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Hydrogen sulfide attenuates surgical trauma-induced inflammatory response and cognitive deficits in mice
j o u r n a l o f s u r g i c a l r e s e a r c h 1 8 3 ( 2 0 1 3 ) 3 3 0 e3 3 6
Available online at www.sciencedirect.com
journal homepage: www.JournalofSurgicalResearch.com
Hydrogen sulfide attenuates surgical trauma-induced inflammatory response and cognitive deficits in mice Qin-Jun Chu, MD,a,1 Long He, MD,a,1 Wei Zhang, MD,a Chun-Lan Liu, MD,a Yan-Qiu Ai, MD,a,* and Qi Zhang, PhDb,** a b
Department of Anesthesiology, First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China Department of pharmacology, School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, Henan, China
article info
abstract
Article history:
Background: It has been increasingly reported that peripheral surgical trauma triggers
Received 2 October 2012
neuroinflammatory processes associated with postoperative cognitive dysfunction, and
Received in revised form
that mitigating the neuroinflammatory effects of surgery prevents surgery-induced
2 December 2012
cognitive dysfunction. Endogenously produced hydrogen sulfide (H2S) has multiple func-
Accepted 3 December 2012
tions in the brain, and an increasing number of studies have demonstrated its anti-
Available online 21 December 2012
inflammatory effects. The present study was designed to investigate the effects of
Keywords:
experience neuroinflammatory changes induced by surgery.
Postoperative cognitive dysfunction
Methods: Each mouse received 5 mg/kg NaHS or volume-matched vehicle administration
Inflammatory response
by intraperitoneal injection once daily, 3 d before surgery, on the day of surgery, and for 3
Hydrogen sulfide
d afterward. We assessed cognitive function using a Morris water maze and evaluated
Mice
expression of proinflammatory cytokines tumor necrosis factor-a, interleukin-1b, and
sodium hydrosulfide (NaHS), an H2S donor, on the cognitive impairment of mice as they
interleukin-6 in the serum and hippocampus. We performed each test 1, 3, and 7 d after surgery. Results: Hippocampal-dependent memory impairment in mice after surgery was associated with increased serum proinflammatory cytokines, as well as proinflammatory cytokine expression in the hippocampus. Presurgery treatment with NaHS, an H2S donor, significantly attenuated surgery-induced memory impairment and expression of proinflammatory cytokines in the serum and hippocampus. Conclusions: These findings suggest that intraperitoneal injections of NaHS could significantly mitigate surgery-induced memory impairment in mice, which is strongly associated with reduced levels of serum and hippocampal proinflammatory cytokines. ª 2013 Elsevier Inc. All rights reserved.
1.
Introduction
After surgery, patients may experience cognitive decline. This complication has been termed postoperative cognitive dysfunction (POCD). It is characterized by disruption of
attention, memory, orientation, information processing, and the sleepewake cycle, which leads to an increased risk of disability and mortality [1,2]. Elderly patients seem to be at high risk of POCD [3], but other age groups are also affected [4]. At present, little is known about the pathophysiological
* Corresponding author. Department of Anesthesiology, First Affiliated Hospital, Zhengzhou University, No. 1 East-Jianshe Road, Zhengzhou 450052, Henan, China. Tel.:þ86 0371 66862024; fax: þ86 0371 66862025. ** Corresponding author. School of Pharmaceutical Sciences, Zhengzhou University, 100 Kexue Avenue, Zhengzhou 450001, Henan, China. Tel.: þ86 0371 67781908; fax: þ86 0371 67781908. E-mail addresses: [email protected] (Y.-Q. Ai), [email protected] (Q. Zhang). 1 Qin-Jun Chu and Long He contributed equally to this research. 0022-4804/$ e see front matter ª 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jss.2012.12.003
j o u r n a l o f s u r g i c a l r e s e a r c h 1 8 3 ( 2 0 1 3 ) 3 3 0 e3 3 6
mechanisms of POCD. Consistent evidence is accumulating for the role of inflammatory processes arising as the result of surgical trauma and subsequent complications [5e7]. Earlier studies have shown that activating the peripheral inflammatory response can profoundly affect the function of the central nervous system (CNS) [8]. It is well known that surgeryinduced tissue damage can activate the peripheral innate immune system, which in turn results in the release of inflammatory mediators. Animal studies show that surgery causes learning and memory impairment that may be associated with increased levels of plasma cytokine, reactive microgliosis, tumor necrosis factor-a (TNF-a), and interleukin1b (IL-1b) transcription and expression in the hippocampus [5,9e11]. Simultaneously, nonspecific attenuation of innate immunity or preemptive treatment with anti-TNF antibodies can prevent neuroinflammatory processes as well as learning and memory dysfunction [9e12]. Hydrogen sulfide (H2S), which is well known traditionally as a toxic gas, has been proven to be produced endogenously in mammalian tissues and has an important role in physiological and pathophysiological conditions [13,14]. In recent years, there has been emerging evidence that the endogenous gaseous molecule can modulate the inflammatory process [15e17]. In the CNS, H2S exhibited extensive neurological effects in a series of in vitro and in vivo models. It was reported that H2S attenuates lipopolysaccharide (LPS)-induced cognitive impairment by reducing the overproduction of proinflammatory mediators [18]. Recently, S-propargyl-cysteine, a novel hydrogen sulfide-modulated agent, attenuated LPS- or Ab-induced spatial learning and memory impairment [19,20]. Therefore, based on these findings, we hypothesized that an H2S-modulated agent may be useful to treat or prevent POCD.
2.
Materials and methods
2.1.
Animals
The Henan Laboratory Animal Center, Henan, China, provided adult male Kunming mice weighing 30e40 g. Animals were housed in a temperature and humidity-controlled room with a 12 h/12 h lightedark cycle for at least 1 wk before the day of operation, and were allowed free access to food and tap water throughout the experiment. All animal studies were performed in accordance with the Regulations of Experimental Animal Administration issued by the State Committee of Science and Technology of the People’s Republic of China and the Institutional Review Committee for the Use of Animals.
2.2.
331
NaHS dosage from pilot studies to ascertain the dose with the highest desired pharmacologic effect. We treated the surgery mice (group S; n ¼ 24) as in the C group, except that they underwent partial hepatectomy under general anesthesia. They were anesthetized with a regimen (fentanyl 0.2 mg/kg and droperidol 5 mg/kg) that we predetermined from pilot studies. Surgery plus NaHS mice (group NS; n ¼ 24) were treated as in the S group, except that they received daily intraperitoneal NaHS solution (5 mg/kg/d) Subjects in all four groups received the same volume of NaHS resolution or placebo. We carried out all surgical procedures under aseptic conditions. Briefly, we exposed the liver through an approximately 2-cm midline abdominal incision. We excised the left lateral lobes of the liver (corresponding to approximately one-third of the organ). We then infiltrated the wound with 0.25% bupivacaine and closed it by sterile suture. We took care to ensure that all mice recovered fully from anesthesia and surgery before being individually housed. We killed the mice on postoperative days 1, 3, and 7 (n ¼ 32/d) after spatial working tests.
2.3.
Cognitive testing: Morris Water Maze
The Morris water maze (MWM) is a hippocampal-dependent test of spatial learning for rodents. The MWM consists of a pool of water (120 cm in diameter and 40 cm high) maintained at a temperature of 26 C 1 C with a hidden submerged platform (1 cm below the surface) in one quadrant. We placed mice in the water in a dimly lit room with various visual clues around the pool. We used a video camera mounted to the ceiling directly above the center of the tank in conjunction with a computerized animal tracking system to monitor swim speed, escape latency, and the distance swum. We placed mice on the platform for 30 s before the start of each trial, and released them into the water facing the wall of the pool from one of four randomly assigned release points (north, south, east, and west). In all trials, mice were allowed to swim until they landed on the platform. If a mouse failed to find the platform within 60 s, it was picked up and placed on the platform for 30 s. We trained mice with the platform in a fixed location for 4 trials/d for 5 consecutive days. Data from 4 trials/ d were averaged to represent a performance block. Animals underwent surgery on day 6. In addition, on postoperative days 1, 3, and 7, were subjected mice to a probe trial; the platform was removed and all but distal visual cues remained constant. Animals were allowed to swim for 60 s. We recorded the percentage of time spent in the east quadrant previously containing the submerged platform and the number of crossings through the target positiondthat is, the specific location of the platform during the acquisition phasedand represented it as an index of memory. All trials lasted between 9 and 16 h.
Surgery and pharmacological treatments
We randomly assigned adult mice (n ¼ 96) to four groups and then trained them on the water maze cognitive test for 5 consecutive days. Control mice (group C; n ¼ 24) received 0.3 mL normal saline (NS) intraperitoneally daily for 3 d before surgery, on the day of surgery, and for the next 3 d. We treated the NaHS mice (group N; n ¼ 24) as in the C group, except that they received intraperitoneal NaHS (Sigma, St. Louis, MO) resolution at a dose of 5 mg/kg/d daily. We determined the
2.4. Enzyme-linked immunosorbent assay for serum TNF-a, IL-1b, and IL-6 To characterize TNF-a, IL-1b, and IL-6 expression, we collected blood samples from below the canthus of the eye of each mouse (from different treatment groups). We pooled together sera of mice from two separate experiments (from different treatment groups) and subsequently used quantitative direct sandwich enzyme-linked immunosorbent assay (ELISA) (R&D
332
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Systems; Minneapolis, MN) to determine the concentration of TNF-a, IL-1b, and IL-6 (in picograms per milliliter). We conducted the procedure according to the manufacturer’s instructions.
2.5. Western blotting for hippocampal TNF-a, IL-1b, and IL-6 protein We homogenized hippocampus in a lysis buffer (20 mmol/L Tris, pH 7.5; 150 mmol/L sodium chloride; 1% Triton X-100; 1% pyrophosphate; 1% b-glycophosphate; 1% Nonidet P-40; 0.1% sodium dodecyl sulfate; 1 mmol/L ethylenediaminetetra acetic acid; 100 mg/mL phenylmethanesulfonylfluoride; and 1 mg/mL aprotinin). We processed the homogenates in a centrifuge for 15 min at 12,000 g at 4 C. We determined the quantity of protein in the supernatants using a bicinchoninic acid protein assay kit (Pierce Biotechnology, Rockford, IL). Protein extracts (30 mg/sample) denatured in Laemmli sample loading buffer at 100 C for 5 min were separated by 12% sodium dodecyl sulfateepolyacrylamide gel electrophoresis and then transferred onto nitrocellulose membranes (Hybond ECL; Amersham, Piscataway, NJ). We blocked membranes with 5% skim milk in Tris-buffered salineeTween 20 for 2 h and incubated them with rabbit polyclonal anti-TNF-a (1:1000; Abcam), rabbit polyclonal anti-IL-1b (1:700; Abcam), or rabbit polyclonal anti-IL-6 (1:500; Abcam) overnight at 4 C. After thorough washing, we incubated membranes in Tris-buffered salineeTween 20 with appropriate secondary antibodies (goat anti-rabbit, conjugated with alkaline phosphatase; Jacksonimmuno Research Laboratories, Inc, PA) diluted 1:5000 for 2 h at room temperature. Visualization of the proteins was accomplished using an enhanced chemiluminescence detection kit (ECL; Amersham) and quantified by densitometric analysis (Kodak Digital Science and ID Image Analysis software; Rochester, NY). We stripped the membrane blots and reincubated them with monoclonal anti-b-actin (AC-15; Sigma, St. Louis, MO). Relative expression levels of protein were normalized by the ratio of target protein (TNF-a, IL-1b, and IL-6) to b-actin.
2.6.
Statistical analysis
All data are expressed as mean standard error of the mean. We analyzed the behavioral data by repeated measures analysis of variance, using the Bonferroni post hoc test to determine specific group differences. We tested intergroup differences of the rest of the data using one-way analysis of variance followed by the least significant difference t-test for multiple comparisons. We performed statistical analysis with SPSS 17.0 software (SPSS Inc, Chicago, IL). In all tests, P < 0.05 indicated statistical significance.
3.
Results
3.1. Effect of NaHS on partial hepatectomy-induced spatial memory impairment To elucidate whether cognitive impairment as defined by a decline in hippocampal-dependent spatial learning and memory could be induced by partial hepatectomy, and whether this impairment could be prevented by NaHS
Fig. 1 e Performance of mice during 5-d acquisition phase in the Morris water maze. (A) Latency to the platform across training days. (B) Swimming distance to the platform across training days. (C) Swimming speed during training days. Data are represented as mean ± standard error of the mean.
administration, we conducted the MWM test. Mice underwent water maze training for 5 consecutive days (acquisition trials). As expected, animals in all groups were able to rescue themselves, and the escape latency and swimming distance gradually decreased over the 4 consecutive training days. Repeated-measures analysis of variance of swim data revealed significant effects by training day for both escape latency and swimming distance (P < 0.05) (Fig. 1A and B) but not speed (P > 0.05) (Fig. 1C). These results indicated that mice showed improvement in spatial learning and memory over time.
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Fig. 2 e Effect of NaHS on hepatectomy, resulting in cognitive impairment in mice in the Morris water maze during the probe trial test. (A) Percentage of time searching each quadrant. (B) Number of crossings over the platform. (C) Swimming speed during probe trial test. Data are represented as mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point. #P < 0.05; ##P < 0.05 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
After the acquisition trials, on postsurgical days 1, 3, and 7, we subjected mice to a probe trial. Probe trials of the MWM test showed that NaHS treatment in the NaHS group did not affect the cognitive function of mice, whereas the learning memory abilities of hepato-lobectomized mice were significantly impaired compared with those in the control group.
333
Fig. 3 e Effect of NaHS on surgery-induced systemic inflammation. We measured TNF-a, IL-1b, and IL-6 levels in serum by ELISA at 1, 3, and 7 d after surgery. (A) Tumor necrosis factor-a, (B) IL-1b, and (C) IL-6. Data are represented as mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point. #P < 0.05; ##P < 0.01 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
The percentage of time spent in the quadrant with the previously located hidden platform and the number of crossings over the platform were significantly less in hepatolobectomized mice (P < 0.05) (Fig. 2A and B), indicating cognitive impairment. However, surgery-induced learning and memory deficits were significantly alleviated by treatment with NaHS compared with the surgery group (P < 0.05) (Fig. 2A and B). On postsurgical days 1, 3, and 7, there was no
334
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Fig. 4 e Effect of NaHS on surgery-induced neuroinflammation in hippocampus. We measured TNF-a, IL-1b, and IL-6 protein by Western blot at 1, 3, and 7 d after surgery. (A) Tumor necrosis factor-a, (B) IL-1b, and (C) IL-6. Data are represented as
j o u r n a l o f s u r g i c a l r e s e a r c h 1 8 3 ( 2 0 1 3 ) 3 3 0 e3 3 6
significant difference in average swimming speed among groups, which suggests that the poorer performance of surgical mice did not result from a lack of motivation or reduced motor ability (P > 0.05) (Fig. 2C). These results indicate that partial hepatectomy induced significant cognitive impairment and that NaHS treatment before and after surgery attenuated this surgically induced impairment.
3.2. Effect of NaHS on partial hepatectomy-elevated serum concentration of proinflammatory cytokines TNF-a, IL-1b, and IL-6 To investigate the effect of partial hepatectomy and/or NaHS administration on the production of serum proinflammatory cytokines after the behavioral experiments, we measured serum TNF-a, IL-1b, and IL-6 by ELISA. Compared with the control group, NaHS alone did not affect the production of serum TNF-a, IL-1b, and IL-6 at any time (P > 0.05) (Fig. 3). Injury caused by partial hepatectomy significantly increased production of serum TNF-a, IL-1b, and IL-6 1 d after surgery, and IL-1b and IL-6 up to 3 d after surgery, but these were still higher than that seen in the NS group (P < 0.01) (Fig. 3). Presurgical and postsurgical administration of NaHS significantly decreased the plasma concentration of TNF-a, IL-1b, and IL-6 (P < 0.05) (Fig. 3).
3.3. Effect of NaHS on expression of TNF-a, IL-1b, and IL-6 proteins in hippocampus After the behavioral experiments, we measured the expression of hippocampal TNF-a, IL-1b, and IL-6 by Western blot. Consistent with the expression of serum proinflammatory cytokines, NaHS did not affect the expression of hippocampal TNF-a, IL-1b, and IL-6 protein on days 1, 3, and 7 after surgery (P > 0.05) (Fig. 4AeC), but partial hepatectomy significantly amplified and prolonged the expression of hippocampal TNF-a, IL-1b, and IL-6 protein (P < 0.05) (Fig. 4AeC). However, after treatment with NaHS, hippocampal TNF-a, IL-1b, and IL-6 expression significantly decreased compared with the surgery group (P < 0.01) (Fig. 4AeC).
4.
Discussion
In this preliminary study, we found that surgical trauma caused by partial hepatectomy led to cognitive deficit detected by the spatial learning and memory test (MWM), and that this cognitive deficit could be significantly alleviated by prophylactic administration of NaHS, an H2S donor. Simultaneously, partial hepatectomy induces an increase in TNF-a, IL-1b, and IL-6 in the serum and in hippocampal expression of these cytokines in adult mice; correspondingly, these changes could be significantly attenuated by preoperative NaHS treatment. Previous studies indicate that surgery induces an increase in proinflammatory cytokines in the serum and hippocampus
335
in adult mice, which may be associated with postoperative cognitive decline [5e7,9e11]. Mounting evidence suggests that neuroinflammation in the CNS, which is mediated by glial activation and the production of inflammatory mediators, may contribute to the neuropathogenesis of POCD [5e7]. Cytokines are central mediators of inflammation after events such as peripheral trauma, and have broad physiological effects both on the periphery and in the CNS [21]. Although low levels of immune activation are necessary for regulating normal cognitive function, high levels of proinflammatory factors associated with trauma have been shown to affect cognitive processes detrimentally [8]. In patient studies, tissue damage associated with surgery activated the peripheral innate immune system, leading to the release of inflammatory mediators such as cytokines, reactive oxygen species, and complements [22e24]. These increases in peripheral levels of inflammatory factors after surgery, such as TNF-a, IL-1b, and IL-6, have been related to a decline in cognitive performance [11,25e27]. In this study, we found surgery induced an increase in TNF-a, IL-1b, and IL-6 in the serum and hippocampus on an early postoperative day and returned to normal 3 d after surgery. In accordance with these findings, surgery was associated with glia activation that lasted up to 3 d after surgery, as well as hippocampal increases in the expression of proinflammatory mediators [5,7,9,11,28]. Impaired hippocampal-dependent learning and memory after partial hepatectomy were similarly observed after infection in a model of peripheral inflammation caused by LPS [28]. Hydrogen sulfide protects neurons against oxidative stress [29] and attenuate production and release of NO and TNFa induced by LPS. It is also a potent trigger of inflammation in primary cultured microglia and astrocytes, as well as murine immortalized BV2 microglial cells [17]. Consistent with this report, Lee et al demonstrated that the anti-neuroinflammatory role of H2S and three H2S-releasing compounds (anethole trithione hydroxide, S-diclofenac, and S-aspirin) [30] attenuates LPS or Ab-induced spatial learning and memory impairment [18e20]. Likewise, Gong et al [18,19] demonstrated that pre- and post-LPS injection treatment with NaHS or S-propargyl-cysteine, a novel H2S-modulated agent, significantly attenuates LPS-induced cognitive deficits and neuronal ultrastructure damage, with concomitant inhibition of TNFa and TNF receptor 1 production, as well as repressed LPSinduced IkB-a degradation and thus activation of nuclear factor-kB. In the present study, pre- and postsurgery treatment with NaHS at the dose of 5 mg/kg is safe, as reported elsewhere [18]. Sodium hydrosulfide significantly attenuated surgery-induced learning and memory deficits and decreased proinflammatory cytokines in the serum and hippocampus. The present study demonstrates that partial hepatectomy triggers neurocognitive decline in adult mice, which appears to be related to systemic inflammation and neuroinflammation. Administration of NaHS prevented this abnormal expression of cytokines and neurofunctional impairment.
= mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point.
# P < 0.05; P < 0.01 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
##
336
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Acknowledgment This work was supported by National Natural Science Foundation of China grant 30901413, to Q.-J.C.; and by Provincial Health Bureau Program of Henan grant 2011020009, to C.-L.L.
references
[1] Bekker AY, Weeks EJ. Cognitive function after anaesthesia in the elderly. Best Pract Res Clin Anaesthesiol 2003;17:259. [2] Steinmetz J, Funder KS, Dahl BT, Rasmussen LS. Depth of anaesthesia and post-operative cognitive dysfunction. Acta Anaesthesiol Scand 2010;54:162. [3] Moller J, Cluitmans P, Rasmussen L, et al. Long-term postoperative cognitive dysfunction in the elderly: ISPOCD1 study. Lancet 1998;351:857. [4] Johnson T, Monk T, Rasmussen LS, et al. Postoperative cognitive dysfunction in middle-aged patients. Anesthesiology 2002;96:1351. [5] Wan Y, Xu J, Ma D, et al. Postoperative impairment of cognitive function in rats: a possible role for cytokinemediated inflammation in the hippocampus. Anesthesiology 2007;106:436. [6] Cao XZ, Ma H, Wang JK, et al. Postoperative cognitive deficits and neuroinflammation in the hippocampus triggered by surgical trauma are exacerbated in aged rats. Prog Neuropsychopharmacol Biol Psychiatry 2010;34:1426. [7] Rosczyk H, Sparkman N, Johnson R. Neuroinflammation and cognitive function in aged mice following minor surgery. Exp Gerontol 2008;43:840. [8] Yirmiya R. Goshen I immune modulation of learning, memory, neural plasticity and neurogenesis. Brain Behav Immun 2011;25:181. [9] Wan Y, Xu J, Meng F, et al. Cognitive decline following major surgery is associated with gliosis, ß-amyloid accumulation, and t phosphorylation in old mice. Crit Care Med 2010;38: 2190. [10] Terrando N, Monaco C, Ma D, et al. Tumor necrosis factoralpha triggers a cytokine cascade yielding postoperative cognitive decline. Proc Natl Acad Sci U S A 2010;107:20518. [11] Cibelli M, Fidalgo AR, Terrando N, et al. Role of interleukin-1b in postoperative cognitive dysfunction. Ann Neurol 2010;68:360. [12] Laalou FZ, Jochum D, Pain L. Postoperative cognitive dysfunction (POCD): strategy of prevention, assessment and management. Ann Fr Anesth Reanim 2011;30:e49. [13] Kimura H, Nagai Y, Umemura K, Kimura Y. Physiological roles of hydrogen sulfide: synaptic modulation, neuroprotection, and smooth muscle relaxation. Antioxid Redox Signal 2005;7:795.
[14] Abe K, Kimura H. The possible role of hydrogen sulfide as an endogenous neuromodulator. J Neurosci 1996;16:1066. [15] Li L, Bhatia M, Moore PK. Hydrogen sulphideda novel mediator of inflammation? Curr Opin Pharmacol 2006;6:125. [16] Li L, Bhatia M, Zhu YZ, et al. Hydrogen sulfide is a novel mediator of lipopolysaccharide-induced inflammation in the mouse. FASEB J 2005;19:1196. [17] Hu LF, Wong PT, Moore PK, Bian JS. Hydrogen sulfide attenuates lipopolysaccharide-induced inflammation by inhibition of p38 mitogen-activated protein kinase in microglia. J Neurochem 2007;100:1121. [18] Gong QH, Wang Q, Pan LL, et al. Hydrogen sulfide attenuates lipopolysaccharide-induced cognitive impairment: a proinflammatory pathway in rats. Pharmacol Biochem Behav 2010;96:52. [19] Gong QH, Wang Q, Pan LL, et al. S-Propargyl-cysteine, a novel hydrogen sulfide-modulated agent, attenuates lipopolysaccharide-induced spatial learning and memory impairment: involvement of TNF signaling and NF-[kappa] B pathway in rats. Brain Behav Immun 2011; 25:110. [20] Gong QH, Pan LL, Liu XH, et al. S-propargyl-cysteine (ZYZ802), a sulphur-containing amino acid, attenuates betaamyloid-induced cognitive deficits and pro-inflammatory response: involvement of ERK1/2 and NF-kB pathway in rats. Amino Acids 2011;40:601. [21] Oppenheim JJ, Feldman M, Durum SK. Cytokine reference: a compendium of cytokines and other mediators of host defense. Waltham, Massachusetts: Academic Press; 2001. [22] Giannoudis PV, Dinopoulos H, Chalidis B, Hall GM. Surgical stress response. Injury 2006;37:S3. [23] Karlidag R, Unal S, Sezer OH, et al. The role of oxidative stress in postoperative delirium. Gen Hosp Psychiatry 2006; 28:418. [24] Levy JH, Tanaka KA. Inflammatory response to cardiopulmonary bypass. Annal Thoracic Surg 2003;75:S715. [25] Beloosesky Y, Hendel D, Weiss A, et al. Cytokines and Creactive protein production in hip-fracture-operated elderly patients. J Gerontol A Biol Sci Med Sci 2007;62:420. [26] Ramlawi B, Rudolph JL, Mieno S, et al. C-Reactive protein and inflammatory response associated to neurocognitive decline following cardiac surgery. Surgery 2006;140:221. [27] Yaffe K, Lindquist K, Penninx B, et al. Inflammatory markers and cognition in well-functioning African-American and white elders. Neurology 2003;61:76. [28] Fidalgo AR, Cibelli M, White JPM, et al. Systemic inflammation enhances surgery-induced cognitive dysfunction in mice. Neurosci Lett 2011;498:63. [29] Kimura Y, Kimura H. Hydrogen sulfide protects neurons from oxidative stress. FASEB J 2004;18:1165. [30] Lee M, Sparatore A, Del Soldato P, et al. Hydrogen sulfidereleasing NSAIDs attenuate neuroinflammation induced by microglial and astrocytic activation. Glia 2010;58:103.
Available online at www.sciencedirect.com
journal homepage: www.JournalofSurgicalResearch.com
Hydrogen sulfide attenuates surgical trauma-induced inflammatory response and cognitive deficits in mice Qin-Jun Chu, MD,a,1 Long He, MD,a,1 Wei Zhang, MD,a Chun-Lan Liu, MD,a Yan-Qiu Ai, MD,a,* and Qi Zhang, PhDb,** a b
Department of Anesthesiology, First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China Department of pharmacology, School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, Henan, China
article info
abstract
Article history:
Background: It has been increasingly reported that peripheral surgical trauma triggers
Received 2 October 2012
neuroinflammatory processes associated with postoperative cognitive dysfunction, and
Received in revised form
that mitigating the neuroinflammatory effects of surgery prevents surgery-induced
2 December 2012
cognitive dysfunction. Endogenously produced hydrogen sulfide (H2S) has multiple func-
Accepted 3 December 2012
tions in the brain, and an increasing number of studies have demonstrated its anti-
Available online 21 December 2012
inflammatory effects. The present study was designed to investigate the effects of
Keywords:
experience neuroinflammatory changes induced by surgery.
Postoperative cognitive dysfunction
Methods: Each mouse received 5 mg/kg NaHS or volume-matched vehicle administration
Inflammatory response
by intraperitoneal injection once daily, 3 d before surgery, on the day of surgery, and for 3
Hydrogen sulfide
d afterward. We assessed cognitive function using a Morris water maze and evaluated
Mice
expression of proinflammatory cytokines tumor necrosis factor-a, interleukin-1b, and
sodium hydrosulfide (NaHS), an H2S donor, on the cognitive impairment of mice as they
interleukin-6 in the serum and hippocampus. We performed each test 1, 3, and 7 d after surgery. Results: Hippocampal-dependent memory impairment in mice after surgery was associated with increased serum proinflammatory cytokines, as well as proinflammatory cytokine expression in the hippocampus. Presurgery treatment with NaHS, an H2S donor, significantly attenuated surgery-induced memory impairment and expression of proinflammatory cytokines in the serum and hippocampus. Conclusions: These findings suggest that intraperitoneal injections of NaHS could significantly mitigate surgery-induced memory impairment in mice, which is strongly associated with reduced levels of serum and hippocampal proinflammatory cytokines. ª 2013 Elsevier Inc. All rights reserved.
1.
Introduction
After surgery, patients may experience cognitive decline. This complication has been termed postoperative cognitive dysfunction (POCD). It is characterized by disruption of
attention, memory, orientation, information processing, and the sleepewake cycle, which leads to an increased risk of disability and mortality [1,2]. Elderly patients seem to be at high risk of POCD [3], but other age groups are also affected [4]. At present, little is known about the pathophysiological
* Corresponding author. Department of Anesthesiology, First Affiliated Hospital, Zhengzhou University, No. 1 East-Jianshe Road, Zhengzhou 450052, Henan, China. Tel.:þ86 0371 66862024; fax: þ86 0371 66862025. ** Corresponding author. School of Pharmaceutical Sciences, Zhengzhou University, 100 Kexue Avenue, Zhengzhou 450001, Henan, China. Tel.: þ86 0371 67781908; fax: þ86 0371 67781908. E-mail addresses: [email protected] (Y.-Q. Ai), [email protected] (Q. Zhang). 1 Qin-Jun Chu and Long He contributed equally to this research. 0022-4804/$ e see front matter ª 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.jss.2012.12.003
j o u r n a l o f s u r g i c a l r e s e a r c h 1 8 3 ( 2 0 1 3 ) 3 3 0 e3 3 6
mechanisms of POCD. Consistent evidence is accumulating for the role of inflammatory processes arising as the result of surgical trauma and subsequent complications [5e7]. Earlier studies have shown that activating the peripheral inflammatory response can profoundly affect the function of the central nervous system (CNS) [8]. It is well known that surgeryinduced tissue damage can activate the peripheral innate immune system, which in turn results in the release of inflammatory mediators. Animal studies show that surgery causes learning and memory impairment that may be associated with increased levels of plasma cytokine, reactive microgliosis, tumor necrosis factor-a (TNF-a), and interleukin1b (IL-1b) transcription and expression in the hippocampus [5,9e11]. Simultaneously, nonspecific attenuation of innate immunity or preemptive treatment with anti-TNF antibodies can prevent neuroinflammatory processes as well as learning and memory dysfunction [9e12]. Hydrogen sulfide (H2S), which is well known traditionally as a toxic gas, has been proven to be produced endogenously in mammalian tissues and has an important role in physiological and pathophysiological conditions [13,14]. In recent years, there has been emerging evidence that the endogenous gaseous molecule can modulate the inflammatory process [15e17]. In the CNS, H2S exhibited extensive neurological effects in a series of in vitro and in vivo models. It was reported that H2S attenuates lipopolysaccharide (LPS)-induced cognitive impairment by reducing the overproduction of proinflammatory mediators [18]. Recently, S-propargyl-cysteine, a novel hydrogen sulfide-modulated agent, attenuated LPS- or Ab-induced spatial learning and memory impairment [19,20]. Therefore, based on these findings, we hypothesized that an H2S-modulated agent may be useful to treat or prevent POCD.
2.
Materials and methods
2.1.
Animals
The Henan Laboratory Animal Center, Henan, China, provided adult male Kunming mice weighing 30e40 g. Animals were housed in a temperature and humidity-controlled room with a 12 h/12 h lightedark cycle for at least 1 wk before the day of operation, and were allowed free access to food and tap water throughout the experiment. All animal studies were performed in accordance with the Regulations of Experimental Animal Administration issued by the State Committee of Science and Technology of the People’s Republic of China and the Institutional Review Committee for the Use of Animals.
2.2.
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NaHS dosage from pilot studies to ascertain the dose with the highest desired pharmacologic effect. We treated the surgery mice (group S; n ¼ 24) as in the C group, except that they underwent partial hepatectomy under general anesthesia. They were anesthetized with a regimen (fentanyl 0.2 mg/kg and droperidol 5 mg/kg) that we predetermined from pilot studies. Surgery plus NaHS mice (group NS; n ¼ 24) were treated as in the S group, except that they received daily intraperitoneal NaHS solution (5 mg/kg/d) Subjects in all four groups received the same volume of NaHS resolution or placebo. We carried out all surgical procedures under aseptic conditions. Briefly, we exposed the liver through an approximately 2-cm midline abdominal incision. We excised the left lateral lobes of the liver (corresponding to approximately one-third of the organ). We then infiltrated the wound with 0.25% bupivacaine and closed it by sterile suture. We took care to ensure that all mice recovered fully from anesthesia and surgery before being individually housed. We killed the mice on postoperative days 1, 3, and 7 (n ¼ 32/d) after spatial working tests.
2.3.
Cognitive testing: Morris Water Maze
The Morris water maze (MWM) is a hippocampal-dependent test of spatial learning for rodents. The MWM consists of a pool of water (120 cm in diameter and 40 cm high) maintained at a temperature of 26 C 1 C with a hidden submerged platform (1 cm below the surface) in one quadrant. We placed mice in the water in a dimly lit room with various visual clues around the pool. We used a video camera mounted to the ceiling directly above the center of the tank in conjunction with a computerized animal tracking system to monitor swim speed, escape latency, and the distance swum. We placed mice on the platform for 30 s before the start of each trial, and released them into the water facing the wall of the pool from one of four randomly assigned release points (north, south, east, and west). In all trials, mice were allowed to swim until they landed on the platform. If a mouse failed to find the platform within 60 s, it was picked up and placed on the platform for 30 s. We trained mice with the platform in a fixed location for 4 trials/d for 5 consecutive days. Data from 4 trials/ d were averaged to represent a performance block. Animals underwent surgery on day 6. In addition, on postoperative days 1, 3, and 7, were subjected mice to a probe trial; the platform was removed and all but distal visual cues remained constant. Animals were allowed to swim for 60 s. We recorded the percentage of time spent in the east quadrant previously containing the submerged platform and the number of crossings through the target positiondthat is, the specific location of the platform during the acquisition phasedand represented it as an index of memory. All trials lasted between 9 and 16 h.
Surgery and pharmacological treatments
We randomly assigned adult mice (n ¼ 96) to four groups and then trained them on the water maze cognitive test for 5 consecutive days. Control mice (group C; n ¼ 24) received 0.3 mL normal saline (NS) intraperitoneally daily for 3 d before surgery, on the day of surgery, and for the next 3 d. We treated the NaHS mice (group N; n ¼ 24) as in the C group, except that they received intraperitoneal NaHS (Sigma, St. Louis, MO) resolution at a dose of 5 mg/kg/d daily. We determined the
2.4. Enzyme-linked immunosorbent assay for serum TNF-a, IL-1b, and IL-6 To characterize TNF-a, IL-1b, and IL-6 expression, we collected blood samples from below the canthus of the eye of each mouse (from different treatment groups). We pooled together sera of mice from two separate experiments (from different treatment groups) and subsequently used quantitative direct sandwich enzyme-linked immunosorbent assay (ELISA) (R&D
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Systems; Minneapolis, MN) to determine the concentration of TNF-a, IL-1b, and IL-6 (in picograms per milliliter). We conducted the procedure according to the manufacturer’s instructions.
2.5. Western blotting for hippocampal TNF-a, IL-1b, and IL-6 protein We homogenized hippocampus in a lysis buffer (20 mmol/L Tris, pH 7.5; 150 mmol/L sodium chloride; 1% Triton X-100; 1% pyrophosphate; 1% b-glycophosphate; 1% Nonidet P-40; 0.1% sodium dodecyl sulfate; 1 mmol/L ethylenediaminetetra acetic acid; 100 mg/mL phenylmethanesulfonylfluoride; and 1 mg/mL aprotinin). We processed the homogenates in a centrifuge for 15 min at 12,000 g at 4 C. We determined the quantity of protein in the supernatants using a bicinchoninic acid protein assay kit (Pierce Biotechnology, Rockford, IL). Protein extracts (30 mg/sample) denatured in Laemmli sample loading buffer at 100 C for 5 min were separated by 12% sodium dodecyl sulfateepolyacrylamide gel electrophoresis and then transferred onto nitrocellulose membranes (Hybond ECL; Amersham, Piscataway, NJ). We blocked membranes with 5% skim milk in Tris-buffered salineeTween 20 for 2 h and incubated them with rabbit polyclonal anti-TNF-a (1:1000; Abcam), rabbit polyclonal anti-IL-1b (1:700; Abcam), or rabbit polyclonal anti-IL-6 (1:500; Abcam) overnight at 4 C. After thorough washing, we incubated membranes in Tris-buffered salineeTween 20 with appropriate secondary antibodies (goat anti-rabbit, conjugated with alkaline phosphatase; Jacksonimmuno Research Laboratories, Inc, PA) diluted 1:5000 for 2 h at room temperature. Visualization of the proteins was accomplished using an enhanced chemiluminescence detection kit (ECL; Amersham) and quantified by densitometric analysis (Kodak Digital Science and ID Image Analysis software; Rochester, NY). We stripped the membrane blots and reincubated them with monoclonal anti-b-actin (AC-15; Sigma, St. Louis, MO). Relative expression levels of protein were normalized by the ratio of target protein (TNF-a, IL-1b, and IL-6) to b-actin.
2.6.
Statistical analysis
All data are expressed as mean standard error of the mean. We analyzed the behavioral data by repeated measures analysis of variance, using the Bonferroni post hoc test to determine specific group differences. We tested intergroup differences of the rest of the data using one-way analysis of variance followed by the least significant difference t-test for multiple comparisons. We performed statistical analysis with SPSS 17.0 software (SPSS Inc, Chicago, IL). In all tests, P < 0.05 indicated statistical significance.
3.
Results
3.1. Effect of NaHS on partial hepatectomy-induced spatial memory impairment To elucidate whether cognitive impairment as defined by a decline in hippocampal-dependent spatial learning and memory could be induced by partial hepatectomy, and whether this impairment could be prevented by NaHS
Fig. 1 e Performance of mice during 5-d acquisition phase in the Morris water maze. (A) Latency to the platform across training days. (B) Swimming distance to the platform across training days. (C) Swimming speed during training days. Data are represented as mean ± standard error of the mean.
administration, we conducted the MWM test. Mice underwent water maze training for 5 consecutive days (acquisition trials). As expected, animals in all groups were able to rescue themselves, and the escape latency and swimming distance gradually decreased over the 4 consecutive training days. Repeated-measures analysis of variance of swim data revealed significant effects by training day for both escape latency and swimming distance (P < 0.05) (Fig. 1A and B) but not speed (P > 0.05) (Fig. 1C). These results indicated that mice showed improvement in spatial learning and memory over time.
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Fig. 2 e Effect of NaHS on hepatectomy, resulting in cognitive impairment in mice in the Morris water maze during the probe trial test. (A) Percentage of time searching each quadrant. (B) Number of crossings over the platform. (C) Swimming speed during probe trial test. Data are represented as mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point. #P < 0.05; ##P < 0.05 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
After the acquisition trials, on postsurgical days 1, 3, and 7, we subjected mice to a probe trial. Probe trials of the MWM test showed that NaHS treatment in the NaHS group did not affect the cognitive function of mice, whereas the learning memory abilities of hepato-lobectomized mice were significantly impaired compared with those in the control group.
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Fig. 3 e Effect of NaHS on surgery-induced systemic inflammation. We measured TNF-a, IL-1b, and IL-6 levels in serum by ELISA at 1, 3, and 7 d after surgery. (A) Tumor necrosis factor-a, (B) IL-1b, and (C) IL-6. Data are represented as mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point. #P < 0.05; ##P < 0.01 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
The percentage of time spent in the quadrant with the previously located hidden platform and the number of crossings over the platform were significantly less in hepatolobectomized mice (P < 0.05) (Fig. 2A and B), indicating cognitive impairment. However, surgery-induced learning and memory deficits were significantly alleviated by treatment with NaHS compared with the surgery group (P < 0.05) (Fig. 2A and B). On postsurgical days 1, 3, and 7, there was no
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Fig. 4 e Effect of NaHS on surgery-induced neuroinflammation in hippocampus. We measured TNF-a, IL-1b, and IL-6 protein by Western blot at 1, 3, and 7 d after surgery. (A) Tumor necrosis factor-a, (B) IL-1b, and (C) IL-6. Data are represented as
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significant difference in average swimming speed among groups, which suggests that the poorer performance of surgical mice did not result from a lack of motivation or reduced motor ability (P > 0.05) (Fig. 2C). These results indicate that partial hepatectomy induced significant cognitive impairment and that NaHS treatment before and after surgery attenuated this surgically induced impairment.
3.2. Effect of NaHS on partial hepatectomy-elevated serum concentration of proinflammatory cytokines TNF-a, IL-1b, and IL-6 To investigate the effect of partial hepatectomy and/or NaHS administration on the production of serum proinflammatory cytokines after the behavioral experiments, we measured serum TNF-a, IL-1b, and IL-6 by ELISA. Compared with the control group, NaHS alone did not affect the production of serum TNF-a, IL-1b, and IL-6 at any time (P > 0.05) (Fig. 3). Injury caused by partial hepatectomy significantly increased production of serum TNF-a, IL-1b, and IL-6 1 d after surgery, and IL-1b and IL-6 up to 3 d after surgery, but these were still higher than that seen in the NS group (P < 0.01) (Fig. 3). Presurgical and postsurgical administration of NaHS significantly decreased the plasma concentration of TNF-a, IL-1b, and IL-6 (P < 0.05) (Fig. 3).
3.3. Effect of NaHS on expression of TNF-a, IL-1b, and IL-6 proteins in hippocampus After the behavioral experiments, we measured the expression of hippocampal TNF-a, IL-1b, and IL-6 by Western blot. Consistent with the expression of serum proinflammatory cytokines, NaHS did not affect the expression of hippocampal TNF-a, IL-1b, and IL-6 protein on days 1, 3, and 7 after surgery (P > 0.05) (Fig. 4AeC), but partial hepatectomy significantly amplified and prolonged the expression of hippocampal TNF-a, IL-1b, and IL-6 protein (P < 0.05) (Fig. 4AeC). However, after treatment with NaHS, hippocampal TNF-a, IL-1b, and IL-6 expression significantly decreased compared with the surgery group (P < 0.01) (Fig. 4AeC).
4.
Discussion
In this preliminary study, we found that surgical trauma caused by partial hepatectomy led to cognitive deficit detected by the spatial learning and memory test (MWM), and that this cognitive deficit could be significantly alleviated by prophylactic administration of NaHS, an H2S donor. Simultaneously, partial hepatectomy induces an increase in TNF-a, IL-1b, and IL-6 in the serum and in hippocampal expression of these cytokines in adult mice; correspondingly, these changes could be significantly attenuated by preoperative NaHS treatment. Previous studies indicate that surgery induces an increase in proinflammatory cytokines in the serum and hippocampus
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in adult mice, which may be associated with postoperative cognitive decline [5e7,9e11]. Mounting evidence suggests that neuroinflammation in the CNS, which is mediated by glial activation and the production of inflammatory mediators, may contribute to the neuropathogenesis of POCD [5e7]. Cytokines are central mediators of inflammation after events such as peripheral trauma, and have broad physiological effects both on the periphery and in the CNS [21]. Although low levels of immune activation are necessary for regulating normal cognitive function, high levels of proinflammatory factors associated with trauma have been shown to affect cognitive processes detrimentally [8]. In patient studies, tissue damage associated with surgery activated the peripheral innate immune system, leading to the release of inflammatory mediators such as cytokines, reactive oxygen species, and complements [22e24]. These increases in peripheral levels of inflammatory factors after surgery, such as TNF-a, IL-1b, and IL-6, have been related to a decline in cognitive performance [11,25e27]. In this study, we found surgery induced an increase in TNF-a, IL-1b, and IL-6 in the serum and hippocampus on an early postoperative day and returned to normal 3 d after surgery. In accordance with these findings, surgery was associated with glia activation that lasted up to 3 d after surgery, as well as hippocampal increases in the expression of proinflammatory mediators [5,7,9,11,28]. Impaired hippocampal-dependent learning and memory after partial hepatectomy were similarly observed after infection in a model of peripheral inflammation caused by LPS [28]. Hydrogen sulfide protects neurons against oxidative stress [29] and attenuate production and release of NO and TNFa induced by LPS. It is also a potent trigger of inflammation in primary cultured microglia and astrocytes, as well as murine immortalized BV2 microglial cells [17]. Consistent with this report, Lee et al demonstrated that the anti-neuroinflammatory role of H2S and three H2S-releasing compounds (anethole trithione hydroxide, S-diclofenac, and S-aspirin) [30] attenuates LPS or Ab-induced spatial learning and memory impairment [18e20]. Likewise, Gong et al [18,19] demonstrated that pre- and post-LPS injection treatment with NaHS or S-propargyl-cysteine, a novel H2S-modulated agent, significantly attenuates LPS-induced cognitive deficits and neuronal ultrastructure damage, with concomitant inhibition of TNFa and TNF receptor 1 production, as well as repressed LPSinduced IkB-a degradation and thus activation of nuclear factor-kB. In the present study, pre- and postsurgery treatment with NaHS at the dose of 5 mg/kg is safe, as reported elsewhere [18]. Sodium hydrosulfide significantly attenuated surgery-induced learning and memory deficits and decreased proinflammatory cytokines in the serum and hippocampus. The present study demonstrates that partial hepatectomy triggers neurocognitive decline in adult mice, which appears to be related to systemic inflammation and neuroinflammation. Administration of NaHS prevented this abnormal expression of cytokines and neurofunctional impairment.
= mean ± standard error of the mean. *P < 0.05; **P < 0.01 versus NS group at the corresponding time point.
# P < 0.05; P < 0.01 versus surgery group at the corresponding time point. P1, P2, and P3 [ surgery group on days 1, 3, and 7 after surgery.
##
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Acknowledgment This work was supported by National Natural Science Foundation of China grant 30901413, to Q.-J.C.; and by Provincial Health Bureau Program of Henan grant 2011020009, to C.-L.L.
references
[1] Bekker AY, Weeks EJ. Cognitive function after anaesthesia in the elderly. Best Pract Res Clin Anaesthesiol 2003;17:259. [2] Steinmetz J, Funder KS, Dahl BT, Rasmussen LS. Depth of anaesthesia and post-operative cognitive dysfunction. Acta Anaesthesiol Scand 2010;54:162. [3] Moller J, Cluitmans P, Rasmussen L, et al. Long-term postoperative cognitive dysfunction in the elderly: ISPOCD1 study. Lancet 1998;351:857. [4] Johnson T, Monk T, Rasmussen LS, et al. Postoperative cognitive dysfunction in middle-aged patients. Anesthesiology 2002;96:1351. [5] Wan Y, Xu J, Ma D, et al. Postoperative impairment of cognitive function in rats: a possible role for cytokinemediated inflammation in the hippocampus. Anesthesiology 2007;106:436. [6] Cao XZ, Ma H, Wang JK, et al. Postoperative cognitive deficits and neuroinflammation in the hippocampus triggered by surgical trauma are exacerbated in aged rats. Prog Neuropsychopharmacol Biol Psychiatry 2010;34:1426. [7] Rosczyk H, Sparkman N, Johnson R. Neuroinflammation and cognitive function in aged mice following minor surgery. Exp Gerontol 2008;43:840. [8] Yirmiya R. Goshen I immune modulation of learning, memory, neural plasticity and neurogenesis. Brain Behav Immun 2011;25:181. [9] Wan Y, Xu J, Meng F, et al. Cognitive decline following major surgery is associated with gliosis, ß-amyloid accumulation, and t phosphorylation in old mice. Crit Care Med 2010;38: 2190. [10] Terrando N, Monaco C, Ma D, et al. Tumor necrosis factoralpha triggers a cytokine cascade yielding postoperative cognitive decline. Proc Natl Acad Sci U S A 2010;107:20518. [11] Cibelli M, Fidalgo AR, Terrando N, et al. Role of interleukin-1b in postoperative cognitive dysfunction. Ann Neurol 2010;68:360. [12] Laalou FZ, Jochum D, Pain L. Postoperative cognitive dysfunction (POCD): strategy of prevention, assessment and management. Ann Fr Anesth Reanim 2011;30:e49. [13] Kimura H, Nagai Y, Umemura K, Kimura Y. Physiological roles of hydrogen sulfide: synaptic modulation, neuroprotection, and smooth muscle relaxation. Antioxid Redox Signal 2005;7:795.
[14] Abe K, Kimura H. The possible role of hydrogen sulfide as an endogenous neuromodulator. J Neurosci 1996;16:1066. [15] Li L, Bhatia M, Moore PK. Hydrogen sulphideda novel mediator of inflammation? Curr Opin Pharmacol 2006;6:125. [16] Li L, Bhatia M, Zhu YZ, et al. Hydrogen sulfide is a novel mediator of lipopolysaccharide-induced inflammation in the mouse. FASEB J 2005;19:1196. [17] Hu LF, Wong PT, Moore PK, Bian JS. Hydrogen sulfide attenuates lipopolysaccharide-induced inflammation by inhibition of p38 mitogen-activated protein kinase in microglia. J Neurochem 2007;100:1121. [18] Gong QH, Wang Q, Pan LL, et al. Hydrogen sulfide attenuates lipopolysaccharide-induced cognitive impairment: a proinflammatory pathway in rats. Pharmacol Biochem Behav 2010;96:52. [19] Gong QH, Wang Q, Pan LL, et al. S-Propargyl-cysteine, a novel hydrogen sulfide-modulated agent, attenuates lipopolysaccharide-induced spatial learning and memory impairment: involvement of TNF signaling and NF-[kappa] B pathway in rats. Brain Behav Immun 2011; 25:110. [20] Gong QH, Pan LL, Liu XH, et al. S-propargyl-cysteine (ZYZ802), a sulphur-containing amino acid, attenuates betaamyloid-induced cognitive deficits and pro-inflammatory response: involvement of ERK1/2 and NF-kB pathway in rats. Amino Acids 2011;40:601. [21] Oppenheim JJ, Feldman M, Durum SK. Cytokine reference: a compendium of cytokines and other mediators of host defense. Waltham, Massachusetts: Academic Press; 2001. [22] Giannoudis PV, Dinopoulos H, Chalidis B, Hall GM. Surgical stress response. Injury 2006;37:S3. [23] Karlidag R, Unal S, Sezer OH, et al. The role of oxidative stress in postoperative delirium. Gen Hosp Psychiatry 2006; 28:418. [24] Levy JH, Tanaka KA. Inflammatory response to cardiopulmonary bypass. Annal Thoracic Surg 2003;75:S715. [25] Beloosesky Y, Hendel D, Weiss A, et al. Cytokines and Creactive protein production in hip-fracture-operated elderly patients. J Gerontol A Biol Sci Med Sci 2007;62:420. [26] Ramlawi B, Rudolph JL, Mieno S, et al. C-Reactive protein and inflammatory response associated to neurocognitive decline following cardiac surgery. Surgery 2006;140:221. [27] Yaffe K, Lindquist K, Penninx B, et al. Inflammatory markers and cognition in well-functioning African-American and white elders. Neurology 2003;61:76. [28] Fidalgo AR, Cibelli M, White JPM, et al. Systemic inflammation enhances surgery-induced cognitive dysfunction in mice. Neurosci Lett 2011;498:63. [29] Kimura Y, Kimura H. Hydrogen sulfide protects neurons from oxidative stress. FASEB J 2004;18:1165. [30] Lee M, Sparatore A, Del Soldato P, et al. Hydrogen sulfidereleasing NSAIDs attenuate neuroinflammation induced by microglial and astrocytic activation. Glia 2010;58:103.