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Helminthic parasites inhibit spontaneous colitis in IL-10 deficient mice
AGAAB63
April 2000
nuclear cells decreased slightly (mean 3.65 to 3.20 p'" 0.011. While fasting gastric pH had no predictive value regarding mucosal damage, there was an inverse correlation between endoscopic score and the pH of the gastric juice post-therapy (R '" 68, p=0.22). Conclusions: None of the pre-NSAID parameters studied related to acute inflammation changed or were associated with the severity of NSAID-induced gastric mucosal damage. The prior association of PMNs with NSAID ulcer was likely because PMNs were surrogate for an Hp infection increased risk of NSAID ulcers.
4801 APPENDECTOMY SUPPRESSES INTESTINAL INFLAMMATION IN A MURINE MODEL OF DSS-INDUCED COLITIS THROUGH MODULATION OF MUCOSAL IMMUNE SYSTEMS. Nobuo Tomiyasu, Keiichi Mitsuyama, Satoshi Matsumoto, Nami Watanabe, Tsutomu Nishiyama, Asuka Suzuki, Kosuke Takagi, Atsushi Toyonaga, Yoshinori Umesaki, Michio Sara, Kurume Univ Sch of Medicine, Kurume, Japan; Yakult Cent Institute, Kunitachi, Japan. Background: The low prevalence of appendectomy in ulcerative colitis and the prevention of colitis in T cell receptoro-rnutant mice by appendectomy support the hypothesis that appendectomy is a potential protective factor for the inflammatory response of the colon. However, the mechanism remains unknown. To elucidate this issue, we removed the appendix, and examined its effect on colonic inflammation in dextran sulfate sodium (DSS)-treated mice [Expt.1] and on mucosal immune systems in normal mice [Expt.2]. Methods: [Expt.1] Female Balb/c mice were appendectomized or sham-operated at different ages (2 weeks and 6 weeks), and colitis was induced by feeding 4% DSS in drinking water at 10 weeks of age (n> 18 in each group). An acute colitis model was induced by a 7 day DSS and killed at II weeks, and a chronic model was induced by 4 cycles of treatment (a 7 day DSS and a 10 day interval) and killed at 19 weeks. The disease severity was assessed by the established clinical score (0-12) and pathological score (0-12). [Expt.2] Mice were appendectomized or shamoperated at different ages (2 weeks and 6 weeks), and killed at 10 weeks of age without colitis induction (n> 12 in each group). Cecal contents were cultured for bacterial analysis. Cytokine secretion from cultured lamina propria lymphocytes (LPLs) was determined by ELISA. The proliferative response of LPLs towards bacterial component was determined by 3H_ thymidine incorporation. Results: [Expt.1] Appendectomy at 2 weeks resulted in a mild improvement of disease severity in the acute model (clinical score, NS; pathological score, p<.05 vs sham-op) and a marked improvement in the chronic model (clinical score, p<'OO5; pathological score, p<'OOI), while appendectomy at 6 weeks did not. [Expt.2] Appendectomy at 2 weeks, but not at 6 weeks, significantly decreased IFN')!Secretion (p<.05 vs sham-op) and increased IL-4 secretion (p<.05) from cultured LPLs with an elimination of Bacteroides species from enteric microflora (p<.05). The proliferation rate by LPLs towards self-bacterial component was significantly reduced in appendectomized mice compared to sham-operated mice (p<.05). Conclusions: These findings suggest for the first time that appendectomy may alter bacterial flora and modulate mucosal immune systems, and thereby suppress the development of chemically induced colitis.
4802 HELMINTIDC PARASITES INHIBIT SPONTANEOUS COLITIS IN IL-I0 DEFICffiNT MICE. David E. Elliott, Cathy Crawford, Jie Li, Arthur Blum, Ahmed Metwali, Khurram Qadir, Joseph F. Urban, Jr., Joel V. Weinstock, Univ of Iowa, Iowa City, IA; USDA, Beltsville, MD. Populations at the highest risk for inflammatory bowel disease (lBD) have the lowest prevalence of helminthic parasite exposure. IBD is rare where helminthic colonization is common. Crohn's disease results from excessive intestinal inflammation with dysregulated Thl (IFN')', IL-12, TNFa) cytokine production. Helminths elicit strong Th2 (lL-4, IL-13, IL-5, IL-IO) responses. Strong Th2 responses impede Th I reactions. Thus, we hypothesize that exposure to helminthic parasites protects individuals from developing Crohn' s disease. IL-IO is a critical immunoregulatory cytokine that inhibits macrophage accessory cell function needed by Thl cells. Mice lacking IL-IO develop spontaneous colitis by 2-3 months of age. The colitis, triggered by intestinal bacteria, is transmural, progressive, and fatal. Colitis in IL-IO deficient mice is attenuated by anti-Il-Ny or anti-IL-12 antibody treatment showing that the inflammation results from a dysregulated Th1 response. To test the hypothesis that helminthic parasites prevent Thl-mediated colitis, we treated IL-IO deficient (C57BLl6_ILlO tm I Cg n ) mice with the small intestinal parasitic worm Heligmosomoides polygyrus. Eight week old mice were inoculated per os with 3rd stage larvae. Control IL-IO deficient mice received gavage lacking parasites. Thirty five days after inoculation the mice had persistent colonization with mature worms in the jejunum. At this time, colons were removed, fixed, and sectioned for histologic examination. The sections were coded and examined by two investigators blinded to the treatment group. Each group contained at least 5 animals and the experiment was repeated three times. Inflammation was graded on a four point scale; O=no inflammation, I =Iow level, 2=intermediate level, 3=high level inflammation with wall thickening, and 4=transmural infiltration. Sham-exposed IL-IO -1- mice spontaneously develop severe colitis (average score 3.67 ::t:: 0.17). Mice that had been exposed to H. polygyrus had significantly less intestinal inflammation (2.2 ::t:: 0.44, p
Conclusion: Colonization with parasitic helminths inhibits colitis in IL-IO deficient mice. Helminths can inhibit spontaneous colitis that results from dysregulated Th I responses. Helminthic parasites can provide protection even in the absence of the critical immunoregulatory cytokine IL-IO. Support: CCFA, DK38327, DK02428, DK25295, VAMC
4803 BLOCKADE OF IL·6 TRANSSIGNALING ABROGATES ESTABLISHED EXPERIMENTAL COLITIS IN MICE BY SUPPRESSION OF T CELL RESISTANCE AGAINST APOPTOSIS. Raja Atreya, Susetta Finotto, Jonas Mudter, Jochen Mullberg, Thomas Jostock, Martin Holtmann, Tasaku Kishimoto, Peter R. Galle, Stefan Rose-John, Markus F. Neurath, I Med Clin Univ, Mainz, Germany; Osaka Univ, Osaka, Japan. T cell resistance against apoptosis appears to contribute to disease perpetuation and the requirement for long-term immunosuppressive therapy in Crohn's disease. Here, we show that a neutralizing IL-6 receptor (IL-6R) antibody causes rapid suppression of established experimental colitis in various THI-mediated animal models of Crohn's disease (IL-IO knockout mice, SCID transfer model, TNBS-induced colitis) by inducing apoptosis of lamina propria T cells. Similarly, specific neutralization of the soluble IL-6 receptor (sIL-6R) in vivo by a newly designed gpl30 Fc fusion protein caused suppression of colitis activity and induction of apoptosis suggesting a key role for the sIL-6R in preventing mucosal T cell apoptosis. In patients with Crohn's disease, we observed that IL-6 is a predominant cytokine produced by lamina propria T cells. These cells showed strong evidence for IL-6 transsignaling associated with activation of STAT-3, bcl-2 and bcl-xl. Finally, blockade oflL-6 transsignaling caused T cell apoptosis providing direct evidence for the involvment of IL-61 sIL-6R in the resistance of lamina propria T cells against apoptosis in Crohn's disease. Taken together, these data suggest that an IL-6/sIL-6R driven pathway of T cell activation strongly contributes to the perpetuation of chronic intestinal inflammation. Specific targeting of this pathway appears to be a promising novel approach for the treatment of patients with Crohn's disease.
4804 A NOVEL, ORALLY-ACTIVE INIDBITOR OF ICAM·l AND ESELECTIN IS EFFICACIOUS IN THE TREATMENT OF COLITIS IN RHESUS MACAQUES. Cynthia H. Jurgensen, Susan M. Daluge, Jeffrey L. Selph, Steven S. Good, Harry Marr, Marie A. Iannone, Brian E. Huber, Allen W. Mangel, Nicholas W. Lerche, Kay Washington, Gerald Wolberg, Glaxo Wellcome Research & Development, RTP, NC; UC Davis Primate Ctr, Davis, CA; Vanderbilt Univ Sch of Medicine, Nashville, TN. Leukocyte adhesion to the vascular endothelium is an essential step in inflammatory processes. This adhesion is dependent on the induction or upregulation of adhesion molecules such as ICAM-I and E-selectin on the endothelium, and represents an important target for inflammatory disorders. The adhesion molecules ICAM-I and E-se1ectinare expressed at sites of active inflammation in patients with inflammatory bowel disease (IBD). Therefore, a small molecule inhibitor of ICAM-I and E-selectin expression could be promising in the treatment of IBD. GI270384X is a novel inhibitor of cytokine-induced adhesiveness of the endothelium for leukocytes, with an ICso=30 nM. This compound acts in vitro through inhibition of the induction of ICAM-I and E-selectin expression by TNF, IL-I, or LPS on human umbilical vein endothelial cells, without altering levels of constitutively-expressed adhesion molecules. The inhibitory activity of GI270384X targets a late event in the induction pathway, interfering with a post-transcriptional process. In studies with orally-dosed compound (25 mglkg/day)in 5 animal species, no systemic exposure to parent compound and minimal exposure to metabolites was observed. The majority of the compound was excreted in the feces. GI270384X was evaluated in rhesus macaques with spontaneous colitis, a condition sharing many of the features of human IBD. Significant improvement was observed in the compound-treated animals (12.5 mg/kg bid for 14 days) versus those receiving placebo. Response to treatment was measured as resolution of diarrhea and increased food intake, corresponding with overall improvement in the histological assessment of inflammation in tissues from colon biopsies. In addition, immunohistochemical analysis of these tissue specimens demonstrated decreased ICAM-I and E-selectin expression in the vasculature of responding animals. Compound disposition in the rhesus monkey was characterized using [14C]-labeled GI270384X, and results showed no increased exposure to radiolabeled species in diseased animals. These findings indicate that G1270384X is a potentially promising new treatment for IBD, representing a novel, orally-active approach to topical treatment of the gastrointestinal tract.
April 2000
nuclear cells decreased slightly (mean 3.65 to 3.20 p'" 0.011. While fasting gastric pH had no predictive value regarding mucosal damage, there was an inverse correlation between endoscopic score and the pH of the gastric juice post-therapy (R '" 68, p=0.22). Conclusions: None of the pre-NSAID parameters studied related to acute inflammation changed or were associated with the severity of NSAID-induced gastric mucosal damage. The prior association of PMNs with NSAID ulcer was likely because PMNs were surrogate for an Hp infection increased risk of NSAID ulcers.
4801 APPENDECTOMY SUPPRESSES INTESTINAL INFLAMMATION IN A MURINE MODEL OF DSS-INDUCED COLITIS THROUGH MODULATION OF MUCOSAL IMMUNE SYSTEMS. Nobuo Tomiyasu, Keiichi Mitsuyama, Satoshi Matsumoto, Nami Watanabe, Tsutomu Nishiyama, Asuka Suzuki, Kosuke Takagi, Atsushi Toyonaga, Yoshinori Umesaki, Michio Sara, Kurume Univ Sch of Medicine, Kurume, Japan; Yakult Cent Institute, Kunitachi, Japan. Background: The low prevalence of appendectomy in ulcerative colitis and the prevention of colitis in T cell receptoro-rnutant mice by appendectomy support the hypothesis that appendectomy is a potential protective factor for the inflammatory response of the colon. However, the mechanism remains unknown. To elucidate this issue, we removed the appendix, and examined its effect on colonic inflammation in dextran sulfate sodium (DSS)-treated mice [Expt.1] and on mucosal immune systems in normal mice [Expt.2]. Methods: [Expt.1] Female Balb/c mice were appendectomized or sham-operated at different ages (2 weeks and 6 weeks), and colitis was induced by feeding 4% DSS in drinking water at 10 weeks of age (n> 18 in each group). An acute colitis model was induced by a 7 day DSS and killed at II weeks, and a chronic model was induced by 4 cycles of treatment (a 7 day DSS and a 10 day interval) and killed at 19 weeks. The disease severity was assessed by the established clinical score (0-12) and pathological score (0-12). [Expt.2] Mice were appendectomized or shamoperated at different ages (2 weeks and 6 weeks), and killed at 10 weeks of age without colitis induction (n> 12 in each group). Cecal contents were cultured for bacterial analysis. Cytokine secretion from cultured lamina propria lymphocytes (LPLs) was determined by ELISA. The proliferative response of LPLs towards bacterial component was determined by 3H_ thymidine incorporation. Results: [Expt.1] Appendectomy at 2 weeks resulted in a mild improvement of disease severity in the acute model (clinical score, NS; pathological score, p<.05 vs sham-op) and a marked improvement in the chronic model (clinical score, p<'OO5; pathological score, p<'OOI), while appendectomy at 6 weeks did not. [Expt.2] Appendectomy at 2 weeks, but not at 6 weeks, significantly decreased IFN')!Secretion (p<.05 vs sham-op) and increased IL-4 secretion (p<.05) from cultured LPLs with an elimination of Bacteroides species from enteric microflora (p<.05). The proliferation rate by LPLs towards self-bacterial component was significantly reduced in appendectomized mice compared to sham-operated mice (p<.05). Conclusions: These findings suggest for the first time that appendectomy may alter bacterial flora and modulate mucosal immune systems, and thereby suppress the development of chemically induced colitis.
4802 HELMINTIDC PARASITES INHIBIT SPONTANEOUS COLITIS IN IL-I0 DEFICffiNT MICE. David E. Elliott, Cathy Crawford, Jie Li, Arthur Blum, Ahmed Metwali, Khurram Qadir, Joseph F. Urban, Jr., Joel V. Weinstock, Univ of Iowa, Iowa City, IA; USDA, Beltsville, MD. Populations at the highest risk for inflammatory bowel disease (lBD) have the lowest prevalence of helminthic parasite exposure. IBD is rare where helminthic colonization is common. Crohn's disease results from excessive intestinal inflammation with dysregulated Thl (IFN')', IL-12, TNFa) cytokine production. Helminths elicit strong Th2 (lL-4, IL-13, IL-5, IL-IO) responses. Strong Th2 responses impede Th I reactions. Thus, we hypothesize that exposure to helminthic parasites protects individuals from developing Crohn' s disease. IL-IO is a critical immunoregulatory cytokine that inhibits macrophage accessory cell function needed by Thl cells. Mice lacking IL-IO develop spontaneous colitis by 2-3 months of age. The colitis, triggered by intestinal bacteria, is transmural, progressive, and fatal. Colitis in IL-IO deficient mice is attenuated by anti-Il-Ny or anti-IL-12 antibody treatment showing that the inflammation results from a dysregulated Th1 response. To test the hypothesis that helminthic parasites prevent Thl-mediated colitis, we treated IL-IO deficient (C57BLl6_ILlO tm I Cg n ) mice with the small intestinal parasitic worm Heligmosomoides polygyrus. Eight week old mice were inoculated per os with 3rd stage larvae. Control IL-IO deficient mice received gavage lacking parasites. Thirty five days after inoculation the mice had persistent colonization with mature worms in the jejunum. At this time, colons were removed, fixed, and sectioned for histologic examination. The sections were coded and examined by two investigators blinded to the treatment group. Each group contained at least 5 animals and the experiment was repeated three times. Inflammation was graded on a four point scale; O=no inflammation, I =Iow level, 2=intermediate level, 3=high level inflammation with wall thickening, and 4=transmural infiltration. Sham-exposed IL-IO -1- mice spontaneously develop severe colitis (average score 3.67 ::t:: 0.17). Mice that had been exposed to H. polygyrus had significantly less intestinal inflammation (2.2 ::t:: 0.44, p
Conclusion: Colonization with parasitic helminths inhibits colitis in IL-IO deficient mice. Helminths can inhibit spontaneous colitis that results from dysregulated Th I responses. Helminthic parasites can provide protection even in the absence of the critical immunoregulatory cytokine IL-IO. Support: CCFA, DK38327, DK02428, DK25295, VAMC
4803 BLOCKADE OF IL·6 TRANSSIGNALING ABROGATES ESTABLISHED EXPERIMENTAL COLITIS IN MICE BY SUPPRESSION OF T CELL RESISTANCE AGAINST APOPTOSIS. Raja Atreya, Susetta Finotto, Jonas Mudter, Jochen Mullberg, Thomas Jostock, Martin Holtmann, Tasaku Kishimoto, Peter R. Galle, Stefan Rose-John, Markus F. Neurath, I Med Clin Univ, Mainz, Germany; Osaka Univ, Osaka, Japan. T cell resistance against apoptosis appears to contribute to disease perpetuation and the requirement for long-term immunosuppressive therapy in Crohn's disease. Here, we show that a neutralizing IL-6 receptor (IL-6R) antibody causes rapid suppression of established experimental colitis in various THI-mediated animal models of Crohn's disease (IL-IO knockout mice, SCID transfer model, TNBS-induced colitis) by inducing apoptosis of lamina propria T cells. Similarly, specific neutralization of the soluble IL-6 receptor (sIL-6R) in vivo by a newly designed gpl30 Fc fusion protein caused suppression of colitis activity and induction of apoptosis suggesting a key role for the sIL-6R in preventing mucosal T cell apoptosis. In patients with Crohn's disease, we observed that IL-6 is a predominant cytokine produced by lamina propria T cells. These cells showed strong evidence for IL-6 transsignaling associated with activation of STAT-3, bcl-2 and bcl-xl. Finally, blockade oflL-6 transsignaling caused T cell apoptosis providing direct evidence for the involvment of IL-61 sIL-6R in the resistance of lamina propria T cells against apoptosis in Crohn's disease. Taken together, these data suggest that an IL-6/sIL-6R driven pathway of T cell activation strongly contributes to the perpetuation of chronic intestinal inflammation. Specific targeting of this pathway appears to be a promising novel approach for the treatment of patients with Crohn's disease.
4804 A NOVEL, ORALLY-ACTIVE INIDBITOR OF ICAM·l AND ESELECTIN IS EFFICACIOUS IN THE TREATMENT OF COLITIS IN RHESUS MACAQUES. Cynthia H. Jurgensen, Susan M. Daluge, Jeffrey L. Selph, Steven S. Good, Harry Marr, Marie A. Iannone, Brian E. Huber, Allen W. Mangel, Nicholas W. Lerche, Kay Washington, Gerald Wolberg, Glaxo Wellcome Research & Development, RTP, NC; UC Davis Primate Ctr, Davis, CA; Vanderbilt Univ Sch of Medicine, Nashville, TN. Leukocyte adhesion to the vascular endothelium is an essential step in inflammatory processes. This adhesion is dependent on the induction or upregulation of adhesion molecules such as ICAM-I and E-selectin on the endothelium, and represents an important target for inflammatory disorders. The adhesion molecules ICAM-I and E-se1ectinare expressed at sites of active inflammation in patients with inflammatory bowel disease (IBD). Therefore, a small molecule inhibitor of ICAM-I and E-selectin expression could be promising in the treatment of IBD. GI270384X is a novel inhibitor of cytokine-induced adhesiveness of the endothelium for leukocytes, with an ICso=30 nM. This compound acts in vitro through inhibition of the induction of ICAM-I and E-selectin expression by TNF, IL-I, or LPS on human umbilical vein endothelial cells, without altering levels of constitutively-expressed adhesion molecules. The inhibitory activity of GI270384X targets a late event in the induction pathway, interfering with a post-transcriptional process. In studies with orally-dosed compound (25 mglkg/day)in 5 animal species, no systemic exposure to parent compound and minimal exposure to metabolites was observed. The majority of the compound was excreted in the feces. GI270384X was evaluated in rhesus macaques with spontaneous colitis, a condition sharing many of the features of human IBD. Significant improvement was observed in the compound-treated animals (12.5 mg/kg bid for 14 days) versus those receiving placebo. Response to treatment was measured as resolution of diarrhea and increased food intake, corresponding with overall improvement in the histological assessment of inflammation in tissues from colon biopsies. In addition, immunohistochemical analysis of these tissue specimens demonstrated decreased ICAM-I and E-selectin expression in the vasculature of responding animals. Compound disposition in the rhesus monkey was characterized using [14C]-labeled GI270384X, and results showed no increased exposure to radiolabeled species in diseased animals. These findings indicate that G1270384X is a potentially promising new treatment for IBD, representing a novel, orally-active approach to topical treatment of the gastrointestinal tract.