Products of Cyclooxygenase-2 Catalysis Regulate Postoperative Bowel Motility

Journal of Surgical Research 86, 50 –54 (1999) Article ID jsre.1999.5692, available online at http://www.idealibrary.com on

Products of Cyclooxygenase-2 Catalysis Regulate Postoperative Bowel Motility Michael D. Josephs, M.D., Guozhang Cheng, M.D., Riadh Ksontini, M.D., Lyle L. Moldawer, Ph.D., and Michael P. Hocking, M.D. Department of Surgery, University of Florida College of Medicine and Veterans Affairs Medical Center, Gainesville, Florida 32610-0286 Submitted for publication November 18, 1998

INTRODUCTION Laparotomy involving manipulation of the small intestine causes injury, initiating an inflammatory cascade in the small bowel wall, which generates eicosanoids and proinflammatory cytokines. We have shown that ketorolac and salsalate, nonselective cyclooxygenase (COX) inhibitors, ameliorate postoperative small bowel ileus in a rodent model. Others have shown that interleukin-1 receptor antagonism improves postoperative gastric emptying. We examined whether inhibition of the proinflammatory cytokines, tumor necrosis factor a (TNFa) and interleukin-1 (IL-1), or selective blockade of cyclooxygenase-2 (COX2), the COX isoform induced during inflammation, would accelerate postoperative small bowel transit in our model. Duodenostomy tubes were inserted into male Sprague–Dawley rats. One week later, animals were randomized to receive TNF-binding protein (TNF-bp), IL-1 receptor antagonist (IL-1ra), or saline (NS) prior to standardized laparotomy. Additional rats were gavaged preoperatively with a selective COX-2 inhibitor (NS-398) or NS. Small intestinal transit was measured as the geometric center (GC) of distribution of 51CrO 4 at 30 min, 3 h, or 6 h (n 5 5–9 rats/group) following laparotomy. Selective inhibition of COX-2 significantly increased postoperative small bowel transit compared to controls (GC 2.9 6 0.3 vs 2.2 6 0.1 at 30 min, GC 2.9 6 0.3 vs 2.5 6 0.2 at 3 h, and GC 3.3 6 0.3 vs 2.8 6 0.2 at 6 h, P < 0.05). In contrast, neither TNF-bp nor IL-1ra altered postoperative small intestinal transit in this model. Use of selective COX-2 inhibitors may accelerate recovery of postoperative bowel dysmotility without the undesirable effects (e.g., gastrointestinal irritation and anti-platelet effect) of nonselective COX inhibitors. © 1999 Academic Press Key Words: COX-2; TNFa; IL-1ra; ileus; motility.

For over a century it has been recognized that abdominal surgery inhibits bowel motility [1]. This inhibition has been referred to as physiologic [2] or functional [3] and usually resolves spontaneously within 4 to 5 days. Ileus has generally not been a concern unless prolonged, when it is termed paralytic ileus [4]. However, even routine, uncomplicated postoperative ileus prolongs patient discomfort, lengthens hospitalization, and increases medical costs [5]. Despite nearly a hundred years of study, no effective therapy has been developed to specifically prevent or shorten postoperative ileus, and treatment remains primarily supportive [6]. Functional ileus is thought to occur via excitation of afferent sensory neurons, with resulting reflex activation of sympathetic efferent pathways via the prevertebral ganglia and central nervous system [6]. The majority of previous clinical and experimental approaches have been designed to intervene with efferent pathways using sympatholytic or cholemimetic agents to counteract increased sympathetic tone [6]. The beneficial effect from these drugs, however, has largely been overshadowed by their systemic side effects [6]. An alternative approach has been the use of prokinetic agents such as metoclopramide, erythromycin, and cisapride to stimulate postoperative gastrointestinal motility. Trials using these agents, however, have shown them to be ineffective [7–9]. Few studies have explored the afferent pathways of postoperative ileus. Tissue trauma, as occurs with abdominal surgery, initiates an inflammatory cascade in the small bowel wall, generating eicosanoids such as prostacyclin (PGI 2 ) [10] and proinflammatory cytokines such as interleukin-1 (IL-1) and tumor necrosis factor a (TNFa) [11–13]. Eicosanoids are liberated from arachidonic acid (AA) by the catalytic effects of

Presented at the Annual Meeting of the Association for Academic Surgery, Seattle, Washington, November 18 –22, 1998

0022-4804/99 $30.00 Copyright © 1999 by Academic Press All rights of reproduction in any form reserved.

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JOSEPHS ET AL.: ROLE OF COX-2 IN POSTOPERATIVE ILEUS

cyclooxygenase (COX). COX exists as two isoforms, COX-1 and COX-2. Both metabolize AA to form the prostaglandins and thromboxanes; however, COX-1 is a constitutively expressed gene, whereas expression of COX-2 is induced by a variety of cytokines, hormones, and growth factors, including the inflammatory mediators IL-1 and TNFa [14 –16]. We have demonstrated that the nonsteroidal anti-inflammatory drugs ketorolac and salsalate, nonselective COX inhibitors, improve small bowel transit and myoelectric activity in a rodent model of postoperative ileus [17, 18]. However, a problem with using these agents clinically is their anti-platelet effect, which could increase postoperative bleeding, as well as their gastric irritant effect, even when administered parenterally. Others have shown that IL-1 receptor antagonism improves postoperative gastric emptying [19]. The current study examined whether inhibition of TNFa, IL-1, or COX-2, the COX isoform induced during inflammatory states, accelerates postoperative small bowel transit in a rodent model of postoperative ileus. METHODS Animals. Male Sprague–Dawley rats (Harlan Sprague–Dawley, Inc., Indianapolis, IN) weighing 275–300 g were acclimated to the facility for 7–10 days. Animals were maintained under conditions of controlled temperature and illumination and were provided with chow and water ad libitum. Prior to all surgical procedures, they were deprived of food for 18 –20 h. This study protocol was approved by both the University of Florida College of Medicine Institutional Animal Care and Use Committee and the Gainesville Veteran’s Administration Animal Care and Use Committee. Small intestinal transit studies. Under ketamine anesthesia (75 mg/kg body weight [BW]) with acepromazine (1 mg/kg BW), silastic catheters were inserted into the duodenum via a midline laparotomy, tunneled through the lateral abdominal wall, and exteriorized between the scapulae dorsally. The catheters were temporarily occluded and secured into place for future use. After recovery, animals were returned to their cages and feeding was resumed. One week following placement of the duodenostomy tubes, rats were randomized into one of three treatment groups. Group I animals received tumor necrosis factor-binding protein (TNF-bp, 5 mg/kg BW intraperitoneally [ip]; Amgen, Inc., Boulder, CO) or physiologic saline (NS) 30 min prior to a standardized laparotomy. Group II rats were administered interleukin-1 receptor antagonist (IL-1ra, 100 mg/kg BW ip; Amgen, Inc.) or NS 30 min before laparotomy. Both TNF-bp and IL-1ra have been shown to neutralize the physiologic effects of TNF and IL-1, respectively, in rodent models of inflammation [20, 21]. Group III animals were gavaged with NS-398 (1 mg/kg BW; Cayman Chemical Co., Ann Arbor, MI), a selective COX-2 inhibitor, or NS 1 h prior to laparotomy. The standardized laparotomy consisted of a midline incision under ketamine anesthesia, with manipulation of the stomach, small bowel, and colon for 1 min each, followed by small bowel evisceration for 5 min, and abdominal closure. Thirty minutes, 3 h, or 6 h following laparotomy (n 5 5–9 rats/time point), the animals were administered an enteral bolus of 0.2 ml Na 51CrO 4 (Amersham Bioscience, Boston, MA) in NS via the duodenostomy catheter. Na 51CrO 4 is not absorbed by the small intestine and has been validated as a marker of intestinal propulsion [15, 16, 22]. Animals were sacrificed with sodium pentobarbital (100 mg/kg BW ip) 25 min following Na 51CrO 4 administration. The small intes-

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FIG. 1. Small intestinal transit in rats treated with TNF-binding protein (TNF-bp) or physiologic saline (NS). TNFa antagonism with TNF-bp failed to improve small intestinal transit after standardized laparotomy. GC, geometric center of radiolabel distribution. tine was ligated proximal to the duodenostomy tubes and at the terminal ileum, carefully excised, divided into 10 equal segments, and placed into scintillation vials. The radioactivity in each segment was determined with a Beckman Gamma 5500 (Beckman Instruments, Irvine, CA) gamma counter and expressed as the percentage of activity per segment. Small bowel transit was calculated as the geometric center (GC) of distribution of 51CrO 4, using the formula

O 1

GC 5

(fraction of radioactivity in ROI n) n,

n

where ROI is the segment number, and n varies from 1 to 10 for the 10 equal segments. The GC accurately reflects the transit of small bowel contents in this model [23, 24], with higher values indicating more distal passage of intraluminal contents. Statistical analysis. All data are expressed as means 6 SEM. Differences in small bowel transit were compared using two-way analysis of variance and Student–Newman–Keuls post hoc analysis. A 95% confidence interval (P , 0.05) was used to determine statistical significance.

RESULTS

There was no operative mortality from either duodenostomy tube insertion or standardized laparotomy. Preoperative administration of the cytokine inhibitors TNF-bp and IL-1ra had no effect on small intestinal transit following laparotomy (Figs. 1 and 2). In contrast, selective inhibition of COX-2 with NS-398 resulted in significantly increased small bowel transit after laparotomy compared to control animals (Fig. 3, P , 0.05). Furthermore, this effect was not timedependent, as the GC of radiolabel distribution was improved in the NS-398 groups compared to controls at all time points evaluated. DISCUSSION

Postoperative ileus, defined as a decrease or temporary loss of organized aboral progression of gastroin-

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testinal contents following abdominal surgery [6, 25], remains a major cause of morbidity in surgical patients. This disorder is directly responsible for substantial patient discomfort and prolonged hospital stays due to abdominal pain, nausea, and vomiting. In addition, current modalities of treatment, which remain supportive and include nasogastric tubes and intravenous catheters, add to this discomfort and further increase medical costs. The trend toward increased use of laparoscopy is thought to have had an impact on postoperative length of stay. However, despite the general impression that laparoscopic approaches are not associated with ileus, currently there is no evidence that laparoscopic bowel resection is associated with a shortened period of ileus compared to open surgery [26, 27]. Furthermore, at present, it does not appear that all abdominal operations will be amenable to a laparoscopic approach. Despite Oschner’s description in 1930 of the treatment of ileus with medication [28], nearly 70 years later, no definitive therapies are available for this incapacitating condition. Thus, postoperative ileus will likely remain a common reason for hospitalization for the foreseeable future. Previous studies from our laboratory have shown that nonselective COX inhibitors increase small bowel transit in a rodent model of postoperative ileus [18, 19]. The mechanism of action of these drugs in stimulating postoperative small bowel motility is unknown, but is likely related to their inhibitory effect on eicosanoid synthesis. PGI 2 delays transit and inhibits intestinal contractions [29, 30], similar to the motility changes which occur following surgery. This inhibitory effect is presumably mediated by PGI 2’s direct effect on smooth muscle cells, as well as by stimulation of visceral afferent sensory nerves [31]. The importance of visceral sensory nerves in the development of postoperative ileus has previously been demonstrated by Holzer et al.

FIG. 2. Small intestinal transit in rats treated with IL-1 receptor antagonist (IL-1ra) or physiologic saline (NS). IL-1 antagonism with IL-1ra failed to improve small intestinal transit after standardized laparotomy. GC, geometric center of radiolabel distribution.

FIG. 3. Small intestinal transit in rats treated with a selective COX-2 antagonist (NS-398) or physiologic saline (NS). The COX-2 inhibitor significantly improved postoperative ileus after standardized laparotomy. GC, geometric center of radiolabel distribution. *P , 0.05 versus NS.

[32]. Their group showed that selective ablation of afferent sensory neurons from the gut following neonatal administration of the selective sensory neurotoxin, capsaicin, subsequently led to markedly improved postoperative small bowel transit [32]. More recently, we confirmed that local application of capsaicin to the celiac/SMA ganglion complex, selectively destroying sensory afferent fibers originating from the foregut, also improved postoperative small bowel transit [33]. Here we have shown that selective inhibition of COX-2 ameliorates postoperative ileus in a rat model. This is likely due to the inhibition of PGI 2 synthesis, although the production of other prostanoids, such as PGE 2, is inhibited with NS-398 as well. Eicosanoids, in particular PGI 2 and PGE 2, exert a hyperalgesic effect by decreasing the threshold of afferent nerve fibers to painful stimuli by noxious agents, including histamine and bradykinin [34]. Stimulation of these pain fibers diminishes small intestinal motility through adrenergic reflex pathways [35]. Interruption of the afferent and efferent limbs of this pathway by capsaicin and 6-hydroxydopamine or splanchnicectomy ameliorates postoperative ileus [32, 35, 36]. This effect may be attenuated by the inhibition of prostanoid synthesis with nonsteroidal anti-inflammatory drugs [31, 37] or, as we have demonstrated, with a selective COX-2 inhibitor. We were intrigued to find significant improvement of ileus at 30 min postoperatively, since our selective COX-2 inhibitor, NS-398 should inhibit only the inducible form of COX. Synthesis of mRNA for COX-2 should take a minimum of 2 h, and thus we expected to see minimal improvement in early postoperative small bowel transit. This improvement in early postoperative transit can be explained by the fact that COX-2 mRNA is expressed basally in human and, presumably, rat

JOSEPHS ET AL.: ROLE OF COX-2 IN POSTOPERATIVE ILEUS

small intestine [38]. Blockade of this fraction of COX appears to reduce production of prostanoids enough to ameliorate postoperative ileus. Constitutively expressed COX-1-induced prostaglandin synthesis would be uninhibited by NS-398. This may explain why, in a previous study, pretreatment with the nonselective COX inhibitor ketorolac returned transit at 30 min postoperatively to control, nonoperated values [18], while in the present study the effect of the selective COX-2 inhibitor NS-398 was more modest. The advantage of the latter, however, is that its beneficial effect on postoperative ileus should not be accompanied by the undesirable effects of nonselective COX inhibition, such as an anti-platelet effect and a deleterious effect on the gastric mucosa. We were surprised that neither the TNF-bp nor the IL-1ra improved postoperative small bowel transit in our model. Exogenous IL-1 and TNF have been shown to induce gastroparesis in rodents in a dose-dependent fashion [39, 40], and Coimbra and Plourde demonstrated that IL-1ra improved postoperative gastric emptying [19]. The reason for the lack of effect of the IL-1ra on postoperative small bowel ileus in the present study is unknown. Coimbra and Plourde demonstrated that IL-1 acted via the release of calcitonin gene-related peptide (CGRP). Our laboratory previously demonstrated that a CGRP receptor antagonist did not improve postoperative small bowel transit [33], despite the study by Plourde et al. [41] that documented improvement in postoperative gastric emptying with a CGRP receptor antagonist. We attribute this disparity to the differing innervation of the stomach and small bowel [33]. In summary, this study demonstrates that preoperative administration of a selective COX-2 inhibitor significantly improves postoperative ileus. Although the mechanism of this effect is unknown, it likely involves inhibition of production of PGI 2 and PGE 2, with prevention of their direct inhibitory effect on intestinal smooth muscle, as well as reduction of visceral afferent nerve fiber stimulation and secondary activation of sympathetic efferent fibers. More insight into the mechanism of postoperative ileus is necessary to develop new therapeutic approaches to change management from a supportive role to a curative or preventative one.

5.

6. 7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

REFERENCES 1.

Douglas, D. M., and Mann, F. C. The effect of peritoneal irritation on the activity of the intestine. Br. Med. J. 1: 227, 1941. 2. Markowitz, J., and Campbell, W. C. The relief of experimental ileus by spinal anesthesia. Am. J. Physiol. 8: 101, 1927. 3. Oschsner, A. Postoperative treatment based on physiologic principles. South. Surg. 4: 197, 1935. 4. Neely, J., and Catchpole, B. Ileus: The restoration of alimentary-tract motility by pharmacological means. Br. J. Surg. 58: 21, 1971.

19.

20.

21.

53

Moss, G., Regal, M. E., and Lichtig, L. L. Reducing postoperative pain, narcotics and length of hospitalization. Surgery 99: 206, 1986. Livingston, E. H., 2nd, and Passaro, E. P., Jr. Postoperative ileus. Dig. Dis. Sci. 35: 121, 1990. Cheape, J. D., Wexner, S. D., James, K., and Jagelman, D. G. Does metaclopramide reduce the length of ileus after colorectal surgery? A prospective randomized trial. Dis. Colon Rectum 34: 437, 1991. Bonacini, M., Quiason, S., Gaddis, M., Reynolds, M., Pemberton, L. D., and Smith, O. J. Therapy of postoperative ileus with intravenous erythromycin: Preliminary results of a double blind placebo-controlled study. Gastroenterology 100: A423, 1991. Benson, M. J., Roberts, J. P., Wingate, D. L., Rogers, J., Deeks, J. J., Castillo, F. D., and Williams, N. S. Small bowel motility following major intra-abdominal surgery: The effect of opiates and rectal cisapride. Gastroenterology 106: 924, 1994. Forstermann, U., and Neufang, B. The role of the kidney in the metabolism of prostacyclin by the 15-hydroxyprostaglandin dehydrogenase pathway in vivo. Biochem. Biophys. Acta 793: 338, 1984. Wyble, C. W., Desai, T. R., Clark, E. T., Hynes, K. L., and Gewertz, B. L. Physiologic concentrations of TNFa and IL-1b released from reperfused human intestine upregulate E-selectin and ICAM-1. J. Surg. Res. 63: 333, 1996. Reimund, J. M., Wittersheim, C., Dumont, S., Muller, C. D., Kenney, J. S., Baumann, R., Poindron, P., and Duclos, B. Increased production of tumour necrosis factor-alpha, interleukin-1 beta, and interleukin-6 by morphologically normal intestinal biopsies from patients with Crohn’s disease. Gut 39: 684, 1996. Fukumura, D., Miura, S., Kurose, I., Higuchi, H., Suzuki, H., Ebinuma, H., Han, J. Y., Watanabe, N., Wakabayashi, G., Kitajima, M., and Ishii, H. IL-1 is an important mediator for microcirculatory changes in endotoxin-induced intestinal mucosal damage. Dig. Dis. Sci. 41: 2482, 1996. Angel, J., Berenbaum, F., Le Denmat, C., Nevalainen, T., Masliah, J., and Fournier, C. Interleukin-1-induced prostaglandin E2 biosynthesis in human synovial cells involves the activation of cytosolic phospholipase A2 and cyclooxygenase-2. Eur. J. Biochem. 226: 125, 1994. Ristimaki, A., Garfinkel, S., Wessendorf, J., Maciag, T., and Hla, T. Induction of cyclooxygenase-2 by interleukin-1 alpha. Evidence for post-transcriptional regulation. J. Biol. Chem. 269: 11769, 1994. Pilbeam, C. C., Kawaguchi, H., Hakeda, Y., Voznesensky, O., Alander, C. B., and Raisz, L. G. Differential regulation of inducible and constitutive prostaglandin endoperoxide synthase in osteoblastic MC3T3-E1 cells. J. Biol. Chem. 268: 25643, 1993. Kelley, M. C., Hocking, M. P., Marchand, S. T., and Sninsky, C. A. Ketorolac prevents postoperative small intestinal ileus in rats. Am. J. Surg. 165: 107, 1993. Cheng, G., Cassissi, C., Drexler, P. G., Vogel, S. B., Sninsky, C. A., and Hocking, M. P. Salsalate, morphine, and postoperative ileus. Am. J. Surg. 171: 85, 1996. Coimbra, C. R., and Plourde, V. Abdominal surgery-induced inhibition of gastric emptying is mediated in part by interleukin-1 beta. Am. J. Physiol. 270: R556, 1996. Josephs, M., Ksontini, R., Solorzano, C., Moshyedi, A., Abdalla, E., Colagiovanni, D., Edwards, C., Tannahill, C., MacKay, S., Copeland, E., and Moldawer, L. Endotoxin and D-galactosamine-induced hepatic injury is mediated by TNFa and not by Fas ligand. J. Immunol., in press. Russell, D. A., Tucker, K. K., Chinookoswong, N., Thompson,

54

22.

23.

24.

25.

26.

27.

28. 29.

30.

31.

JOURNAL OF SURGICAL RESEARCH: VOL. 86, NO. 1, SEPTEMBER 1999 R. C., and Kohno, T. Combined inhibition of interleukin-1 and tumor necrosis factor in rodent endotoxemia: Improved survival and organ function. J. Infect. Dis. 171: 1528, 1995. Espat, N. J., Cheng, G., Kelly, M. C., Vogel, S. B., Sninsky, C. A., and Hocking, M. P. Vasoactive intestinal peptide and substance P receptor antagonists improve postoperative ileus. J. Surg. Res. 58: 719, 1995. Miller, M. S., Galligan, J. J., and Burks, T. F. Accurate measurement of intestinal transit in the rat. J. Pharmacol. Methods 6: 211, 1981. Sninsky, C. A. Vincristine alters myoelectric activity and transit of the small intestine in rats. Gastroenterology 92: 472, 1987. Cannon, W. B., and Murphy, F. T. The movements of the stomach and intestines in some surgical conditions. Ann. Surg. 43: 512, 1906. Bernstein, M. A., Dawson, J. W., Reissman, P., Weiss, E. G., Nogueras, J. J., and Wexner, S. D. Is complete laparoscopic colectomy superior to laparoscopic assisted colectomy? Am. Surg. 62: 507, 1996. Carlson, M. A., and Frantzides, C. T. Canine intestinal myoelectric activity after open versus laparoscopically assisted right hemicolectomy. Am. J. Surg. 174: 79, 1997. Oschner, A., Gage, I. M., and Cutting, R. A. The value of drugs in the relief of ileus. Arch. Surg. 21: 924, 1930. Ruwart, M. J., and Rush, B. D. Prostacyclin inhibits gastric emptying and small-intestinal transit in rats and dogs. Gastroenterology 87: 392, 1984. Thor, P., Konturek, J. W., Konturek, S. J., and Anderson, J. H. Role of prostaglandins in control of intestinal motility. Am. J. Physiol. 248: G353, 1985. Doherty, N. S., Beaver, T. H., Chan, K. Y., Coutant, J. E., and Westrich, G. L. The role of prostaglandins in the nociceptive

32.

33.

34.

35. 36.

37.

38.

39.

40.

41.

response induced by intraperitoneal zymosan in mice. Br. J. Pharmacol. 91: 39, 1987. Holzer, P., Lippe, I. T., and Holzer-Petsche, U. Inhibition of gastrointestinal transit due to surgical trauma or peritoneal irritation is reduced in capsaicin-treated rats. Gastroenterology 91: 360, 1986. Freeman, M. E., Cheng, G., and Hocking, M. P. The role of afferent neurons and CGRP in postoperative small bowel ileus. Surg. Forum 47: 188, 1996. Ferreira, S. H., Nakamura, M., and Salete de Abreu Castro, M. The hyperalgesic effects of prostacyclin and prostaglandin E 2. Prostaglandins 16: 31, 1978. Furness, J. B., and Costa, M. Adynamic ileus, its pathogenesis and treatment. Med. Biol. 52: 82, 1974. Dubois, A., Weise, V. K., and Kopin, J. J. Postoperative ileus in the rat: Pathophysiology, etiology, and treatment. Ann. Surg. 178: 781, 1973. Berkenkopf, J. W., and Weichman, B. W. Production of prostacyclin in mice following intraperitoneal injection of acetic acid, phenylbenzoquinone and zymosan: Its role in the writhing response. Prostaglandins 36: 393, 1988. O’Neill, G. P., and Ford-Hutchinson, A. W. Expression of mRNA for cyclooxygenase-1 and cyclooxygenase-2 in human tissues. FEBS Lett. 330: 156, 1993. McCarthy, D. O., and Daun, J. M. The role of prostaglandins in interleukin-1 induced gastroparesis. Physiol. Behav. 52: 351, 1992. Montuschi, P., Preziosi, P., and Navarra, P. Interleukin-1a and tumour necrosis factor inhibit rat gastric fundus motility in vitro. Eur. J. Pharmacol. 234: 303, 1993. Plourde, V., Wong, H. C., Walsh, J. H., and Tache, Y. CGRP antagonists and capsaicin on celiac ganglia partly prevent postoperative gastric ileus. Peptides 14: 1225, 1993.