leuko-
cyte rheology in vascular disease: clinical rationale and methodology. Interna-
tional Society of Clinical Hemorheology. Clin. Hemorheol. Microcirc. 21:7–24
55. Schmid-Schönbein GW. 1987. Rheology of leukocytes. In Handbook of Bioengi-
neering, ed. R Skalak, S Chien. New York: McGraw-Hill
56. Zhelev DV, Alteraifi AM, Hochmuth RM. 1996. F-actin network formation in
tethers and in pseudopods stimulated by chemoattractant. Cell Motil. Cytoskelet.
35:331–44
57. Chodniewicz D, Alteraifi AM, Zhelev DV. 2004. Experimental evidence for
the limiting role of enzymatic reactions in chemoattractant-induced pseudopod
extension in human neutrophils. J. Biol. Chem. 279:24460–66
58. Niggli V. 2003. Signaling to migration in neutrophils: importance of localized
pathways. Int. J. Biochem. Cell Biol. 35:1619–38
59. Alteraifi AM, Zhelev DV. 1997. Transient increase of free cytosolic calcium
during neutrophil motility responses. J. Cell. Sci. 110(Pt. 16):1967–77
60. Chodniewicz D, Zhelev DV. 2003. Novel pathways of f-actin polymerization
in the human neutrophil. Blood 102:2251–58
61. Zhu C, Skalak R. 1988. A continuum model of protrusion of pseudopod in
leukocytes. Biophys. J. 54:1115–37
62. Zhelev DV, Alteraifi AM, Chodniewicz D. 2004. Controlled pseudopod exten-
sion of human neutrophils stimulated with different chemoattractants. Biophys.
J. 87:688–95
63. Kelleher JF, Atkinson SJ, Pollard TD. 1995. Sequences, structural models,
and cellular localization of the actin-related proteins arp2 and arp3 from acan-
thamoeba. J. Cell Biol. 131:385–97
64. Marcus WD, Hochmuth RM. 2002. Experimental studies of membrane tethers
formed from human neutrophils. Ann. Biomed. Eng. 30:1273–80
ANRV281-BE08-04 ARI 12 June 2006 17:8
65. Schmid-Schönbein GW, Skalak R, Sung K-LP, Chien S. 1982. Human leuko-
cytes in the active state. In White Blood Cells, Morphology and Rheology as Related to
Function, ed. U Bagge, GVR Born, P Gaehtgens. The Hague: Martinus Nijhoff
66. Lee J, Schmid-Schönbein GW. 1995. Biomechanics of muscle capillaries:
hemodynamic resistance, endothelial distensibility, and pseudopod formation.
Ann. Biomed. Eng. 23:226–46
67. McGrath JL, Hartwig JH, Tardy Y, Dewey CF Jr. 1998. Measuring actin dy-
namics in endothelial cells. Microsc. Res. Tech. 43:385–94
68. McGrath JL, Tardy Y, Dewey CF Jr, Meister JJ, Hartwig JH. 1998. Simultane-
ous measurements of actin filament turnover, filament fraction, and monomer
diffusion in endothelial cells. Biophys. J. 75:2070–78
69. Paul BZ, Kim S, Dangelmaier C, Nagaswami C, Jin J, et al. 2003. Dynamic reg-
ulation of microtubule coils in adp-induced platelet shape change by p160rock

(rho-kinase). Platelets 14:159–69

70. Winokur R, Hartwig JH. 1995. Mechanism of shape change in chilled human
platelets. Blood 85:1796–804
71. Hartwig JH. 1992. Mechanisms of actin rearrangements mediating platelet ac-
tivation. J. Cell Biol. 118:1421–42
72. Italiano JE Jr, Lecine P, Shivdasani RA, Hartwig JH. 1999. Blood platelets are
assembled principally at the ends of proplatelet processes produced by differ-
entiated megakaryocytes. J. Cell Biol. 147:1299–312
73. Simon SI, Green CE. 2005. Molecular mechanics and dynamics of leukocyte
recruitment during inflammation. Annu. Rev. Biomed. Eng. 7:151–85
74. Ley K, Sperandino M. 2003. Molecular mechanisms of leukocyte adhesion. In
Molecular Basis of Microcirculatory Disorders, ed. GW Schmid-Schönbein, DN
Granger. Paris: Springer
75. Bruehl RE, Springer TA, Bainton DF. 1996. Quantitation of L-selectin dis-
tribution on human leukocyte microvilli by immunogold labeling and electron
microscopy. J. Histochem. Cytochem. 44:835–44
76. van Mourik JA, Romani de Wit T, Voorberg J. 2002. Biogenesis and exocytosis
of Weibel-Palade bodies. Histochem. Cell Biol. 117:113–22
77. Koedam JA, Cramer EM, Briend E, Furie B, Furie BC, Wagner DD. 1992. P-
selectin, a granule membrane protein of platelets and endothelial cells, follows
the regulated secretory pathway in att-20 cells. J. Cell Biol. 116:617–25
78. Alon R, Hammer DA, Springer TA. 1995. Lifetime of the P-selectin-
carbohydrate bond and its response to tensile force in hydrodynamic flow.
Nature 374:539–42
79. Zhu C, Lou J, McEver RP. 2005. Catch bonds: physical models, structural bases,
biological function and rheological relevance. Biorheology 42:443–62
80. Goldsmith HL, Quinn TA, Drury G, Spanos C, McIntosh FA, Simon SI.
2001. Dynamics of neutrophil aggregation in couette flow revealed by videomi-
croscopy: effect of shear rate on two-body collision efficiency and doublet life-
time. Biophys. J. 81:2020–34
81. Evans E, Leung A, Hammer D, Simon S. 2001. Chemically distinct transition
states govern rapid dissociation of single L-selectin bonds under force. Proc.
Natl. Acad. Sci. USA 98:3784–89

ANRV281-BE08-04 ARI 12 June 2006 17:8
82. Takagi J, Springer TA. 2002. Integrin activation and structural rearrangement.
Immunol. Rev. 186:141–63
83. Iigo Y, Suematsu M, Higashida T, Oheda J, Matsumoto K, et al. 1997. Con-
stitutive expression of ICAM-1 in rat microvascular systems analyzed by laser
confocal microscopy. Am. J. Physiol. Heart Circ. Physiol. 273:H138–47
84. Israels SJ, Gerrard JM, Jacques YV, McNicol A, Cham B, et al. 1992. Platelet
dense granule membranes contain both granulophysin and P-selectin (gmp-
140). Blood 80:143–52
85. Katayama T, Ikeda Y, Handa M, Tamatani T, Sakamoto S, et al. 2000. Im-
munoneutralization of glycoprotein ibalpha attenuates endotoxin-induced in-
teractions of platelets and leukocytes with rat venular endothelium in vivo. Circ.
Res. 86:1031–37
86. Furie B, Furie BC, Flaumenhaft R. 2001. A journey with platelet P-selectin:
the molecular basis of granule secretion, signalling and cell adhesion. Thromb.
Haemost. 86:214–21
87. Borthakur G, Cruz MA, Dong JF, McIntire L, Li F, et al. 2003. Sulfatides
inhibit platelet adhesion to von Willebrand factor in flowing blood. J. Thromb.
Haemost. 1:1288–95
88. Romo GM, Dong JF, Schade AJ, Gardiner EE, Kansas GS, et al. 1999. The
glycoprotein IB-IX-V complex is a platelet counterreceptor for P-selectin. J.
Exp. Med. 190:803–14
89. Konstantopoulos K, Kukreti S, McIntire LV. 1998. Biomechanics of cell inter-
actions in shear fields. Adv. Drug Deliv. Rev. 33:141–64
90. Dole VS, Bergmeier W, Mitchell HA, Eichenberger SC, Wagner DD. 2005.
Activated platelets induce Weibel-Palade-body secretion and leukocyte rolling
in vivo: role of P-selectin. Blood 106:2334–39
91. Kuijper PH, Gallardo Torres HI, Houben LA, Lammers JW, Zwaginga JJ,
Koenderman L. 1998. P-selectin and mac-1 mediate monocyte rolling and ad-
hesion to ECM-bound platelets under flow conditions. J. Leukoc. Biol. 64:467–73
92. Fernandes LS, Conde ID, Wayne Smith C, Kansas GS, Snapp KR, et al. 2003.
Platelet-monocyte complex formation: effect of blocking PSGL-1 alone, and in
combination with alphaiibbeta3 and alphambeta2, in coronary stenting. Thromb.
Res. 111:171–77
93. Konstantopoulos K, Neelamegham S, Burns AR, Hentzen E, Kansas GS,
et al. 1998. Venous levels of shear support neutrophil-platelet adhesion and
neutrophil aggregation in blood via P-selectin and beta2-integrin. Circulation
98:873–82
94. McCord JM. 2000. The evolution of free radicals and oxidative stress. Am. J.
Med. 108:652–59
95. Royall JA, Kooy NW, Beckman JS. 1995. Nitric oxide-related oxidants in acute
lung injury. New Horiz. 3:113–22
96. Beckman JS, Ye YZ, Chen J, Conger KA. 1996. The interactions of nitric oxide
with oxygen radicals and scavengers in cerebral ischemic injury. Adv. Neurol.
71:339–50; discussion 350–34
ANRV281-BE08-04 ARI 12 June 2006 17:8
97. Suematsu M, Kurose I, Asako H, Miura S, Tsuchiya M. 1989. In vivo visualiza-

tion of oxyradical-dependent photoemission during endothelium-granulocyte
interaction in microvascular beds treated with platelet-activating factor.
J. Biochem. 106:355–60
98. Suematsu M, Schmid-Schönbein GW, Chavez-Chavez RH, Yee TT, Tamatami
T, et al. 1993. In vivo visualization of oxidative changes in microvessels during
neutrophil activation. Am. J. Physiol.Heart Circ. Physiol. 264:H881–91
99. Grisham MB, Granger DN. 1988. Neutrophil-mediated mucosal injury. Role
of reactive oxygen metabolites. Dig. Dis. Sci. 33:6S–15
100. Smith SM, Grisham MB, Manci EA, Granger DN, Kvietys PR. 1987. Gastric
mucosal injury in the rat. Role of iron and xanthine oxidase. Gastroenterology
92:950–56
101. Wilms H, Delano FA, Schmid-Schönbein GW. 2000. Mechanisms of parenchy-

mal cell death in-vivo after microvascular hemorrhage. Microcirculation 7:1–11

102. Parikh SA, Cho SH, Oh CK. 2003. Preformed enzymes in mast cell granules
and their potential role in allergic rhinitis. Curr. Allergy Asthma Rep. 3:266–72
103. Kubes P, Kanwar S, Niu XF, Gaboury JP. 1993. Nitric oxide synthesis inhibition
induces leukocyte adhesion via superoxide and mast cells. FASEB J. 7:1293–99
104. Kanwar S, Wallace JL, Befus D, Kubes P. 1994. Nitric oxide synthesis inhibition
increases epithelial permeability via mast cells. Am. J. Physiol. Gastrointest. Liver
Physiol. 266:G222–29
105. Kanwar S, Hickey MJ, Kubes P. 1998. Postischemic inflammation: a role for
mast cells in intestine but not in skeletal muscle. Am. J. Physiol.Gastrointest. Liver
Physiol. 275:G212–18
106. Kubes P, Kanwar S. 1994. Histamine induces leukocyte rolling in post-capillary
venules. A P-selectin-mediated event. J. Immunol. 152:3570–77
107. Gaboury JP, Johnston B, Niu XF, Kubes P. 1995. Mechanisms underlying acute
mast cell-induced leukocyte rolling and adhesion in vivo. J. Immunol. 154:804–
13
108. Wagner DD. 1993. The Weibel-Palade body: the storage granule for von Wille-
brand factor and P-selectin. Thromb. Haemost. 70:105–10
109. Fredrickson BJ, Dong JF, McIntire LV, Lopez JA. 1998. Shear-dependent
rolling on von Willebrand factor of mammalian cells expressing the platelet
glycoprotein IB-IX-V complex. Blood 92:3684–93
110. Bainton DF. 1976. Neutrophil granules: a review. Am. J. Med. Technol. 42:15–21
111. Faurschou M, Borregaard N. 2003. Neutrophil granules and secretory vesicles
in inflammation. Microbes Infect. 5:1317–27
112. Pinsky DJ, Naka Y, Liao H, Oz MC, Wagner DD, et al. 1996. Hypoxia-induced
exocytosis of endothelial cell Weibel-Palade bodies. A mechanism for rapid
neutrophil recruitment after cardiac preservation. J. Clin. Invest. 97:493–500
113. Mulivor AW, Lipowsky HH. 2004. Inflammation- and ischemia-induced shed-
ding of venular glycocalyx. Am. J. Physiol. Heart Circ. Physiol. 286:H1672–80
114. van den Berg BM, Vink H, Spaan JA. 2003. The endothelial glycocalyx protects
against myocardial edema. Circ. Res. 92:592–94
115. Mulivor AW, Lipowsky HH. 2002. Role of glycocalyx in leukocyte-endothelial
cell adhesion. Am. J. Physiol. Heart Circ. Physiol. 283:H1282–91

ANRV281-BE08-04 ARI 12 June 2006 17:8
116. Black S, Kushner I, Samols D. 2004. C-reactive protein. J. Biol. Chem.

279:48487–90
117. Lowe GD, Rumley A, Mackie IJ. 2004. Plasma fibrinogen. Ann. Clin. Biochem.
41:430–40
118. Lacy F, O’Connor DT, Schmid-Schönbein GW. 1998. Plasma hydrogen per-
oxide production in hypertensives and normotensive subjects at genetic risk of
hypertension. J. Hypertens. 16:291–303
119. McDonald DM. 1994. Endothelial gaps and permeability of venules in rat
tracheas exposed to inflammatory stimuli. Am. J. Physiol. Endocrinol. Metab.
266:L61–83
120. Gute DC, Ishida T, Yarimizu K, Korthuis RJ. 1998. Inflammatory responses to
ischemia and reperfusion in skeletal muscle. Mol. Cell. Biochem. 179:169–87
121. Michel CC, Curry FE. 1999. Microvascular permeability. Physiol. Rev. 79:703–
61
122. Kubes P, Grisham MB, Barrowman JA, Gaginella T, Granger DN. 1991.
Leukocyte-induced vascular protein leakage in cat mesentery. Am. J. Physiol.
Heart Circ. Physiol. 261:H1872–79
123. Zilberberg J, Harris NR. 2001. Role of shear and leukocyte adherence on venular
permeability in the rat mesentery. Microvasc. Res. 62:215–25
124. Sarelius IH, Kuebel JM, Wang J, Huxley VH. 2005. Macromolecule perme-
ability of in situ and excised rodent skeletal muscle arterioles and venules. Am.
125. McDonald DM, Thurston G, Baluk P. 1999. Endothelial gaps as sites for plasma
leakage in inflammation. Microcirculation 6:7–22
126. Baluk P, Hirata A, Thurston G, Fujiwara T, Neal CR, et al. 1997. Endothelial
gaps: time course of formation and closure in inflamed venules of rats. Am. J.
Physiol. Endocrinol. Metab. 272:L155–70
127. Neal CR, Michel CC. 1997. Transcellular openings through frog microvascular
endothelium. Exp. Physiol. 82:419–22
128. Neal CR, Michel CC. 2000. Effects of temperature on the wall strength and
compliance of frog mesenteric microvessels. J. Physiol. 526(Pt. 3):613–22
129. Schmid-Schönbein GW, Kosawada T, Skalak R, Chien S. 1995. Membrane
model of endothelial cells and leukocytes. A proposal for the origin of a cortical
stress. J. Biomech. Eng. 117:171–78
130. Waugh RE, Sassi M. 1986. An in vitro model of erythroid egress in bone marrow.
Blood 68:250–57
131. Huber AR, Weiss SJ. 1989. Disruption of the subendothelial basement mem-
brane during neutrophil diapedesis in an in vitro construct of a blood vessel
wall. J. Clin. Invest. 83:1122–36
132. Steadman R, St John PL, Evans RA, Thomas GJ, Davies M, et al. 1997. Human
neutrophils do not degrade major basement membrane components during
chemotactic migration. Int. J. Biochem. Cell Biol. 29:993–1004
133. Bakowski B, Tschesche H. 1992. Migration of polymorphonuclear leukocytes
through human amnion membrane—a scanning electron microscopic study.
Biol. Chem. Hoppe Seyler 373:529–46
ANRV281-BE08-04 ARI 12 June 2006 17:8
134. Schmid-Schönbein GW, Usami S, Skalak R, Chien S. 1980. The interaction of

leukocytes and erthrocytes in capillary and postcapillary. Microvasc. Res. 19:45–
70
135. Bagge U, Blixt A, Strid K-G. 1983. The initiation of post-capillary margination
of leukocytes: studies in vitro on the influence of erythrocyte concentration and
flow velocity. Int. J. Microcirc. Clin. Exp. 2:215–27
136. Bagge U, Karlsson R. 1980. Maintenance of white cell margination at the pas-
sage through small venular junctions. Microvasc. Res. 20:92–95
137. Damiano ER, Westheider J, Tozeren A, Ley K. 1996. Variation in the velocity,
deformation, and adhesion energy density of leukocytes rolling within venules.
Circ. Res. 79:1122–30
138. Lipowsky HH, Scott DA, Cartmell JS. 1996. Leukocyte rolling velocity and
its relation to leukocyte-endothelium adhesion and cell deformability. Am. J.

Physiol. Heart Circ. Physiol. 270:H1371–80

139. Firrell JC, Lipowsky HH. 1989. Leukocyte margination and deformation in
mesenteric venules of rat. Am. J. Physiol. Heart Circ. Physiol. 256:H1667–74
140. Greenberg AW, Brunk DK, Hammer DA. 2000. Cell-free rolling mediated
by L-selectin and sialyl Lewis(X) reveals the shear threshold effect. Biophys. J.
79:2391–402
141. Gaehtgens P, Ley K, Pries AR, Müller R. 1985. Mutual interactions between
leukocytes and microvascular blood flow. Prog. Appl. Microcirc. 7:15–28
142. Goldsmith HL, Spain S. 1984. Margination of leukocytes in blood flow through
small tubes. Microvasc. Res. 27:204–22
143. Pearson MJ, Lipowsky HH. 2004. Effect of fibrinogen on leukocyte margination
and adhesion in postcapillary venules. Microcirculation 11:295–306
144. Munn LL, Melder RJ, Jain RK. 1996. Role of erythrocytes in leukocyte-
endothelial interactions: mathematical model and experimental validation. Bio-
phys. J. 71:466–78
145. Lawrence MB, Springer TA. 1993. Neutrophils roll on E-selectin. J. Immunol.
151:6338–46
146. Burns AR, Bowden RA, Abe Y, Walker DC, Simon SI, et al. 1999. P-selectin
mediates neutrophil adhesion to endothelial cell borders. J. Leukoc. Biol. 65:299–
306
147. Norman KE, Moore KL, McEver RP, Ley K. 1995. Leukocyte rolling in vivo
is mediated by P-selectin glycoprotein ligand-1. Blood 86:4417–21
148. Maslin CL, Kedzierska K, Webster NL, Muller WA, Crowe SM. 2005.
Transendothelial migration of monocytes: the underlying molecular mecha-
nisms and consequences of HIV-1 infection. Curr. HIV Res. 3:303–17
149. Dore M, Korthuis RJ, Granger DN, Entman ML, Smith CW. 1993. P-selectin
mediates spontaneous leukocyte rolling in vivo. Blood 82:1308–16
150. Mayadas TN, Johnson RC, Rayburn H, Hynes RO, Wagner DD. 1993. Leuko-
cyte rolling and extravasation are severely compromised in P selectin-deficient
mice. Cell 74:541–54
151. Kunkel EJ, Dunne JL, Ley K. 2000. Leukocyte arrest during cytokine-
dependent inflammation in vivo. J. Immunol. 164:3301–8

ANRV281-BE08-04 ARI 12 June 2006 17:8
152. Ley K. 2002. Integration of inflammatory signals by rolling neutrophils. Im-

munol. Rev. 186:8–18
153. Tozeren A, Ley K. 1992. How do selectins mediate leukocyte rolling in venules?
Biophys. J. 63:700–9
154. Schmid-Schönbein GW. 1988. A theory of blood flow in skeletal muscle.
J. Biomech. Eng. 110:20–26
155. Lei X, Lawrence MB, Dong C. 1999. Influence of cell deformation on leukocyte
rolling adhesion in shear flow. J. Biomech. Eng. 121:636–43
156. Dong C, Lei XX. 2000. Biomechanics of cell rolling: shear flow, cell-surface
adhesion, and cell deformability. J. Biomech. 33:35–43
157. Dembo M, Torney DC, Saxman K, Hammer D. 1988. The reaction-limited
kinetics of membrane-to-surface adhesion and detachment. Proc. R. Soc. London
B. Biol. Sci. 234:55–83

158. Zhao Y, Chien S, Skalak R. 1995. A stochastic model of leukocyte rolling.
Biophys. J. 69:1309–20
159. Hammer DA, Apte SM. 1992. Simulation of cell rolling and adhesion on sur-
faces in shear flow: general results and analysis of selectin-mediated neutrophil
adhesion. Biophys. J. 63:35–57
160. Hammer DA, Tempelman LA, Apte SM. 1993. Statistics of cell adhesion under
hydrodynamic flow: simulation and experiment. Blood Cells 19:261–75; discus-
sion 275–67
161. Goetz DJ, el-Sabban ME, Pauli BU, Hammer DA. 1994. Dynamics of neu-
trophil rolling over stimulated endothelium in vitro. Biophys. J. 66:2202–9
162. Brunk DK, Hammer DA. 1997. Quantifying rolling adhesion with a cell-free
assay: E-selectin and its carbohydrate ligands. Biophys. J. 72:2820–33
163. Caputo KE, Hammer DA. 2005. Effect of microvillus deformability on leuko-
cyte adhesion explored using adhesive dynamics simulations. Biophys. J. 89:187–
200
164. King MR, Kim MB, Sarelius IH, Hammer DA. 2003. Hydrodynamic interac-
tions between rolling leukocytes in vivo. Microcirculation 10:401–9
165. Lawrence MB, Springer TA. 1991. Leukocytes roll on a selectin at physiologic
flow rates: distinction from and prerequisite for adhesion through integrins. Cell
65:859–73
166. Von Adrian UH, Chambers JD, McEvoy LM, Bargatze RF, Arfors K-E, Butcher
EC. 1991. Two-step model of leukocyte-endothelial cell interaction in inflam-
mation: distinct roles for LECAM-1 and leukocyte β2 integrins in vivo. Proc.
Natl. Acad. Sci. USA 88:7538
167. Jones DA, McIntire LV, Smith CW, Picker LJ. 1994. A two-step adhesion
cascade for T cell/endothelial cell interactions under flow conditions. J. Clin.
Invest. 94:2443–50
168. Norman MU, Kubes P. 2005. Therapeutic intervention in inflammatory dis-
eases: a time and place for anti-adhesion therapy. Microcirculation 12:91–98
169. Moazzam F, DeLano FA, Zweifach BW, Schmid-Schönbein GW. 1997. The
leukocyte response to fluid stress. Proc. Natl. Acad. Sci. USA 94:5338–43
ANRV281-BE08-04 ARI 12 June 2006 17:8
170. Fukuda S, Yasu T, Predescu DN, Schmid-Schönbein GW. 2000. Mechanisms

for regulation of fluid shear stress response in circulating leukocytes. Circ. Res.
86:E13–18
171. Cinamon G, Shinder V, Alon R. 2001. Shear forces promote lymphocyte mi-
gration across vascular endothelium bearing apical chemokines. Nat. Immunol.
2:515–22
172. Coughlin MF, Schmid-Schönbein GW. 2004. Pseudopod projection and cell
spreading of passive leukocytes in response to fluid shear stress. Biophys. J.
87:2035–42
173. Lien DC, Henson PM, Capen RL, Henson JE, Hanson WL, et al. 1991. Neu-
trophil kinetics in the pulmonary microcirculation during acute inflammation.
Lab. Invest. 65:145–59
174. Engelson ET, Schmid-Schönbein GW, Zweifach BW. 1986. The microvascu-
lature in skeletal muscle. II. Arteriolar network anatomy in normotensive and

hypertensive rats. Microvasc. Res. 31:356–74
175. Schmid-Schönbein GW, Lee J. 1995. Leukocytes in capillary flow. Int. J. Mi-
crocirc. Clin. Exp. 15:255–64
176. Skalak R, Öskaya N. 1987. Models of erythrocyte and leukocyte flow in capil-
laries. In Physiological Fluid Dynamics II, ed. LS Srinath, M Singh. New Delhi:
Tata McGraw Hill
177. Worthen GS, Schwab B 3rd, Elson EL, Downey GP. 1989. Mechanics of stimu-
lated neutrophils: cell stiffening induces retention in capillaries. Science 245:183–
86
178. Harris AG, Skalak TC. 1993. Leukocyte cytoskeletal structure is a determinant
of capillary plugging and flow resistance in skeletal muscle. Am. J. Physiol. Heart
179. Harris AG, Skalak TC, Hatchell DL. 1994. Leukocyte-capillary plugging and
network resistance are increased in skeletal muscle of rats with streptozotocin-
induced hyperglycemia. Int. J. Microcirc. Clin. Exp. 14:159–66
180. Sutton DW, Schmid-Schönbein GW. 1992. Elevation of organ resistance due
to leukocyte perfusion. Am. J. Physiol. Heart Circ. Physiol. 262:H1646–50
181. Harris AG, Skalak TC. 1996. Effects of leukocyte capillary plugging in skele-
tal muscle ischemia-reperfusion injury. Am. J. Physiol. Heart Circ. Physiol.
271:H2653–60
182. Korthuis RJ, Grisham MB, Granger DN. 1988. Leukocyte depletion attenuates
vascular injury in postischemic skeletal muscle. Am. J. Physiol. Heart Circ. Physiol.
254:H823–27
183. Dutka AJ, Kochanek PM, Hallenbeck JM. 1989. Influence of granulocytopenia
on canine cerebral ischemia induced by air embolism. Stroke 20:390–95
184. Engler RL, Dahlgren MD, Morris DD, Peterson MA, Schmid-Schönbein GW.
1986. Role of leukocytes in the response to acute myocardial ischemia and reflow
in dogs. Am. J. Physiol. Heart Circ. Physiol. 251:H318–23
185. Ohashi KL, Tung DK, Wilson J, Zweifach BW, Schmid-Schönbein GW. 1996.
Transvascular and interstitial migration of neutrophils in rat mesentery. Micro-
circulation 3:199–210

ANRV281-BE08-04 ARI 12 June 2006 17:8
186. Furie MB, Naprstek BL, Silverstein SC. 1987. Migration of neutrophils across
monolayers of cultured microvascular endothelial cells. An in vitro model of
leucocyte extravasation. J. Cell. Sci. 88(Pt. 2):161–75
187. Marchesi VT, Florey HW. 1961. Electron micrographic observations on the
emigration of leukocytes. Q. J. Exp. Physiol. 45:343–61
188. Florey HW, Grant LH. 1961. Leucocyte migration from small blood vessels
stimulated with ultraviolet light: an electron-microscope study. J. Pathol. Bac-
teriol. 82:13–17
189. Muller WA, Weigl SA, Deng X, Phillips DM. 1993. PECAM-1 is required for
transendothelial migration of leukocytes. J. Exp. Med. 178:449–60
190. Vaporciyan AA, DeLisser HM, Yan HC, Mendiguren II, Thom SR, et al. 1993.
Involvement of platelet-endothelial cell adhesion molecule-1 in neutrophil re-
cruitment in vivo. Science 262:1580–82

191. Burns AR, Smith CW, Walker DC. 2003. Unique structural features that in-
fluence neutrophil emigration into the lung. Physiol. Rev. 83:309–36
192. Xia M, Leppert D, Hauser SL, Sreedharan SP, Nelson PJ, et al. 1996. Stimulus
specificity of matrix metalloproteinase dependence of human T cell migration
through a model basement membrane. J. Immunol. 156:160–67
193. Leppert D, Hauser SL, Kishiyama JL, An S, Zeng L, Goetzl EJ. 1995. Stimula-
tion of matrix metalloproteinase-dependent migration of T cells by eicosanoids.
FASEB J. 9:1473–81
194. Weiss SJ, Peppin G, Ortiz X, Ragsdale C, Test ST. 1985. Oxidative autoactiva-
tion of latent collagenase by human neutrophils. Science 227:747–49
195. Young RE, Thompson RD, Larbi KY, La M, Roberts CE, et al. 2004. Neu-
trophil elastase (NE)-deficient mice demonstrate a nonredundant role for NE
in neutrophil migration, generation of proinflammatory mediators, and phago-
cytosis in response to zymosan particles in vivo. J. Immunol. 172:4493–502
196. Hoshi O, Ushiki T. 2004. Neutrophil extravasation in rat mesenteric venules
induced by the chemotactic peptide n-formyl-methionyl-luecylphenylalanine
(FMLP), with special attention to a barrier function of the vascular basal lamina
for neutrophil migration. Arch. Histol. Cytol. 67:107–14
197. Carman CV, Springer TA. 2004. A transmigratory cup in leukocyte diapedesis
both through individual vascular endothelial cells and between them. J. Cell
Biol. 167:377–88
198. Garcia JG, Verin AD, Herenyiova M, English D. 1998. Adherent neutrophils
activate endothelial myosin light chain kinase: role in transendothelial migra-
tion. J. Appl. Physiol. 84:1817–21
199. Gawlowski DM, Benoit JN, Granger HJ. 1993. Microvascular pressure and
albumin extravasation after leukocyte activation in hamster cheek pouch. Am.
200. Guo M, Wu MH, Granger HJ, Yuan SY. 2005. Focal adhesion kinase in
neutrophil-induced microvascular hyperpermeability. Microcirculation 12:223–
32
201. Allport JR, Muller WA, Luscinskas FW. 2000. Monocytes induce reversible
focal changes in vascular endothelial cadherin complex during transendothelial
migration under flow. J. Cell Biol. 148:203–16
ANRV281-BE08-04 ARI 12 June 2006 17:8
202. Tung DK, Bjursten LM, Zweifach BW, Schmid-Schönbein GW. 1997. Leuko-
cyte contribution to parenchymal cell death in an experimental model of in-
flammation. J. Leukoc. Biol. 62:163–75
203. Ohashi KL, Tung DK-L, Wilson JM, Schmid-Schönbein GW. 1993. Kinetics
of neutrophil migration across rat mesenteric venules using time-lapse video
photography. FASEB J. 7:M127
204. Werr J, Xie X, Hedqvist P, Ruoslahti E, Lindbom L. 1998. Beta1 integrins
are critically involved in neutrophil locomotion in extravascular tissue in vivo.
J. Exp. Med. 187:2091–96
205. Loike JD, Cao L, Budhu S, Hoffman S, Silverstein SC. 2001. Blockade of alpha 5
beta 1 integrins reverses the inhibitory effect of tenascin on chemotaxis of human
monocytes and polymorphonuclear leukocytes through three-dimensional gels
of extracellular matrix proteins. J. Immunol. 166:7534–42

206. Loike JD, Cao L, Budhu S, Marcantonio EE, El Khoury J, et al. 1999. Dif-
ferential regulation of beta1 integrins by chemoattractants regulates neutrophil
migration through fibrin. J. Cell Biol. 144:1047–56
207. Walker DC, Behzad AR, Chu F. 1995. Neutrophil migration through preex-
isting holes in the basal laminae of alveolar capillaries and epithelium during
streptococcal pneumonia. Microvasc. Res. 50:397–416
208. Doerschuk CM, Markos J, Coxson HO, English D, Hogg JC. 1994. Quanti-
tation of neutrophil migration in acute bacterial pneumonia in rabbits. J. Appl.
Physiol. 77:2593–99
209. Quehenberger O. 2005. Thematic review series: the immune system and athero-
genesis. Molecular mechanisms regulating monocyte recruitment in atheroscle-
rosis. J. Lipid Res. 46:1582–90
210. Navab M, Hama SY, Nguyen TB, Fogelman AM. 1994. Monocyte adhesion
and transmigration in atherosclerosis. Coron. Artery Dis. 5:198–204
211. Schröder S, Palinski W, Schmid-Schönbein GW. 1991. Activated monocytes
and granulocytes, capillary non-perfusion and neovascularization in diabetic
retinopathy. Am. J. Pathol. 139:81–100
212. de Villiers WJ, Smart EJ. 1999. Macrophage scavenger receptors and foam cell
formation. J. Leukoc. Biol. 66:740–46
213. Bobryshev YV. 2006. Monocyte recruitment and foam cell formation in
atherosclerosis. Micron 37:208–22
214. Zlotnik A, Yoshie O. 2000. Chemokines: a new classification system and their
role in immunity. Immunity 12:121–27
215. Boring L, Gosling J, Cleary M, Charo IF. 1998. Decreased lesion formation in
ccr2-/- mice reveals a role for chemokines in the initiation of atherosclerosis.
Nature 394:894–97
216. Ley K, Kansas GS. 2004. Selectins in T-cell recruitment to non-lymphoid tissues
and sites of inflammation. Nat. Rev. Immunol. 4:325–35
217. Miyamoto YJ, Andruss BF, Mitchell JS, Billard MJ, McIntyre BW. 2003. Di-
verse roles of integrins in human T lymphocyte biology. Immunol. Res. 27:71–84
218. Dailey MO. 1998. Expression of T lymphocyte adhesion molecules: regulation
during antigen-induced T cell activation and differentiation. Crit. Rev. Immunol.
18:153–84

ANRV281-BE08-04 ARI 12 June 2006 17:8
219. Burger D, Dayer JM. 2002. The role of human T-lymphocyte-monocyte contact
in inflammation and tissue destruction. Arthritis Res. 4(Suppl. 3):S169–76
220. D’Ambrosio D, Mariani M, Panina-Bordignon P, Sinigaglia F. 2001.
Chemokines and their receptors guiding T lymphocyte recruitment in lung
inflammation. Am. J. Respir. Crit. Care Med. 164:1266–75
221. Schmid-Schönbein GW, DeLano FA, Costa J, Harris AG. 1999. Parenchymal
cell death and leukocyte-endothelial cell interaction in acute experimental in-
flammation. In Inflammatory Cells and Mediators in CNS Diseases, ed. RR Ruffolo
Jr, GZ Feuerstein, AJ Hunter, G Poste, BW Metcalf. Australia: Harwood Acad.
Publ.
222. Suematsu M, DeLano FA, Poole D, Engler RL, Miyasaka M, et al. 1994. Spatial
and temporal correlation between leukocyte behavior and cell injury in postis-
chemic rat skeletal muscle microcirculation. Lab. Invest. 70:684–95

223. Hermann C, Zeiher AM, Dimmeler S. 1997. Shear stress inhibits H2 O2 -induced
apoptosis of human endothelial cells by modulation of the glutathione redox
cycle and nitric oxide synthase. Arterioscler. Thromb. Vasc. Biol. 17:3588–92
224. Dimmeler S, Haendeler J, Rippmann V, Nehls M, Zeiher AM. 1996. Shear
stress inhibits apoptosis of human endothelial cells. FEBS Lett. 399:71–74
225. Coxon A, Tang T, Mayadas TN. 1999. Cytokine-activated endothelial cells de-
lay neutrophil apoptosis in vitro and in vivo. A role for granulocyte/macrophage
colony-stimulating factor. J. Exp. Med. 190:923–34
226. Mayadas TN, Cullere X. 2005. Neutrophil beta2 integrins: moderators of life
or death decisions. Trends Immunol. 26:388–95
227. Hall SE, Savill JS, Henson PM, Haslett C. 1994. Apoptotic neutrophils are
phagocytosed by fibroblasts with participation of the fibroblast vitronectin
receptor and involvement of a mannose/fucose-specific lectin. J. Immunol.
153:3218–27
228. Savill JS, Henson PM, Haslett C. 1989. Phagocytosis of aged human neutrophils
by macrophages is mediated by a novel “charge-sensitive” recognition mecha-
nism. J. Clin. Invest. 84:1518–27
229. Ezaki T, Baluk P, Thurston G, La Barbara A, Woo C, McDonald DM. 2001.
Time course of endothelial cell proliferation and microvascular remodeling in
chronic inflammation. Am. J. Pathol. 158:2043–55
230. Chen IIH, Prewitt RL, Dowell RF. 1981. Microvascular rarefaction in spon-
taneously hypertensive rat cremaster muscle. Am. J. Physiol.Heart Circ. Physiol.
241:H306–10
231. Vogt CJ, Schmid-Schönbein GW. 2001. Microvascular endothelial cell death
and rarefaction in the glucocorticoid-induced hypertensive rat. Microcirculation
8:129–39
232. Zawicki DF, Jain RK, Schmid-Schönbein GW, Chien S. 1981. Dynamics of
neovascularization in normal tissue. Microvasc. Res. 21:27–47
233. Schmid-Schönbein GW, Kistler EB, Hugli TE. 2001. Mechanisms for cell
activation and its consequences for biorheology and microcirculation: Multi-
organ failure in shock. Biorheology 38:185–201
234. McCord JM. 1987. Oxygen-derived radicals: a link between reperfusion injury
and inflammation. Fed. Proc. 46:2402–6
ANRV281-BE08-04 ARI 12 June 2006 17:8
235. Yamakawa Y, Takano M, Patel M, Tien N, Takada T, Bulkley GB. 2000. In-
teraction of platelet activating factor, reactive oxygen species generated by
xanthine oxidase, and leukocytes in the generation of hepatic injury after
shock/resuscitation. Ann. Surg. 231:387–98
236. Lefer AM, Lefer DJ. 1993. Pharmacology of the endothelium in ischemia-
reperfusion and circulatory shock. Ann. Rev. Pharm. Tox. 33:71–90
237. Movat HZ, Cybulsky MI, Colditz IG, Chan MK, Dinarello CA. 1987. Acute in-
flammation in gram-negative infection: endotoxin, interleukin 1, tumor necrosis
factor, and neutrophils. Fed. Proc. 46:97–104
238. Smedegård G, Cui LX, Hugli TE. 1989. Endotoxin-induced shock in the rat.
A role for C5A. Am. J. Pathol. 135:489–97
239. Loscalzo J. 1996. The macrophage and fibrinolysis. Semin. Thromb. Hemost.
22:503–6
240. Stecher VJ, Sorkin E. 1972. The chemotactic activity of fibrin lysis products.
Int. Arch. Allergy Appl. Immunol. 43:879–86
241. Garcia JG, Pavalko FM, Patterson CE. 1995. Vascular endothelial cell activation
and permeability responses to thrombin. Blood Coagul. Fibrinolysis 6:609–26
242. Cirino G, Cicala C, Bucci MR, Sorrentino L, Maraganore JM, Stone SR. 1996.
Thrombin functions as an inflammatory mediator through activation of its re-
ceptor. J. Exp. Med. 183:821–27
243. Dahlen SE, Bjork J, Hedqvist P, Arfors KE, Hammarstrom S, et al. 1981.
Leukotrienes promote plasma leakage and leukocyte adhesion in postcapillary
venules: in vivo effects with relevance to the acute inflammatory response. Proc.
244. Liao L, Aw TY, Kvietys PR, Granger DN. 1995. Oxidized LDL-induced
microvascular dysfunction. Dependence on oxidation procedure. Arterioscler.
Thromb. Vasc. Biol. 15:2305–11
245. Lehr HA, Seemuller J, Hubner C, Menger MD, Messmer K. 1993. Oxidized
LDL-induced leukocyte/endothelium interaction in vivo involves the receptor
for platelet-activating factor. Arterioscler. Thromb. 13:1013–18
246. Kubes P, Suzuki M, Granger DN. 1991. Nitric oxide: an endogenous modula-
tion of leukocyte adhesion. Proc. Natl. Acad. Sci. USA 88:4651–55
247. Kaminski PM, Proctor KG. 1989. Attenuation of no-reflow phenomenon, neu-
trophil activation, and reperfusion injury in intestinal microcirculation by top-
ical adenosine. Circ. Res. 65:426–35
248. Zanotti S, Kumar A. 2002. Cytokine modulation in sepsis and septic shock.
Expert Opin. Investig. Drugs 11:1061–75
249. Zimmerman GA, McIntyre TM, Prescott SM. 1996. Adhesion and signaling in
vascular cell–cell interactions. J. Clin. Invest. 98:1699–702
250. Gonzalez NC, Wood JG. 2001. Leukocyte-endothelial interactions in environ-
mental hypoxia. Adv. Exp. Med. Biol. 502:39–60
251. Gilcrease MZ, Hoover RL. 1991. Neutrophil adhesion to endothelium follow-
ing hyperosmolar insult. Diabetes Res. 16:149–57
252. Blake DR, Winyard PG, Marok R. 1994. The contribution of hypoxia-
reperfusion injury to inflammatory synovitis: the influence of reactive oxygen

ANRV281-BE08-04 ARI 12 June 2006 17:8
intermediates on the transcriptional control of inflammation. Ann. N.Y. Acad.

Sci. 723:308–17
253. Altay T, Gonzales ER, Park TS, Gidday JM. 2004. Cerebrovascular inflamma-
tion after brief episodic hypoxia: modulation by neuronal and endothelial nitric
oxide synthase. J. Appl. Physiol. 96:1223–30; discussion 1196
254. Shah S, Allen J, Wood JG, Gonzalez NC. 2003. Dissociation between skeletal
muscle microvascular pO2 and hypoxia-induced microvascular inflammation.
J. Appl. Physiol. 94:2323–29
255. Whalen MJ, Carlos TM, Clark RS, Marion DW, DeKosky ST, et al. 1997. The
effect of brain temperature on acute inflammation after traumatic brain injury
in rats. J. Neurotrauma 14:561–72
256. Kessler F, Schmidt KL. 1984. Thermometry in experimental inflammation. II.
Studies of the effect of local cold application on the skin temperature of the rat’s
paw in experimental inflammation. Z. Rheumatol. 43:83–88

257. Lefer AM. 1974. Myocardial depressant factor and circulatory shock. Klin.
Wochenschr. 52:358–70
258. McMillen MA, Huribal M, Sumpio B. 1993. Common pathway of endothelial-
leukocyte interaction in shock, ischemia, and reperfusion. Am. J. Surg. 166:557–
62
259. Tompkins SD, Gregory S, Hoyt DB, Ozkan AN. 1990. In vitro inhibition of
IL-2 biosynthesis in activated human peripheral blood mononuclear cells by a
trauma-induced glycopeptide. Immunol. Lett. 23:205–9
260. Lefer AM. 1970. Role of a myocardial depressant factor in the pathogenesis of
circulatory shock. Fed. Proc. 29:1836–47
261. Emerit I, Garban F, Vassy J, Levy A, Filipe P, Freitas J. 1996. Superoxide-
mediated clastogenesis and anticlastogenic effects of exogenous superoxide dis-
mutase. Proc. Natl. Acad. Sci. USA 93:12799–804
262. Junger WG, Hoyt DB, Liu FC, Loomis WH, Coimbra R. 1996. Immunosup-
pression after endotoxin shock: the result of multiple anti-inflammatory factors.
J. Trauma 40:702–9
263. Migeotte I, Riboldi E, Franssen JD, Gregoire F, Loison C, et al. 2005. Identi-
fication and characterization of an endogenous chemotactic ligand specific for
FPRL2. J. Exp. Med. 201:83–93
264. Kistler EB, Hugli TE, Schmid-Schönbein GW. 2000. The pancreas as a source
of cardiovascular cell activating factors. Microcirculation 7:183–92
265. Barroso-Aranda J, Schmid-Schönbein GW, Zweifach BW, Mathison JC. 1991.
Contribution of polymorphonuclear neutrophils to lethality induced by bacte-
rial lipopolysaccaride. Circ. Res. 69:1196–206
266. Barroso-Aranda J, Zweifach BW, Mathison JC, Schmid-Schönbein GW. 1995.
Neutrophil activation, tumor necrosis factor, and survival after endotoxic and
hemorrhagic shock. J. Cardiovasc. Pharmacol. 25(Suppl. 2):S23–29
267. Wells CL, Hess DJ, Erlandsen SL. 2004. Impact of the indigenous flora in
animal models of shock and sepsis. Shock 22:562–68
268. Deitch EA, Berg R. 1987. Bacterial translocation from the gut: a mechanism of
infection. J. Burn Care Rehabil. 8:475–82
ANRV281-BE08-04 ARI 12 June 2006 17:8
269. Takesue Y, Kakehashi M, Ohge H, Uemura K, Imamura Y, et al. 2005. Bacterial

translocation: not a clinically relevant phenomenon in colorectal cancer. World
J. Surg. 29:198–202
270. Roumen RM, Hendriks T, Wevers RA, Goris JA. 1993. Intestinal permeability
after severe trauma and hemorrhagic shock is increased without relation to
septic complications. Arch. Surg. 128:453–57
271. Penn AH. 2005. Digestive enzymes in the generation of cytotoxic mediators during
shock. PhD thesis, Dep. Bioeng., Univ. Calif., San Diego
272. Korhonen H, Pihlanto A. 2003. Food-derived bioactive peptides—
opportunities for designing future foods. Curr. Pharm. Des. 9:1297–308
273. Parker F, Migliore-Samour D, Floc’h F, Zerial A, Werner GH, et al. 1984. Im-
munostimulating hexapeptide from human casein: amino acid sequence, syn-
thesis and biological properties. Eur. J. Biochem. 145:677–82

274. Meisel H, FitzGerald RJ. 2003. Biofunctional peptides from milk proteins: min-
eral binding and cytomodulatory effects. Curr. Pharm. Des. 9:1289–95
275. Otani H, Hata I. 1995. Inhibition of proliferative responses of mouse spleen
lymphocytes and rabbit Peyer’s patch cells by bovine milk caseins and their
digests. J. Dairy Res. 62:339–48
276. Martin MA, Monnnai M, Otani H. 2000. Isolation and characterization of a
cytotoxic pentapeptide, k-casecidin, from bovine k-casein digested with bovine
trypsin. Animal Sci. J. 71:197–207
277. Martin MA, Otani H. 2001. Antimicrobial and cytotoxic peptides released from
milk proteins by the action of mammalian gastrointestinal proteinases. Cur. Res.
Adv. Agric. Biol. Chem. 1:23–36
278. Riderknecht H. 1986. Activation of pancreatic zymogens. Normal activation,
premature intrapancreatic activation, protective mechanisms against inappro-
priate activation. Dig. Dis. Sci. 31:314–21
279. Doucet JJ, Hoyt DB, Coimbra R, Schmid-Schönbein GW, Junger WG, et al.
2004. Inhibition of enteral enzymes by enteroclysis with nafamostat mesilate
reduces neutrophil activation and transfusion requirements after hemorrhagic
shock. J. Trauma 56:501–10; discussion 510–11
280. Rollwagen FM, Davis TA, Li YY, Pacheco ND, Zhu XL. 2004. Orally admin-
istered IL-6 induces elevated intestinal GM-CSF gene expression and splenic
CFU-GM. Cytokine 27:107–12
281. Pappenheimer JR. 1990. Paracellular intestinal absorption of glucose, creati-
nine, and mannitol in normal animals: relation to body size. Am. J. Physiol.
Gastrointest. Liver Physiol. 259:G290–99
282. Langer JC, Sohal SS, Blennerhassett P. 1995. Mucosal permeability after sub-
clinical intestinal ischemia-reperfusion injury: an exploration of possible mech-
anisms. J. Pediatr. Surg. 30:568–72
283. Childs EW, Udobi KF, Hunter FA. 2005. Hypothermia reduces microvascular
permeability and reactive oxygen species expression after hemorrhagic shock.
J. Trauma 58:271–77
284. Garcia Soriano F, Liaudet L, Marton A, Hasko G, Batista Lorigados C, et al.
2001. Inosine improves gut permeability and vascular reactivity in endotoxic
shock. Crit. Care Med. 29:703–8

ANRV281-BE08-04 ARI 12 June 2006 17:8
285. Wattanasirichaigoon S, Menconi MJ, Delude RL, Fink MP. 1999. Effect of
mesenteric ischemia and reperfusion or hemorrhagic shock on intestinal mu-
cosal permeability and atp content in rats. Shock 12:127–33
286. Russell DH, Barreto JC, Klemm K, Miller TA. 1995. Hemorrhagic shock in-
creases gut macromolecular permeability in the rat. Shock 4:50–55
287. Rollwagen FM, Li YY, Pacheco ND, Dick EJ, Kang YH. 2000. Microvascu-
lar effects of oral interleukin-6 on ischemia/reperfusion in the murine small
intestine. Am. J. Pathol. 156:1177–82
288. Bounous G. 1986. Pancreatic proteases and oxygen-derived free radicals in acute
ischemic enteropathy. Surgery 99:92–94
289. Kistler EB, Hugli TE, Schmid-Schönbein GW. 2000. The pancreas as a source
of cardiovascular cell activating factors. Microcirculation 7:183–92
290. Schmid-Schönbein GW, Hugli TE, Kistler EB, Sofianos A, Mitsuoka H. 2001.
Pancreatic enzymes and microvascular cell activation in multiorgan failure. Mi-

crocirculation 8:5–14
291. Waldo SW, Rosario HS, Penn AH, Schmid-Schönbein GW. 2003. Pancreatic
digestive enzymes are potent generators of mediators for leukocyte activation
and mortality. Shock 20:138–43
292. Suematsu M. 1991. Intravital observations of granulocyte-mediated oxidative
stress on microvascular endothelium during endotoxemia. Jap. Coll. Angiol.
31:339–46
293. Kanwar S, Kubes P. 1994. Ischemia/reperfusion-induced granulocyte influx is
a multistep process mediated by mast cells. Microcirculation 1:175–82
294. Kanwar S, Kubes P. 1994. Mast cells contribute to ischemia-reperfusion-
induced granulocyte infiltration and intestinal dysfunction. Am. J. Physiol. Gas-
trointest. Liver Physiol. 267:G316–21
295. Aoyama T, Ino Y, Ozeki M, Oda M, Sato T, et al. 1984. Pharmacological studies
of FUT-175, nafamstat mesilate. I. Inhibition of protease activity in in vitro and
in vivo experiments. Jpn. J. Pharmacol. 35:203–27
296. Mitsuoka H, Kistler EB, Schmid-Schönbein GW. 2000. Generation of in vivo
activating factors in the ischemic intestine by pancreatic enzymes. Proc. Natl.
Acad. Sci. USA 97:1772–77
297. Mitsuoka H, Kistler EB, Schmid-Schönbein GW. 2002. Protease inhibition in
the intestinal lumen: attenuation of systemic inflammation and early indicators
of multiple organ failure in shock. Shock 17:205–9
298. Schimmmeyer SM. 2005. Intraintestinal enzyme inhibition in splanchnic arterial
occlusion shock. Master thesis, Dep. Bioeng., Univ. Calif., San Diego
299. Fitzal F, DeLano FA, Young C, Rosario HS, Schmid-Schönbein GW. 2002.
Pancreatic protease inhibition during shock attenuates cell activation and pe-
ripheral inflammation. J. Vasc. Res. 39:320–29
300. Deitch ED, Shi HP, Lu Q, Feketeova E, Xu DZ. 2003. Serine proteases are
involved in the pathogenesis of trauma-hemorrhagic shock-induced gut and
lung injury. Shock 19:542–46
301. Fitzal F, Delano FA, Young C, Rosario HS, Junger WG, Schmid-Schönbein
GW. 2003. Pancreatic enzymes sustain systemic inflammation after an initial
endotoxin challenge. Surgery 134:446–56
ANRV281-BE08-04 ARI 12 June 2006 17:8
302. Kloner RA, Jennings RB. 2001. Consequences of brief ischemia: Stunning,
preconditioning, and their clinical implications: Part 2. Circulation 104:3158–
67
303. Kloner RA, Jennings RB. 2001. Consequences of brief ischemia: stunning, pre-
conditioning, and their clinical implications: Part 1. Circulation 104:2981–89
304. Davis JM, Gute DC, Jones S, Krsmanovic A, Korthuis RJ. 1999. Ischemic pre-
conditioning prevents postischemic P-selectin expression in the rat small intes-
tine. Am. J. Physiol. Heart Circ. Physiol. 277:H2476–81
305. Dayton C, Yamaguchi T, Warren A, Korthuis RJ. 2002. Ischemic precondi-
tioning prevents postischemic arteriolar, capillary, and postcapillary venular
dysfunction: signaling pathways mediating the adaptive metamorphosis to a
protected phenotype in preconditioned endothelium. Microcirculation 9:73–89
306. Kamada K, Dayton CB, Yamaguchi T, Korthuis RJ. 2004. Antecedent ethanol
ingestion prevents postischemic microvascular dysfunction. Pathophysiology

10:131–37
307. Janoff A, Zweifach BW. 1963. Effect of endotoxin-tolerance, cortisone, and
thorotrast on release of enzymes for subcellular particles of mouse liver. Proc.
Soc. Exp. Biol. Med. 114:695–98
308. Zweifach BW. 1960. The contribution of the reticuloendothelial system to the
development of tolerance to experimental shock. Ann. N.Y. Acad. Sci. 88:203–12
309. Barroso-Aranda J, Schmid-Schönbein GW, Zweifach BW, Mathison JC. 1991.
Polymorphonuclear neutrophil contribution to induced tolerance by bacterial
lipopolysaccharide. Circ. Res. 69:1196–206
310. Eising GP, Mao L, Schmid-Schönbein GW, Engler RL, Ross J. 1996. Effects
of induced tolerance to bacterial lipopolysaccharide on myocardial infarct size
in rats. Cardiovasc. Res. 31:73–81
311. Szabo C, Thiemermann C, Wu CC, Perretti M, Vane JR. 1994. Attenuation of
the induction of nitric oxide synthase by endogenous glucocorticoids accounts
for endotoxin tolerance in vivo. Proc. Natl. Acad. Sci. USA 91:271–75
312. Gimbrone MA Jr, Topper JN, Nagel T, Anderson KR, Garcia-Cardena G.
2000. Endothelial dysfunction, hemodynamic forces, and atherogenesis. Ann.
N.Y. Acad. Sci. 902:230–39; discussion 239–40
313. Resnick N, Yahav H, Shay-Salit A, Shushy M, Schubert S, et al. 2003. Fluid shear
stress and the vascular endothelium: for better and for worse. Prog. Biophys. Mol.
Biol. 81:177–99
314. Ogunrinade O, Kameya GT, Truskey GA. 2002. Effect of fluid shear stress on
the permeability of the arterial endothelium. Ann. Biomed. Eng. 30:430–46
315. Langille BL. 2001. Morphologic responses of endothelium to shear stress: re-
organization of the adherens junction. Microcirculation 8:195–206
316. Busse R, Fleming I. 1998. Pulsatile stretch and shear stress: physical stimuli
determining the production of endothelium-derived relaxing factors. J. Vasc.
Res. 35:73–84
317. Topper JN, Cai J, Falb D, Gimbrone MA Jr. 1996. Identification of vas-
cular endothelial genes differentially responsive to fluid mechanical stimuli:
cyclooxygenase-2, manganese superoxide dismutase, and endothelial cell nitric

ANRV281-BE08-04 ARI 12 June 2006 17:8
oxide synthase are selectively up-regulated by steady laminar shear stress. Proc.
318. Lin K, Hsu PP, Chen BP, Yuan S, Usami S, et al. 2000. Molecular mechanism
of endothelial growth arrest by laminar shear stress. Proc. Natl. Acad. Sci. USA
97:9385–89
319. Chen BP, Li YS, Zhao Y, Chen KD, Li S, et al. 2001. DNA microarray analysis
of gene expression in endothelial cells in response to 24-h shear stress. Physiol.
Genomics 7:55–63
320. Chiu JJ, Chen LJ, Chang SF, Lee PL, Lee CI, et al. 2005. Shear stress inhibits
smooth muscle cell-induced inflammatory gene expression in endothelial cells:
role of NF-κβ. Arterioscler. Thromb. Vasc. Biol. 25:963–69
321. Lerner-Marmarosh N, Yoshizumi M, Che W, Surapisitchat J, Kawakatsu H,
et al. 2003. Inhibition of tumor necrosis factor-[alpha]-induced SHP-2 phos-

phatase activity by shear stress: a mechanism to reduce endothelial inflamma-

tion. Arterioscler. Thromb. Vasc. Biol. 23:1775–81
322. Sheikh S, Rainger GE, Gale Z, Rahman M, Nash GB. 2003. Exposure to fluid
shear stress modulates the ability of endothelial cells to recruit neutrophils in
response to tumor necrosis factor-alpha: a basis for local variations in vascular
sensitivity to inflammation. Blood 102:2828–34
323. Wung BS, Cheng JJ, Hsieh HJ, Shyy YJ, Wang DL. 1997. Cyclic strain-induced
monocyte chemotactic protein-1 gene expression in endothelial cells involves
reactive oxygen species activation of activator protein 1. Circ. Res. 81:1–7
324. Resnick N, Yahav H, Khachigian LM, Collins T, Anderson KR, et al. 1997. En-
dothelial gene regulation by laminar shear stress. Adv. Exp. Med. Biol. 430:155–
64
325. SenBanerjee S, Lin Z, Atkins GB, Greif DM, Rao RM, et al. 2004. Klf2 is
a novel transcriptional regulator of endothelial proinflammatory activation.
J. Exp. Med. 199:1305–15
326. Sugihara-Seki M, Schmid-Schönbein GW. 2003. The fluid shear stress distri-
bution on the membrane of leukocytes in the microcirculation. J. Biomech. Eng.
125:628–38
327. Makino A, Glogauer M, Bokoch GM, Chien S, Schmid-Schönbein GW. 2005.
Control of neutrophil pseudopods by fluid shear: role of Rho family GTPases.
Am. J. Physiol. Cell Physiol. 288:C863–71
328. Yap B, Kamm RD. 2005. Cytoskeletal remodeling and cellular activation during
deformation of neutrophils into narrow channels. J. Appl. Physiol. 99:2323–30
329. Yap B, Kamm RD. 2005. Mechanical deformation of neutrophils into narrow
channels induces pseudopod projection and changes in biomechanical proper-
ties. J. Appl. Physiol. 98:1930–39
330. Fukuda S, Schmid-Schönbein GW. 2003. Regulation of cd18 expression
on neutrophils in response to fluid shear stress. Proc. Natl. Acad. Sci. USA
100:13152–57
331. Komai Y, Schmid-Schönbein GW. 2005. De-activation of neutrophils in sus-
pension by fluid shear stress: a requirement for erythrocytes. Ann. Biomed. Eng.
33:1375–86
ANRV281-BE08-04 ARI 12 June 2006 17:8
332. Marschel P, Schmid-Schönbein GW. 2002. Control of fluid shear response in

circulating leukocytes by integrins. Ann. Biomed. Eng. 30:333–43
333. Wang Y, Chang J, Li YC, Li YS, Shyy JY, Chien S. 2004. Shear stress and VEGF
activate IKK via the FLK-1/CBL/AKT signaling pathway. Am. J. Physiol. Heart
334. Makino A, Prossnitz ER, Bünemann, Wang JM, Yao W, Schmid-Schönbein
GW. 2006. G protein-coupled receptors serve as mechanosensors for fluid shear
stress in neutrophils. Am. J. Physiol. Cell Physiol. 290:In press
335. Suematsu M, Oshio C, Miura S, Tsuchiya M. 1987. Real-time visualization of
oxyradical burst from single neutrophil by using ultrasensitive video intensifier
microscopy. Biochem. Biophys. Res. Commun. 149:1106–10
336. Fukuda S, Schmid-Schönbein GW. 2002. Centrifugation attenuates the fluid
shear response of circulating leukocytes. J. Leukoc. Biol. 72:133–39


Contents ARI 16 June 2006 15:26
Annual Review
of Biomedical
Contents Engineering
Volume 8, 2006
Fluorescence Molecular Imaging

Vasilis Ntziachristos p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 1
Multimodality In Vivo Imaging Systems: Twice the Power

or Double the Trouble?

Simon R. Cherry p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p35
Bioimpedance Tomography (Electrical Impedance Tomography)
R.H. Bayford p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p63
Analysis of Inflammation
Geert W. Schmid-Schönbein p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p93
Drug-Eluting Bioresorbable Stents for Various Applications
Meital Zilberman and Robert C. Eberhart p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 153
Glycomics Approach to Structure-Function Relationships
of Glycosaminoglycans
Ram Sasisekharan, Rahul Raman, and Vikas Prabhakar p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 181
Mathematical Modeling of Tumor-Induced Angiogenesis
M.A.J. Chaplain, S.R. McDougall, and A.R.A. Anderson p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 233
Mechanism and Dynamics of Cadherin Adhesion
Deborah Leckband and Anil Prakasam p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 259
Microvascular Perspective of Oxygen-Carrying and -Noncarrying
Blood Substitutes
Marcos Intaglietta, Pedro Cabrales, and Amy G. Tsai p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 289
Polymersomes
Dennis E. Discher and Fariyal Ahmed p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 323
Recent Approaches to Intracellular Delivery of Drugs and DNA
and Organelle Targeting
Vladimir P. Torchilin p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 343
Running Interference: Prospects and Obstacles to Using Small
Interfering RNAs as Small Molecule Drugs
Derek M. Dykxhoorn and Judy Lieberman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 377
v
Contents ARI 16 June 2006 15:26
Stress Protein Expression Kinetics

Kenneth R. Diller p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 403
Electrical Forces for Microscale Cell Manipulation
Joel Voldman p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 425
Biomechanical and Molecular Regulation of Bone Remodeling
Alexander G. Robling, Alesha B. Castillo, and Charles H. Turner p p p p p p p p p p p p p p p p p p p p p p p 455
Biomechanical Considerations in the Design of Graft:
The Homeostasis Hypothesis
Ghassan S. Kassab and José A. Navia p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 499
Machine Learning for Detection and Diagnosis of Disease
Paul Sajda p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 537
Prognosis in Critical Care

Lucila Ohno-Machado, Frederic S. Resnic, and Michael E. Matheny p p p p p p p p p p p p p p p p p p p p 567
Lab on a CD
Marc Madou, Jim Zoval, Guangyao Jia, Horacio Kido, Jitae Kim, and Nahui Kim p p p 601
INDEXES
Subject Index p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 629

Cumulative Index of Contributing Authors, Volumes 1–8 p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 643
Cumulative Index of Chapter Titles, Volumes 1–8 p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p p 646
ERRATA
An online log of corrections to Annual Review of Biomedical Engineering chapters (if any,
1977 to the present) may be found at http://bioeng.annualreviews.org/
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California San Diego, La Jolla, California 92093-0412; email: gwss@bioeng.ucsd.edu

Annu. Rev. Biomed. Eng. Key Words

Non-resolved chronic inflammation

Inflammatory mediator release

Expression of adhesion molecules

Minutes Hours Days/weeks

INFLAMMATION AND DISEASE

EXAMPLES OF INFLAMMATION IN THE

A Localized Endothelial Injury

Inflammation Generated by Arthus and Shwartzman Reaction

CELLULAR MARKERS OF INFLAMMATION

Cell Activation Observed In Vitro

Pseudopods are essential elements for many inﬂammatory activities of circulating

www.annualreviews.org • Inflammation 101

form, but few caveolae

(C). J: cell junction.

Pseudopod projections are rarely detected on endothelial cells in normal blood

Membrane Adhesion Molecules

www.annualreviews.org • Inflammation 103

ICAM-1 is a member of the immunoglobulin superfamily. Its expression is regu-

lated by transcription through mechanisms in the microcirculation that are largely

Oxygen Free Radical Formation

techniques was introduced by M. Suematsu and his colleagues at Keio University

www.annualreviews.org • Inflammation 105

radical–mediated parenchymal tissue injury—owing to the presence of scavenging

of the extracellular domain of adhesion molecules such as ICAM-1. Even though

Plasma Markers for Inflammation

www.annualreviews.org • Inflammation 107

Individual pores after such

acute stimulation are

repaired by the endothelial cells.

Leukocyte Infiltration into the Microcirculation

www.annualreviews.org • Inflammation 109

toward the endothelium (141–144). Neutrophils roll on P-selectin, E-selectin (145,

www.annualreviews.org • Inflammation 111

The site of leukocyte adhesion in the microcirculation is predominantly in post-

leukocyte to response to ﬂuid mechanical stress. It is a truly interdependent process.

Trapping of Leukocyte in Capillaries

between leukocyte and endothelium during transient plugging of leukocytes. The

www.annualreviews.org • Inflammation 113

In acute inﬂammatory conditions, neutrophils may persist on the basement mem-

transient plasma leakage depending on the particular choice of chemotactic agent

requires attachment of neutrophils to extracellular collagen ﬁbers via a mechanism

Monocyte and Lymphocyte Infiltration

www.annualreviews.org • Inflammation 117

Microvascular Cell Death

www.annualreviews.org • Inflammation 119

Number of PI(+) nuclei / area (n/mm2)

Ischemia (I group) by itself 400 Control

from Reference 222.

Capillary Restructuring Under Inflammation

www.annualreviews.org • Inflammation 121

TRIGGER MECHANISMS FOR INFLAMMATION

mechanisms to activate cells and stimulate an inﬂammatory cascade. It is convenient

THE SOURCE OF INFLAMMATORY

Inflammatory Mediators in Plasma

www.annualreviews.org • Inflammation 123

antihypertensive, opioid-like, and mineral binding) (272–275). Cytotoxic peptides

Pancreatic Enzymes and the Production

The New Autodigestion Hypothesis

Neutrophil mortality (%)

the production of inﬂammatory mediators by pancreatic digestive enzyme is not

www.annualreviews.org • Inflammation 125