April 1995 Immunology, Microbiology, and Inflammatory Disorders A789 RECENTLYRECRUITED MONOCYTE/MACROPHAGES AND NAIVET-CELLSCONTRIBUTETO SUBMUCOSAL PERIVASCULAR INFILTRATES INCROHN'SDISEASE(CD). V. L. ~Jrgio°, S. Fais', M. BoirivantL F. Pallonel" * Istituti di Clinica Medica 1 e *Clinica Medica 2, Univ. "La Sapienza", Roma; ~ Istituto Superirore di Sanit& Roma: "l' Dipartimanto di Medicina Sperimentaie, Uniw R. Calabria, Catanzaro, Italy, Cr~n's disease (CD) is Characterized by transmurai perivascular mononudser cell (MNC) infiltrates. Aim of the present study was to characterize the cellular components of these inflammatory infiltrates, To this purpose we analyzed the expression of both adhesion molecules and their ligands on endothelial cells (EC) and MNC, and characterized the subsets of MNC within the submucosal perivascular infiltrates. Cryostat sections of 10 surgical specimens were examined by immunohistochemistry using the APAAP complex method. CDfla +. mainly L-Selectin+ MNC appeared to be the major component of perivascular infiltrates. The vast majority of these cells were CD68", CD3t+ monocytes/macrophages (Me) surrounded by CD3+, L-Selectin+, CD31", CD45RA* and/or CD45RO+ T lymphocytes. The MNC observed within the dilated vessels lumen displayed the same surface phenotype. Double staining of T lymphocytes revealed that they mainly expressed both CD45RA and CD45RO markers. EC were ICAM-1 + and in great majority CD34*, Most important, strong adhesion between L-Selectin +. CDlla+ intravascular MNC and CD34+. ICAM-1+ EC were observed, These observation suggest that 1) peripheral MNC are activelly recruited in submucosa of CD tissue; 2) EC express adhesion molecules permissive for the recruitment of Me and both naive and memory T cells: 3) both naive and memory T cells are involved in trans endothelial migration and perivascular infiltrates generation; 4) dose membrane contact between migrated Me and naive T cells leads to the T cell transition from the naive to the memory phenotype within the CD submucosal inliltrates. CONSTITUTIVELY INCREASED TGF-$ mR.NA EXPRESSION IN THE MUCOSA OF PATIENTS WITH ULCERATIVE COLITIS. C. Butz, N. Endres, M. Classen, and K. Deusch, It Dept. of Medicine, Technical University, Munich, Germany. The role of cytokine dysregulation in the pathogenesis of inflammatory bowel disease (IBD) has often been discussed. In this study, we investigated the cytokine mRNA pattern corlstitutively expressed in the mucosa of patients with inflammatory bowel disea~. We analysed the cytokine mRNA expression in the colonic mucosa of patients with ulcerative colitis (UC) and Croim's disease (CD) and in normal mucosa. Additionally, we compared the content of cytokine mRNA in biopsies of affected and not affected colonic mucosa of patients with IBD. Colonic biopsies front patients undergoing colonoseopy were collected and the total RNA extracted. A semiquantilafive RT-PCR reaction was carried out using oliginucleotide primers s~c for IL-I, -2, -3, -4, -5, -6, -7, -8, - 10, TNF-c~ and -5, IL-1R, IL-IRA, IL-2R, IYN..y, TGF-B, GM-CSF, ECGF, CD3, CD4, and CDI4. The intensity of the PCR product was quantitied by laser densitomelry scanning of the bands in the agarose gel. For statistical analysis a Mann-WhitneyU test was performed. Stafisticab'y significant results were obtained for IL-3, TNF-o~ GM-CSF, ECGF, and TGF-B. No significant differences could be found for CD, neither between CD and controls nor between affected and not affected colonic mucosa. However, the UC specimens contained significantly more mR.NA for IL-3, TNF-~, GM-CSF, ECGF, and TGF-B. In the affected mucosa particularly TNF-og IL-3, and I"GF-B mRNA was increased, whereas in the not affected mucosa higher levels of GM-CSF, IL-3, and ECGF could be detected. When the affected mucosa was compared to not affected mucosa, the only statistically significant difference was the increased expression of TGF-B in the affected portion of the colon. The enhanced expression of TGF-B in the gut of patients with UC seems especially interesling, as TGF-B is known as a potent inhibitor of proliferation for many epithelial cells. On the other hand, it has also been shown, that TGF-B can augment the production of IL-6 by fibroblasts. Further studies have to elucidate the role of TGF-B in the pathogenesis of UC. O EFFECTS OF AN ULCERATIVE COLITIS NUTRITIONAL FORMULA (UCNF) ON CECAL AND COLONIC MUCOSAL PROSTAGLANDIN (PG) SYNTHESIS IN PIGS. J M Camobell. G C. Fshey, Jr., C.A. Lichtensteiger, K.A. Garleb, and S.J. DeMichele. Division of Nutritional Sciences, University of Illinois, Urbana, IL and Ross Products Division, Abbott Laboratories, Columbus, OH. Growing evidence supports a pathogenic role of arachidonic acid derived inflammatory mediators within the gastrointestinal tract of patients with ulcerative colitis. Dietary intake of n-3 fatty acids (FA) may directly influence the rate and pattern of colonic mucosal PG synthesis. The purpose of this study was to assess the effects of the UCNF containing indigestible oligosaccharides and fish oil on plasma phospholipid FA profiles, gut microbiota, and cecal and colonic mucosal PG profiles in pigs. Forty-eight growing barrows were equally randomized among four sacrifice times (day 0, 7, 14, and 21), two periods, and one of two diets: A control diet and the UCNF. Diets were isocaloric and contained comparable levels of protein, fat, and carbohydrate. Throughout the study, the diets were used to meet 100% of each pig's energy requirement. Pigs were allowed free access to water. Intake and body weight were recorded daily while fecal, blood, and tissue samples were collected at time of sacrifice. Prostaglandins were extracted from the cecal and colonic mucosa and plasma and measured via ELISA kits. Bifidobacteria were unaffected by treatment. Within one week of ingestion of the UCNF, the composition of plasma phospholipid FAs showed an increase in 20:5n3 and 22:6n3 (0.12% vs. 10.2%, P<.0001; 2.2% vs. 7.3%, P<.0001, respectively) and a decrease in 20:4n6 and 18:2n6 (9.8% vs. 6.5%, P<.0001; 22.4% vs. 5.4%, P<.0001, respectively). Plasma PGE2 and 6-keto-PGF~, levels were unaffected by treatment while thromboxane B2 (TXB2) was decreased (P<.O001) with ingestion of the UCNF. Prostaglandin E2, 6-keto-PGF]~, and TXB2 of the cecal mucosa (P<.0001) and colonic mucosa (P<.0001) were decreased due to ingestion of the UCNF. Ingestion of UCNF produced a suppression in the synthesis of proinflammatory PGs by cecal and colonic mucosai cells. These changes may have been mediated by rapid increases of n-3 FAs into cellular phopholipids. Dietary supplementation with UCNF may prove beneficial at modulating colonic PG synthesis in patients with ulcerative colitis. SAFETY AND TOLERANCE OF AN ULCERATIVE COLITIS NUTRITIONAL FORMULA (UCNF) IN HEALTHY ADULTS. I.M. CamDbelk G.C. Fahey, Jr., K.A. Garleb, and S.J. DeMichele. Division of Nutritional Sciences, University of Illinois, Urbana, IL and Ross Products Division, Abbott Laboratories, Columbus, OH. Recent studies have suggested that n-3 fatty acids from fish oil as well as short chain fatty acids produced during fermentation of indigestible oligosancharides may attentuate some of the gut injury and inflammation associated with ulcerative colitis. The purpose of this study was to assess the effects of a liquid nutritional formula containing indigestible oligosaccharides and gum arabic, as well as fish oil, on serum chemistries, hematology, plasma phospholipid fatty acid profiles, urinary constituents, and tolerance in healthy adults. Thirty adult males were randomly assigned to one of two treatments: A control diet (n=10) and the UCNF (n=20). Diets were isocaloric and contained comparable levels of protein, fat, and carbohydrate. For 14 days, the diets were used to meet 100% of each subject's energy requirement. Subjects were allowed free access to diet caffeine-free soda, tap water, and sugafless gum. Intake and tolerance were recorded daily while blood and urine samples were collected weekly on days 0, 7, and 14. No severe adverse reactions were reported. Certain subjects consuming large quantities of UCNF experienced some distention, gas, and loose stools. Sole source intake of UCNF was safe as evident by no untoward effects on serum chemistries, hematology, and urinalysis compared to the control formula. Serum triglycerides showed a week by treatment (P<.001) interaction of decreasing across time with the UCNF. The composition of plasma phospholipid fatty acids showed substantial decreases in 18:2n6 (21.0% to 7.7%; P<.05) and significant increases in 20:5n3 (0.33% to 10.3%; P<.05) and 22:6n3 (2.1% to 6.0%; P<.05) within one week of feeding UCNF. Dietary supplementation with UCNF may prove useful as adjunctive therapy in patients with ulcerative colitis.