Role of cycloexygenasse-1 and cyclooxygenase-2 in ischemia-reperfusion injury in the rat small intestine

administration of FITC-labeledalbumin (25 mg/kg, i.v.) in 5 experimental series: a) time control (CON, n =4), b) in the presenceof histamine as a measureof a maximal permeability response (HIS, 103M, n =5), c) after 20 min ET-1 superfuslon (1O-SM)(n=4), d) after 20 min of ET-1 superfusion in the presence of endothalin-A (ETA) receptor blockade with BQ123(10~M) (n =4), and e) after 20 rain of ET-1 supeffusion in the presenceof PAF inhibition with WEB-2086 (lmg/kg, Lv.) (n=4). ET-1 significantly increased albumin leakage above control levels but the amount of leakagewas only half of the maximal response observed with HIS (p<0.05). The increase in permeabilitywith ET-1 was abolished with both B0-123 (p
regulatorymucin genesequencesin normal and diseasestates will further the understandingof mucin function in epithelial protection.

1019 Th2 Responsesto the Helminth Hellgmosomoldespolygyrus Reduce Thl-promoted Epithelial Hyperplasia in a Mouse Model of Helicobacter hepaticus.aesesioted Inflammatory Bowel Disease (IBD) Mark T. Whary, MA Institute of Technology, Cambridge, MA; Hal Ning Shi, MA Gen Hosp, Boston, MA; Harold White, MA Institute of Technology, Cambridge, MA; Cathryn NeglerAnderson, MA Gen I-Iosp, Boston, MA; James G. Fox, MA Institute of Technology, Cambridge, MA BACKGROUND:Prior studies demonstratedthat Thl-promuted gastric atrophy secondaryto He/icobacterpy/ori infection in mice was ameliorated by the Th2 host responseto intestinal parasitism. Theseresults suggestedthat the low incidenceof gastric cancer in African populations which havea high incidenceof H. pyloriinfection may be attributablein part to parasitism. To determine if the Th2 response to parasites would ameliorate IBD in mice, the effect of coinfection with the murine helminth Heligmosomoidespolygyrus (Hpg) and Helicobactef hepaticus(HI?)on progression of diseasein the ILIO -~- mouse model of IBD was examined. METHODS: Forty wildtype C57BL/lOJ mice (WT) and 40 IL-IO-'- mice (C57BU10J background) were infected with Hpg or Hh alone or sequentially infected with Hpg then Hh for 16 weeks. RESULTS: Mice infected with Hpg developed higher total serum tgG1 and IoE levels and eosinophilia (p
1022 NeutrophiI-Assesioted, NADPH Oxidese-Oerived Superoxide Mediates LymphocyteEndothelial Cell Adhesion In Post-lschomic intestinal MIsrovascuiature. Takeharu Shigernatsu, LSU Health Science Ctr, Shreveport, LA; Chris R. Ross, Coil of Veterinary Medicine, Kansas Slate Univ, Manhattan,KS; Joe M. McCord, Univ of Colorado Health Science Ctr, Denver, CO; D Nell Granger, LSU Health Science Ctr, Shreveport, LA BACKGROUND/OBJECTIVES:While superoxide has been implicated as a major mediator of neutrophil-endothelialcell adhesion in intestinal venules exposedto ischemia-reperfusion (I/ R), it remains unclear whether this reactive oxygen specie also contributes to the recruitment of other leukocyte populations into postischemic intestine. The objectives of this study were to: 1) determinewhether superoxidemediatesI/R-inducedlymphocyteendothelialcell adhesion in mudne intestinal venules, and 2) define the potential role of neutrophil-associatedNAOPH oxJdaseas a source of this superoxide. METHODS:T-lymphocytes, isolated from spleens of C57BL]6 (wild type: WT) mice, were labelled with carboxy-fiuoresceindiacetatesuccinimidyl ester and injected intravenously into mice that were exposedto intestinal I/R. The superior mesentericartery was occludedfor 45min, followed by 6hr reperfusion. Intravital fluorescence microscopy was used to monitor the rolling and firm adherenceof T-cells in mucosal venules of the small intestine in control (WT, sham-operated),I/R-exposedWT or I/R-exposedNAOPH oxidase knock-out (phox47-/-) mice. Other mice were treated with either an extracellular superoxide dismutass (ecSOO) mimic (mutant form of Mn-SOD) or antineutrophil serum. RESULTS: Intestinal I/R resulted in significant lymphocyte-endothelialcell adhesionat 6 hrs of reperfusion in WT mice. This I/R-inducedlymphocyte recruitment responsewas significantly attenuatedin WT mice treated with the ecSOOmimic, and in phox47-/- mice. A significant, but less dramatic, attenuationof T-cell adhesionwas noted after treatment with antineutrophil serum. CONCLUSIONS:Thesefindings implicate superoxida as a mediator of T-lymphocyteendothelialcell adhesionin postisshemicintestinalvenules.The neutrophil-associatedenzyme, NADPH oxidase,may be a major source of the superoxidethat mediatesthis T-cell response. (Supported by HL26441)

1020 The Traffic And Role Of Circulating Dendritic Cell Precursors In A Granulomatoes Liver Disease: Dynamic Regulation By CC Chemokinas Tsuneo Aiba, Hiroyuki Yoneyama,Niigata Univ Sch of Medicine, Niigata Japan; Kenjiro Matsuno, Kumamoto Univ Sch of Medicine, Kumamoto Japan; Yan Yun Zhang, Tokyo Univ, Tokyo Japan; Maeako Mural, Kenji Suzuki, Makoto Naito, Hitoshi Asakura, Niigata Univ Sch of Medicine, Niigata Japan; Kouji Matsushima, Tokyo Univ, Tokyo Japan

lm

BACKGROUND:Dendritic cells (DCs) are the most powerful initiators of immunity. The trafficking of DCs is therefore pivotal in immune surveillance,and may be well regulatedby chemokines. However,the origin and the in vivo trafficking of DC precursors are still poorly understood. AIM: Identification of circulating OC precursors and investigation of their roles and needs for chemokines in hepatic inflammation. METHODS:A granulomatous liver disease was induced in C57BU6 mice by injection of heat-killed Propionibacteriumaches (P.acnes). From 1 hour to 28 days after infection, blood and liver CD11c+ OCs were sorted and tested their phenotypes, allo-stimulating capabilities, phagocytotic activities, chemokine receptor expression profiles, and migration potentials. For in vivo trafficking experiments,Ly5.2 micederived blood CD1lc + DC precursorswere adoptivelytransferred into P.acnes-primedcongeneic Ly5.1 mice. Frequencyand distribution of donor DCs were determined by double color immunohistochemicl staining. To definethe roles of chemokinesin DCtrafficking, neutralizing anti-macrophage inflammatory protein (MIP)-la antibody (Ab) and anti-secondary lymphoid organ chemokine (SLC) Ab were also injected just prior to cell transfer. Finally, to revealthe role of SLC in granuloma formation, we also investigated the effect of anti-SLC Ab on the granuloma-associatedliver injury. RESULTS:During infection, F4/80-B22O-CD11c+ DC precursors appeared in the circulation, and matured within newly formed granulomas. Recruited DCs later migrated to the portal area to interact with T cells in what we term "portal tract-associated lymphoid tissue (PALT)'. MIP-la attracted blood DC precursors to the sinusoidal granulomas, while SLC attracted mature DCs to the newly identified PALT. Anti-SLC Ab diminished PALTexpansion,while exacerbatinggranuloma formation. CONCLUSION: In emergency, distinct circulating DC precursors can migrate into a solid organ like liver, and participate in the granulomatous reaction in response to specific chemokines.

Role Of Cycloesygenase-1 And CyclooxYoenase-2 In Ischemia-Reperfusion Injury In The Rat Small intestine Nenad Martcic, KariheinzEhdich, Brigitta M. Peskar, Univ of Bochum, Bochum Germany In normal rats, small intestinal damage only develops when both cyclooxygenase(COX)-1 and COX-2 are inhibited, whereas selective inhibition of one COX isoform is not ulcerogenic (Gastroenterology 118: A826, 2000). We have previously shown that the non-selective COX inhibitor indomethacin and selectiveCOX-2 inhibitors markedly increase damageinduced by ischemia-reperfusion (I-R) in the rat stomach (Br J Pharmacol 128: 1659, 1999). This study investigates whether indomethacin, the COX-1 inhibitor SC-560 and the COX-2 inhibitors rofecoxib and NS-398 modify damage induced by I-R in the rat small intestine. METHODS: Fasted male Wistar rats, n=4-11 per group, were anaesthetizedwith pentobarbital. The abdomen was opened and the superior mesenteric artery clamped for 30 min followed by reperfusion for 60 min. Drugs were administered orally 60 rain before ischemia.As indication of damagethe small intestinal permeabilityfor Evans blue (15 mg/kg, injected iv 30 min after starting the reperfusion) was assessedby measuringdye concentrationsin the luminal contents of the ileum as described (J Clin Gastroenterol 12 Suppl 1: $158, 1990). RESULTS:The concentration of Evans blue in the small intestinal luminal contents of rats not subjected to I-R was 34-+3 pg/ml. Indomethacin (10 and 20 mg/kg) did not affect the permeability of Evans blue in rats without I-R but dose-dependentlyincreased luminal dye concentrations from 40-+2 in I-R alone to 129-+26 (p
1021 Endothelin-1 (ET-1) Induced Macromolecular Leakage in Submucoesl Misrovessels in Guinea Pig Small Intestine Peter Hugh MacDonald,Chris K. Chapler, Jeff O. Mewburn, Cheryl E. King-Vanvlack, Queen's Univ, Kingston Canada We previously found that ET-1 superfusion (lOeM) decreased blood flow in both proximal (1~/2nd order) and terminal (3~/4~ order) submucosal arterioles and venules in the small intestine of anesthetizedguinea pigs. The decreasein blood flow was associatedwith vasoconstriction in proximal but not in terminal microvessels,however,plateletactivating factor (PAF) inhibition eliminatedthe ET-1 induced reduction in blood flow in the terminal vessels without any effect on proximal vessels.Thesedata suggest that ET-1 stimulated PAF activation which in turn increased microvascular permeability; the accompanying hyperviscosity was most likely responsiblefor the reduction in blood flow. In the current study, we examinedthe effect of ET-1 on submucosal vascular permeability and the mechanisms by which these changes were induced in anesthetized guinea pigs. Microvascular permeability was determined by albumin leakageof fluorescein-isothiocyanate(FITC) labeledalbumin using fluorescence intravital videomicroscopy. Mean optical intensity of fluorescence was measured 5 min after

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