Original article 233
Does analgesia and condition influence immunity after
surgery? Effects of fentanyl, ketamine and clonidine on
natural killer activity at different ages
Patrice Forget, Valerie Collet, Patricia Lavandhomme and Marc De Kock
Background and objective Cellular immunity varies in the
perioperative period. We evaluated the effects of fentanyl,
clonidine and ketamine at different time points after surgery and
in animals in different conditions (young vs. old).
Materials and methods Rats undergoing laparotomy under
sevoflurane anaesthesia were assigned to receive saline,
fentanyl (40 mg kg
1), clonidine (10 mg kg
1) or ketamine
(10 mg kg
1) 1 h before surgery. Natural killer (NK) activity was
quantified at different time points (immediately or after 18, 24,
48, 72 h and 8 days) in vitro by the lysis of YAC-1 cells. In-vivo
assessment included counting the number of lung metastases
induced by the MADB-106 cells.
Results During the first 24 h after surgery, a rapid increase
in NK activity was noted, followed by a significant depression
returning to baseline at 8 days. Analgesics show specific
effects: fentanyl depressed NK activity with or without
surgery. Clonidine depressed NK activity in nonoperated
animals and during the first 24 h after surgery. Ketamine
Introduction
Cellular immunity varies after surgery and anaesthesia.1,2
For unknown reasons,3,4 tissue injury significantly
depresses this immunity in proportion to the degree of
tissue injury.5 This perioperative immunosuppression
may have deleterious consequences on oncological out-
come in cancer patients.1
Anaesthetics and analgesics reduce perioperative stress
reaction and consequently are expected to reduce cellular
immune impairments.6 Paradoxically, analgesics used in
anaesthesia, that is, the synthetic opioids, induce potent
depression of cellular immunity, in particular the activity
of the natural killer (NK) cells.1,7 The NK cells are
cytotoxic lymphocytes which constitute a major com-
ponent of the innate immune system. These cells con-
tribute to the first-line defence against viral infection and
growth of tumour cells.4
Recent human data suggested that the type of anaesthe-
sia may influence changes in immune reactivity and
consequently the incidence of postoperative cancer
recurrence.8 However, human studies are scarce and
no definitive conclusion has been drawn from animal
experiments.1,6,914 The reason may be the complexity
From the Department of Anesthesiology, Universite Catholique de Louvain, St Luc
Hospital, Brussels, Belgium
Correspondence to Patrice Forget, Department of Anesthesiology, St Luc
Hospital, av. Hippocrate 10-1821, 1200 Brussels, Belgium
Tel: +32 2 7641821; e-mail: forgetpatrice@yahoo.fr
0265-0215 2010 Copyright European Society of Anaesthesiology
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
depressed NK activity in nonoperated animals but, after
surgery, this activity varied with the same time course
as saline. Ketamine and clonidine significantly reduced the
number of lung metastases in operated animals. Ketamine
significantly reduced the number of metastases in old
nonoperated animals. Finally, ageing has a significant
negative influence.
Conclusion Surgery, analgesics and co-existing conditions
significantly influence cellular immunity. The importance of these
changes varies with time. Fentanyl had a worse influence than
clonidine and ketamine, but seemed equally protective against
the development of metastases.
Eur J Anaesthesiol 2010;27:233240
Keywords: age factors, analgesia, immune response, natural killer cells,
surgery
Received 18 October 2008 Revised 23 March 2009
Accepted 22 April 2009
of immune changes. These are dynamic and a drug-
induced effect may not necessarily be the same at differ-
ent postoperative time points.7,11,15,16 Further, preopera-
tive co-existing conditions, such as stress, may strongly
influence the results and ageing may modify the response
to an immune challenge.1720
Therefore, using a validated model for cellular immu-
nity,1,12,2124 we evaluated the effects of several analge-
sics (fentanyl, clonidine and ketamine) at different
time points after surgery and in animals in different
conditions.
Methods
Animals
After approval from the Animal Care and Use Committee
of the Catholic University of Louvain, male Wistar rats
(weight >300 g) were acclimatized for at least 4 weeks
before experimentation with free access to food and
water. They were housed four per cage in single-sex
housing and in a reversed 12 h light/12 h dark cycle. They
were handled gently and were not unnecessarily dis-
turbed. Young rats were aged between 14 and 18 weeks
and old rats between 34 and 40 weeks. In accordance with
previous work21 and preliminary data (data not shown),
power analysis showed that at least five animals per
condition were necessary to detect a relative difference
of 50% of the studied parameters. Therefore, seven
animals per condition were studied.
DOI:10.1097/EJA.0b013e32832d540e
 234 Forget et al.
Surgical model
Laparotomy
The model previously described by Page et al.25 was used.
General anaesthesia was induced using 8% sevoflurane
(Sevorane; Abbott Laboratories, Abbott Park, Illinois,
USA) in an induction chamber with a continuous oxygen
flow of 1.5 l min
1 (sevoflurane vaporizer; Drager,
Lubeck, Germany). Three minutes later, the inspired
sevoflurane concentration was reduced to 3%. After
5 min, the animals were drawn from the chamber and
sevoflurane was administered via a plastic cone under
spontaneous respiration. A 4 cm midline incision was
performed after shaving and cleaning the abdomen with
iso-betadine (polyvidone iodine 50 mg ml
1, ethanol
96%) (Viatris, Brussels, Belgium). Over the course of
1 min, the small intestine was rubbed between two pieces
of gauze in four locations to initiate tissue damage. A
saline-soaked gauze pad was placed over this for 3 min.
After this manipulation, the abdominal cavity was rinsed
using saline. The muscle and skin were sutured in two
layers with Vicryl 30 (Novartis, Brussels, Belgium).
Sevoflurane was discontinued and the animals allowed
to recover.
First experiment: natural killer activity (ex vivo) in young
animals
Groups
Rats were randomly assigned to one of the four groups
(saline, fentanyl, clonidine and ketamine) (Fig. 1). All the
animals underwent the surgical procedure at time zero.
The operator was blinded for the group assigned during
the whole experiment. At the end of the study protocol,
the animals were euthanazied by intraperitoneal injection
of an overdose of pentobarbital.
Drug administration
Each group received the study drug 1 h before anaes-
thesia and surgery. Drugs in a volume of 1 ml were
injected intraperitoneally. Doses administered were
fentanyl 40 mg kg
1 (Janssen-Cilag, Berchem, Belgium),
clonidine chlorhydrate 10 mg kg
1 (Catapressan; Boer-
hinger Ingelheim, Ingelheim, Germany), ketamine
10 mg kg
1 (Ketalar; Pfizer, New York, USA) and saline.
Blood for sampling (1 ml heparinized syringe with
25 U ml
1) was withdrawn by intracardiac puncture under
sevoflurane anaesthesia.
Natural killer activity assessment
The NK activity was measured according to the metho-
dology described by Shakhar and Ben-Eliyahu22 immedi-
ately before and at 1, 18, 24, 48, 72 h or 8 days after
surgery.
NK cell activity was quantified by determining the ability
of peripheral blood mononuclear cells (PBMCs) to lyse an
NK-sensitive target cell line YAC-1. Target cells were
labelled by incubation with radioactive chromium,
European Journal of Anaesthesiology 2010, Vol 27 No 3
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
washed, then added to a dilution series of PBMCs and
incubated for 4 h. Supernatants were harvested and the
amount of intracellular chromium-51 released into
the supernatant was measured with a gamma counter.
The amount of chromium-51 released is proportional
to the lytic activity of the NK cells. The percentage
release was calculated by the formula: (sample release-
SP)/(MAX-SP)  100.
Second experiment: MADB-106 (in vivo) in young and
old animals
The MADB-106 cells are a selected variant of a mam-
mary adenocarcinoma (MADB-100) chemically induced
in Fischer 344 rats. The MADB-106 tumour metastasizes
only to the lungs, and the number of tumour cells
retained in the lung 24 h following intravenous inocu-
lation as well as the consequent metastases enumerated
weeks later are highly dependent on NK activity.21,22,24
Groups
Young and old rats were randomly assigned to one of
the four following groups of treatment: saline, fentanyl,
clonidine and ketamine (Fig. 1). All these animals under-
went the surgical procedure previously described at time
zero. Control rats received the study drugs but did not
undergo surgery.
Drug administration
One hour before anaesthesia and surgery (laparotomy) or
anaesthesia alone, drugs were injected intraperitoneally:
fentanyl 40 mg kg
1, clonidine chlorhydrate 10 mg kg
1,
ketamine 10 mg kg
1 or saline depending on the group
assignment.
Pulmonary metastases
For induction and counting of the pulmonary metastases,
the same method as described by Page et al.12 was used.
The cell line was maintained under 5% CO2 at 378C in
cultures in complete medium. Two hours after the drugs
were administered 4  105 MADB-106 cells kg
1 were
injected in the caudal vein in phosphate-buffered saline
0.5 ml. After 21 days, the rats were euthanazied (intra-
peritoneally pentobarbital) and the lungs removed and
placed in Bouins solution [71% saturated picric acid
solution, 23% formaldehyde (37% solution) and 5% gla-
cial acetic acid]. After ethanol washing, metastases were
counted by two observers blinded to the group of treat-
ment assigned. The results were expressed as the total
number of metastases in both lungs.
Statistical analysis
ANOVA and MANOVA were used to detect intragroup
and intergroup differences. Normality of the data was
assessed with ShapiroWilks test. Homoscedasticity
was assessed with Levenes test. If significant, Tukeys
Honestly Significant Differences test was used for
specific intergroup or intragroup comparisons. A P value

Fig. 1
Time of drug administration, natural killer activity assessment and surgery in experiments 1 and 2. IP, intraperitoneally; NK, natural killer.
less than 0.05 was considered statistically significant.
Statistica (data analysis software system) version 7 (Stat-
soft Inc., 2004, Tulsa, Oklahoma, USA) was used for all
the analyses.
Results
No animal died as a result of the surgery or intraperitoneal
drug administration before completion of the assigned
experiment.
Experiment 1: natural killer activity in vitro
Saline group
In control nonoperated animals treated with saline, the
baseline NK activity was measured at 15.6  2.9%
(Fig. 2). Surgical stress (under sevoflurane anaesthesia)
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
Natural killer activity, analgesia and ageing 235
induced a biphasic evolution of the NK activity. First, a
significant increase was noted (peak effect after 18 h,
42.5  1.5 vs. 15.6  2.9%, P  0.01). After this peak,
the NK activity significantly decreased (minimum
4.9  2.3% at 48 h, P  0.05 vs. baseline and peak values).
After 8 days, the NK activity was comparable to that of
the controls.
Fentanyl group
Fentanyl significantly depressed NK activity in non-
operated animals (4.6  3.6 vs. 15.6  2.9%, P < 0.05 vs.
saline group). At 1, 18 and 24 h after surgery, this drug
caused a highly significant depression of NK activity
when compared with saline-treated animals (peak at
European Journal of Anaesthesiology 2010, Vol 27 No 3
 236 Forget et al.
Fig. 2
50
T0
& 1h
18 h
24 h
40 & 48 h
72 h
Natural killer lyse (%)
Day 8
30
20
& *
10 & * *
* *
*
0 animals (200  17 vs. 108  10, P < 0.05). Surprisingly,
Saline Fentanyl
Natural killer activity assessed by specific killing of YAC-1 target cells in
rats without (T0) or after a laparotomy under sevoflurane anaesthesia.
Blood sampling made 1 h after administration of saline, fentanyl
(40 mg kg
1), clonidine (10 mg kg
1) or ketamine (10 mg kg
1) at
different time points (immediately or after 18, 24, 48, 72 h or 8 days).
Data are presented as means  SEM. P < 0.05 in intergroup
comparison, comparison with saline group at the same time. &P < 0.05
in intragroup comparison.
18 h: 42.5  1.5 vs. 2.5  1.3%, P < 0.001). Depression of
NK activity was still noted after 8 days.
Clonidine group
Clonidine significantly depressed NK activity in non-
operated animals (8.3  1.8 vs. 15.6  2.9%, P < 0.05).
In operated animals, clonidine depressed NK activity
during the first 24 h when compared with saline-treated
animals. After this period, it significantly increased the
NK activity. At 8 days, the results obtained were
comparable to those observed in nonoperated clonidine-
treated animals but lower than in saline controls.
Ketamine group
Ketamine significantly depressed NK activity in non-
operated animals (5.4  1.0 vs. 15.6  2.9%, P < 0.05).
After surgery, the evolution of NK activity in ketamine-
treated animals presented with a pattern similar to the one
observed in saline-treated animals. The peaks and troughs
were, however, less pronounced than in saline-treated rats.
Surprisingly, 8 days after surgery, the administration of
ketamine still increased NK activity in comparison with
nonoperated ketamine-treated animals.
Experiment 2: MADB-106 (in vivo)
In young nonoperated animals, fentanyl was the only
drug associated with a significant increase in the number
of lung metastases (n 97  17 vs. 31  11 in the saline
group, P < 0.05) (Fig. 3). No significant difference was
observed in animals treated with clonidine or ketamine
when compared with the saline controls.
European Journal of Anaesthesiology
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
Surgical trauma per se is a potent stimulus for metastatic
growth (n 178  21 in the saline-operated group vs.
31  11 in the nonoperated group, P < 0.05). In contrast
with nonoperated animals, the administration of fentanyl
did not significantly promote development of metastases
in operated animals. In clonidine-treated and ketamine-
*
& treated animals, laparotomy was not associated with an
*
& & increased number of lung metastases (Fig. 3a).
* &
&& In old nonoperated animals, the total number of pulmon-
ary metastases was significantly higher than in young
* *
* animals except in the ketamine group. In this particular
group, the number of metastases was significantly lower
than that observed in the old nonoperated saline-treated
the administration of fentanyl was not associated with
Clonidine Ketamine an increased number of metastases in old nonoperated
animals. As in younger animals, surgery significantly
increased the number of metastases in saline-treated
and fentanyl-treated animals but not in clonidine-treated
and ketamine-treated animals (Fig. 3b).
The results are summarized in Table 1.
Discussion
The purpose of the present work was to study the effects
of various analgesic agents on cellular immunity at several
time points after surgery. The influence of different
coexisting conditions (i.e. young vs. old animals) was also
investigated. In the present study, two different tech-
niques were used to assess the lytic activity of the NK
cells: an in-vitro technique using the lysis of YAC-1 cells,
validated by several publications,1,12,2125 and an in-vivo
process assessing the cytotoxic potency of NK cells by
counting lung metastase following intravenous inocu-
lation of MADB-106 cells. The development of lung
metastases from this type of mammary adenocarcinoma
is highly dependent on NK activity.21,22,24 Using this
methodology, it was possible to study the influence of
drugs and conditions on NK function. The doses were
chosen according to the literature on acute antinocicep-
tion.17,26,27
Our results confirm that both surgery and drugs interfere
with cellular immunity. Surgical stress induces a biphasic
reaction. In the first 24 h, a rapid increase in NK activity is
noted, which is followed by a significant depression
returning to baseline after 8 days. Analgesic drugs show
specific effects and interfere with this time course.
The mu-opiate agonist fentanyl significantly depresses
the lytic activity of the NK cells measured in vitro before
and after surgery. This immune suppressive effect is,
however, less pronounced when considering growth of
metastases. Ageing has a significant negative influence on
growth of metastases. But when compared with the
saline-treated animals, an increased incidence of metas-
tases is reported only in young fentanyl-treated animals
without surgery. After surgery, administration of fentanyl
2010, Vol 27 No 3
 Natural killer activity, analgesia and ageing 237

Fig. 3

(a)
400
Without surgery
350 With laparotomy

300

Metastases in the lungs (n) 250

*&
200
*&
150
*
+
100 +

50

0
Saline Fentanyl Clonidine Ketamine

(b)
+&
400 Without surgery
With laparotomy
350
Metastases in the lungs (n)

300
&#
250
*
*
200

&# &#
150

100

50

0
Saline Fentanyl Clonidine Ketamine

Effects on immunity of surgery and analgesics in young (1418 weeks old) (a) and old rats (3440 weeks old) (b) with or without laparotomy.
Natural killer (NK) activity was assessed in vivo by the number of lung metastases 3 weeks after the injection in the caudal vein of 4  105 MADB-
106 cells kg
1 (mammary adenocarcinoma). Fentanyl (40 mg kg
1), clonidine (10 mg kg
1), ketamine (10 mg kg
1) or saline was injected
intraperitoneally 5 h before cells injection. (a) P < 0.05 in comparison with saline without surgery. P < 0.05 in comparison with saline with
laparotomy. &P < 0.05 in intragroup comparison. (b) P < 0.05 in comparison with young animals without surgery in the same group. P < 0.05 in
comparison with old animals without surgery in the same group. &P < 0.05 in comparison with young animals with surgery. #P < 0.05 in intergroup
comparison with surgery vs. saline. P < 0.05 in intergroup comparison without surgery vs. saline.

Table 1 Natural killer activity assessed in vitro by specific killing of YAC-1 target cells and in vivo by the number of lung metastases 3 weeks
after the injection in the caudal vein of 4 T 105 MADB-106 cells kgS1 (mammary adenocarcinoma) without or after a laparotomy under
sevoflurane anaesthesia
NK activity in vitro Number of lung metastases

Young rats Old rats

Groups Without surgery With surgery Without surgery With surgery Without surgery With surgery

Saline Control "# & Control " & " & " &
Fentanyl (40 mg kg
1) #M #M "M " M& " & # M"&
Clonidine (10 mg kg
1) #M #" M & # M # M # M"&
Ketamine (10 mg kg
1) #M "M& # M # M # M"&

M
P < 0.05 in intergroup comparison, comparison with saline group at the same time. &P < 0.05 in intragroup comparison. For specific comparisons, see the text.

European Journal of Anaesthesiology 2010, Vol 27 No 3

Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.

 238 Forget et al.
does not increase (in young) or even reduce (in old) the
incidence of metastases.
Clonidine depresses in vitro NK activity before surgery
and during the first postoperative hours. In vivo, however,
it significantly reduces the number of metastases devel-
oped after surgery in young animals. In old animals, it
additionally reduces metastases in nonoperated animals
when compared with saline controls.
Finally, the effects of ketamine deserve attention.
It significantly depresses NK activity in nonoperated
animals but, after surgery, this activity varies according
to the same time course as saline-treated animals. Keta-
mine significantly reduces the number of lung metastases
in young operated animals and in old nonoperated and
operated animals.
The time-related evolution in NK activity we observed
has important methodological consequences. It is not
possible to draw any valid conclusion on the effects of
a particular drug from a study considering only one point
on the time-related curve. Moreover, this may explain the
wide discrepancies reported between the results obtained
for the same drug by different studies. For example,
morphines effect on postoperative NK activity has been
shown to be depressive,13,28,29 neutral30,31 or protec-
tive.12,32,33 Similar arguments may be advanced for the
discordant literature existing for fentanyl.1
The complexity increases when considering the inter-
actions of analgesics (e.g. opiates) with the adrenergic
system. Drugs acting on this system are known to influ-
ence cellular immunity. This is not really surprising,
given the numerous functional interactions between
immunity, opiates and the sympathetic nervous sys-
tem.13,5,22,3436 For example, immune competent cells
express similar receptors to sensory neurons (i.e. opiate
and adrenergic receptors).37 Consequently, it may be
expected that drugs acting on these receptors affect both
immunity and sensory functions.
Several studies have considered the effects of adrenergic
agonists/antagonists; however, there are very few data
concerning the alpha-2-adrenergic agonist clonidine. The
drug is widely used as an anxiolytic during anaesthesia, as
a component of balanced anaesthesia or to prevent myo-
cardial ischaemia. Alpha-2-adrenoceptor agonists are
potent modulators of the stress reaction. These effects
are dual. Centrally, they induce analgesia via the presyn-
aptic alpha-2-adrenoreceptor in the descending path-
ways. Peripherally, they increase local apoptosis and
anti-inflammatory products at the site of a lesion.38,39
Moreover alpha-2-adrenergic stimulation transforms
cytokine gene expression from a pro-inflammatory to
an anti-inflammatory profile.40 In summary, clonidine
may blunt the biphasic immune reaction. Consequently,
clonidine may positively influence cellular immunity
after surgical stress.
European Journal of Anaesthesiology 2010, Vol 27 No 3
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
As with the other analgesics studied, ketamine signifi-
cantly reduces NK activity in animals without laparo-
tomy. This is in accordance with another study21 and is
probably mediated by an interaction with the sympath-
etic nervous system. However, that study used very high
doses of ketamine (80 mg kg
1) and did not include a
surgical procedure. After surgery, the depressant effect is
less pronounced. In young and old animals, ketamine
displays the same effects on metastases development as
clonidine. Ketamine is known to interfere with many
determinants of the immune reaction, such as the pro-
duction of cytokines, and, therefore, a positive influence
on cellular immunity is not surprising. This has been
shown in humans with small doses (0.5 mg kg
1) before
maxillofacial surgery.41
In the present study, we were able to confirm that innate
immunity experiences significant changes with advan-
cing age.18 Particularly, as reported by Ogata et al.,42 NK
cells from old animals are less capable of destroying
tumour cells. It is interesting to note that, the effects
of drugs differ between young and old animals. The
deleterious effects of fentanyl in young nonoperated
animals are absent in old animals. In contrast with the
young nonoperated animals, both clonidine and ketamine
protect against metastasis multiplication in nonoperated
old rats. Alterations in intracellular signalling are key
contributors to the tumoricidal deficiency of NK cells
in the old animals.43 This may explain the differential
effect of drugs according to age. The general positive
antistress effects of these drugs became prominent
when compared with the direct negative effect on the
NK cells.
For a long time, the literature has highlighted the import-
ance of cellular immunity depression after tissue destruc-
tion.5 Cellular immunity is mediated by antigen-
sensitized T lymphocytes via lymphokines or direct
cytotoxicity. It consists of the activation of macrophages,
NK cells, cytotoxic T lymphocytes and the release of
various cytokines. This response is directed primarily at
microbes infecting cells and participates in defending
against cancer cell proliferation. Among these effectors,
NK cells have received particular attention in the repres-
sion of cellular immunity after surgery.1 These cells are
considered to be the major effector cells that are active
before the induction of adaptive immunity by T cells in
the early stages of tumour growth.44,45 In numerous
animal models, preservation of NK cytotoxic activity
was positively correlated with resistance to metastasis
multiplication and growth after surgery.1,12,25,32,35,46,47
These experimental results were validated in some stu-
dies considering metastasis-free survival and metastasis
progression in humans.48,49 Nevertheless, it is widely
acknowledged that animal models do not reliably
simulate immunemalignant interactions in humans.50
Models of experimental metastasis are chosen based on
their artificially high immunogenicity, and the course of

natural metastasis development in rodents is markedly
shorter and potentially less complex than in humans.36
Thereby, it precludes any extrapolation of the animal
data on humans. This kind of investigation remains,
however, an important first step to studying the complex
interactions between stress, drugs and immunity.
Which other parameters and which limits need to be
evoked? Influence of sevoflurane was not investigated in
this study. However, suppression of NK activity by halo-
genated anaesthetic agents seems to be relatively marginal
if present.21 Another concern may be the physiological
differences induced by the analgesics. However, the influ-
ence of blood gases on NK activity or haemodynamics
induced by these analgesics has already been refuted.26,51
In conclusion, our results confirm that surgery, analgesics
and co-existing conditions such as age significantly influ-
ence cellular immunity. The importance of the induced
changes varies with time and with the occurrence of
surgery. Fentanyl seems to have a greater depressive
effect on NK activity than clonidine and ketamine.
Fentanyl is the only drug susceptible to increase the
incidence of metastases at the doses we used. Because
immunity is a highly integrated system, it is necessary to
consider several parameters before drawing any definitive
conclusion based on one immune parameter or another.
References
1 Shakhar G, Ben-Eliyahu S. Potential prophylactic measures against
postoperative immunosuppression: could they reduce recurrence rates in
oncological patients? Ann Surg Oncol 2003; 10:972992.
2 Galley HF, DiMatteo MA, Webster NR. Immunomodulation by anaesthetic,
sedative and analgesic agents: Does it matter? Intensive Care Med 2000;
26:267274.
3 Viswanathan K, Daugherty C, Dhabhar FS. Stress as an endogenous
adjuvant: augmentation of the immunization phase of cell-mediated
immunity. Int Immunol 2005; 17:10591069.
4 Orange JS, Ballas ZK. Natural killer cells in human health and disease. Clin
Immunol 2006; 118:110.
5 Desborough JP. The stress response to trauma and surgery. Br J Anaesth
2000; 85:109117.
6 Page GG, Ben-Eliyahu S, Yirmiya R, Liebeskind JC. Morphine attenuates
surgery-induced enhancement of metastatic colonization in rats. Pain
1993; 54:2128.
7 Beilin B, Shavit Y, Hart J, et al. Effects of anaesthesia based on large versus
small doses of fentanyl on natural killer cell cytotoxicity in the perioperative
period. Anesth Analg 1996; 82:492497.
8 Exadaktylos AK, Buggy DJ, Moriarty DC, et al. Can anesthetic technique for
primary breast cancer surgery affect recurrence or metastasis?
Anesthesiology 2006; 105:660664.
9 Whiteside TL, Herberman RB. Role of human natural killer cells in health
and disease. Clin Diagn Lab Immunol 1994; 1:125133.
10 Page GG. Immunologic effects of opioids in the presence or absence of
pain. J Pain Symptom Manage 2005; 29 (S5):S25S31.
11 Nelson CL, Dykstra LA, Lysle DT. Comparison of the time course of
morphines analgesic and immunologic effects. Anesth Analg 1997;
85:620626.
12 Page GG, McDonald JS, Ben-Eliyahu S. Preoperative versus postoperative
administration of morphine: impact on the neuroendocrine, behavioural,
and metastatic-enhancing effects of surgery. Br J Anaesth 1998; 81:216
223.
13 Shavit Y, Lewis JW, Terman GW, et al. Opioid peptides mediate the
suppressive effect of stress on natural killer cell cytotoxicity. Science 1984;
223:188190.
14 Shapiro J, Jersky J, Katzav S, et al. Anesthetic drugs accelerate the
progression of postoperative metastases of mouse tumors. J Clin Invest
1981; 68:678685.
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
Natural killer activity, analgesia and ageing 239
15 Wrona D. Neural-immune interactions: an integrative view of the
bidirectional relationship between the brain and immune systems.
J Neuroimmunol 2006; 172:3858.
16 Greisen J, Hokland M, Grofte T, et al. Acute pain induces an instant
increase in natural killer cell cytotoxicity in humans and this response is
abolished by local anaesthesia. Br J Anaesth 1999; 83:235240.
17 De Kock M, Meert TF. Alpha 2-adrenoreceptor agonists and stress-induced
analgesia in rats: influence of stressors and methods of analysis. Pharmacol
Biochem Behav 1997; 58:109117.
18 Plackett TP, Boehmer ED, Faunce DE, Kovacs EJ. Aging and innate
immune cells. J Leukoc Biol 2004; 76:291299.
19 Albright JF, Albright JF. Age-associated impairment of murine natural killer
activity. Proc Natl Acad Sci U S A 1983; 80:63716375.
20 Mocchegiani E, Malavolta M. NK and NKT cells functions in
immunosenescence. Aging Cell 2004; 3:177184.
21 Melamed R, Bar-Yosef S, Shakhar G, et al. Suppression of natural killer cell
activity and promotion of tumor metastasis by ketamine, thiopental, and
halothane, but not by propofol: mediating mechanisms and prophylactic
measures. Anesth Analg 2003; 97:13311339.
22 Shakhar G, Ben-Eliyahu S. In vivo beta-adrenergic stimulation suppresses
natural killer activity and compromises resistance to tumor metastasis in
rats. J Immunol 1998; 160:32513258.
23 Kiessling R, Klein E, Wigzell H. Natural killer cells in the mouse. I. Cytotoxic
cells with specificity for mouse Moloney leukemia cells. Specificity and
distribution according to genotype. Eur J Immunol 1975; 5:112117.
24 Barlozzari T, Leonhardt J, Wiltrout RH, et al. Direct evidence for the role of
LGL in the inhibition of experimental tumor metastases. J Immunol 1985;
134:27832789.
25 Page GG, Blakely WP, Ben-Eliyahu S. Evidence that postoperative pain is a
mediator of the tumor-promoting effects of surgery in rats. Pain 2001;
90:191199.
26 Docquier MA, Lavandhomme P, Ledermann C, et al. Can determining the
minimum alveolar anesthetic concentration of volatile anesthetic be used as
an objective tool to assess antinociception in animals? Anesth Analg 2003;
97:10331039.
27 Alam S, Saito Y, Kosaka Y. Antinociceptive effects of epidural and
intravenous ketamine to somatic and visceral stimuli in rats. Can J Anaesth
1996; 43:408413.
28 Colacchio TA, Yeager MP, Hildebrandt LW. Perioperative
immunomodulation in cancer surgery. Am J Surg 1994; 167:171179.
29 Saurer TB, Ijames SG, Lysle DT. Neuropeptide Y Y1 receptors mediate
morphine-induced reductions of natural killer cell activity. J Neuroimmunol
2006; 177:1826.
30 Tsai YC, Won SJ, Lin MT. Effects of morphine on immune response in rats
with sciatic constriction injury. Pain 2000; 88:155160.
31 Franchi S, Panerai AE, Sacerdote P. Buprenorphine ameliorates the effect
of surgery on hypothalamuspituitaryadrenal axis, natural killer cell
activity and metastatic colonization in rats in comparison with morphine or
fentanyl treatment. Brain Behav Immun 2007; 21:767774.
32 Bar-Yosef S, Melamed R, Page GG, et al. Attenuation of the tumor-
promoting effect of surgery by spinal blockade in rats. Anesthesiology
2001; 94:10661073.
33 Page GG, Ben-Eliyahu S, Liebeskind JC. The role of LGL/NK cells in
surgery-induced promotion of metastasis and its attenuation by morphine.
Brain Behav Immun 1994; 8:241250.
34 Docquier MA, Lavandhomme P, Boulanger V, et al. Questioning the
cardiocirculatorty effects of opioids under volatile anaesthesia. Br J
Anaesth 2004; 93:408413.
35 Melamed R, Rosenne E, Shakhar K, et al. Marginating pulmonary-NK
activity and resistance to experimental tumor metastasis: suppression
by surgery and the prophylactic use of a beta-adrenergic antagonist
and a prostaglandin synthesis inhibitor. Brain Behav Immun 2005;
19:114126.
36 Ben-Eliyahu S, Shakhar G. The impact of stress, catecholamines, and the
menstrual cycle on NK activity and tumor development: from in vitro studies
to biological significance. In: Ader R, Felten DL, Cohen N, editors.
Psychoneuroimmunology. 3rd ed. San Diego: Academic Press; 2000.
pp. 25452563.
37 Elenkov IJ, Wilders RL, Chrousos GP, Vizi ES. The sympathetic nerve: an
integrative interface between two supersystems the brain and the
immune system. Pharmacol Rev 2000; 52:592638.
38 Juge M, Grimaud N, Petit JY. Involvement of alpha2-adrenergic
mechanisms in experimental analgesic and anti-inflammatory activities of a
benzamide derivative. Pharmacol Res 1997; 36:179185.
39 Romero-Sandoval A, Eisenach JC. Clonidine reduces hypersensitivy and
alters the balance pro- and anti-inflammatory leukocytes after local injection
at the site of inflammatory neuritis. Brain Behav Immun 2007; 21:569
580.
European Journal of Anaesthesiology 2010, Vol 27 No 3
 240 Forget et al.
40 Romero-Sandoval EA, McCall C, Eisenach JC. Alpha2-adrenoreceptor
stimulation transforms immune responses in neuritis and blocks neuritis-
induced pain. J Neurosci 2005; 25:89888994.
41 Bentley MW, Stas JM, Johnson JM, et al. Effects of preincisional
ketamine treatment in natural killer cell activity and postoperative
pain management after oral maxillofacial surgery. AANA J 2005; 73:427
436.
42 Ogata K, Yokose N, Tamura H, et al. Natural killer cells in the late decades
of human life. Clin Immunol Immunopathol 1997; 84:269275.
43 Mariani E, Mariani AR, Meneghetti A, et al. Age-dependent decreases of NK
cell phosphoinositide turnover during spontaneous but not Fc-mediated
cytolitic activity. Int Immunol 1998; 10:981989.
44 Kurosawa S, Matsuzaki G, Harada M, et al. Early appearance and activation
of natural killer cells in tumor-infiltrating lymphoid cells during tumor
development. Eur J Immunol 1993; 23:10291033.
45 Kim R, Emi M, Tanabe K. Cancer immunoediting from immune surveillance
to immune escape. Immunology 2007; 121:114.
European Journal of Anaesthesiology 2010, Vol 27 No 3
Copyright European Society of Anaesthesiology. Unauthorized reproduction of this article is prohibited.
46 Shakhar G, Abudarham N, Melamed R, et al. Amelioration of operation-
induced suppression of marginating pulmonary NK activity using poly IC: a
potential approach to reduce postoperative metastasis. Ann Surg Oncol
2007; 14:841852.
47 Gaspani L, Bianchi M, Limiroli E, et al. The analgesic drug tramadol
prevents the effect of surgery on natural killer cell activity and metastatic
colonization in rats. J Neuroimmunol 2002; 129:1824.
48 Liljefors M, Nilsson B, Hielm Skog AL, et al. Natural killer (NK) cell function
is a strong prognostic factor in colorectal carcinoma patients treated with
the monoclonal antibody 17-1A. Int J Cancer 2003; 105:717723.
49 Whiteside TL, Herberman RB. The role of natural killer cells in human
disease. Clin Immunol Immunopathol 1989; 53:123.
50 Ben-Eliyahu S, Page GG, Schleifer SJ. Stress, NK cells, and cancer: still a
promissory note. Brain Behav Immun 2007; 21:881887.
51 Beilin B, Shavit Y, Cohn S, Kedar E. Narcotic-induced suppression of
natural killer cell activity in ventilated and nonventilated rats. Clin Immunol
Immunopathol 1992; 64:173176.