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Disturbed mRNA-expression of immunoregulatory cytokines in mucosa of patients with IBD
A884
AGA ABSTRACTS
GASTROENTEROLOGY, Vol. 1 0 8 , No. 4
LYMPHOCYTE HOMING TO THE GUT IN INFLAMMATORY BOWEL DISEASE (IBD) K Nakamuta, S Bloom, DP Jewell. Gastroenterology Unit, Radcliffe Infirmary, Oxford, UK Background: The role of homing receptors potentially relevant for the migration of naive and memory (CD45RA/RO) lymphocytes into the intestine in IBD has been investigated. Patients and Methods: Peripheral blood (PBL), lamina propria (LPL) and intraepithelial (IEL) lymphocytes were isolated from patients undergoing colonic resection: 12 ulcerative colitis (UC), 7 Crohn's disease (CD), and 10 carcinoma (control). Using two eolour FACS analysis, cells were characterised as naive (CD45 RA) or memory (CD45RO) and each sub-population anaiysed for expression of CD31, L-Selectin, c~4 integrin, LFA-1, and the integrin OqELI37. Data are shown as mean +/- SE. Results OtlELI37
(XlELI37
naive memory naive memory
Control IEL 63+/-8.0 71+/-7.0 Control LPL 14+/-4.0 17+/-3.0
UC IEL 9+/-3.0* 48+/-8.0 UC LPL 5+/-1.0 10+/-1.5
CD IEL 11+/-2.0" 70+/-6.0 CD LPL 12+/-8.5 14+/-5.0
No differences were seen between groups for PBL. The CD45RA/45RO ratio was similar for PBL, LPL and IEL in UC and CD compared with controls. However, there was a marked reduction in oriEL[37 expression (p<0.01*) in naive IEL in the disease groups. There were no differences in expression of LFA-1 or ct4 integrin on naive compared with memory cells, although LFA-1 generally reduced on IEL. CD31 was not increased.on ]EL or LPL CD45RO cells, suggesting that it is not involved in m*gration of lymphocytes across endothelium. Conclusions: Expression of CqELI37 is higher on antigen-primed intraepithelial lymphocytes in IBD than naive lymphocytes. This molecule may play a role in homing of memory cells to intestinal mucosa following antigenic re-challenge.
INFLUENCE OF H E ~ I C O B A C T E R PYLORI ON GASTRIC INTERLEUKIN-8 SYNTHESIS IN A DANISH AND AN ALBANIAN PATIENT POPULATION. O.H. Nielsen, S. Prifti, P. Peichl, I. Lindley, T. Horn. Depts. of Gastroenterology C & Pathology, Herlev Hospital, Univ. of Copenhagen, Denmark, Dept. of Gastroenterology, Tirana Univ. Hospital No. l, Albania, and Sandoz Research Inst., Vienna, Austria. There is growing evidence that Helicobacter pylori (H. pylQri) may initiate an inflammatory cascade. One important mediator for phagocyte chemotaxis and activation is interleukin-8 (IL-8). The aim was to study the association be ~ tween H. Dylori and IL-8, together with anti-IL-8-1gA antibodies, and f u r t h e r t o compare the results in two distinct patient populations. Totally 154 consecutive patients, who underwent an upper endoscopic examination were enrolled. In the Danish material (n=65) 28 were controls (C) (i.e. irritable bowel syndrome), 6 suffered from gastric ulcer disease (GU), 15 fro m duodenal ulcer (DU), 9 from gastritis (G), and 7 from duodenitis (D). In t h e A l b a n i a n material (n=89) 19 were C, 13 suffered from GU, 36 from DU, 12 from G, and 9 from D. IL-8 and anti-~L-8 IgA antibodies were analyzed in homogenized biopsy specimens by ELISA techniques. Significantly more Albanians, including C and dyspeptic patients (93.3%; 95% confidence limits 85.9-97.5%) were H. pvlori positive (histology and CLOtest) as c o m p a r e d t o the Danish subgroup (40.0%; 28.0-52.9%) (p<0.008). Three of 26 Danes (i1.5%) with a positive'CLOtest had IL-8 concentrations below the detection limit (DL) {18 pg/ml) as compared t o 2 9 of 39 (74.4%) with a negative CLotest (p<0.0O05). The IL-8 concentrations in gastric biopsies were significantly higher in C and D in the Albanian than in the Danish sub-populations (p<0.O01 and p
SALIVARY SECRETORY IMMUNOGLOBULIN A IN PEDIATRIC PATIENTS WITH CHRONIC IDIOPATHIC INFLAMMATORY BOWEL DISEASE. H. Nann, K. McGilligan, R. Reifen*, P. Sherman*, F. Sinatra, D. Thomas, Childrens Hospital Los Angeles, University of Southern California School of Medicine and *The Hospital for Sick Children, Toronto. Secretory immunoglobulinsplay an important role in mucosal defense by inhibition of bacterial adherence to mucosal surfaces, inducing antigenic shift in microorganisms, and controlling the permeability of the intestine to macromolecules. Secretory IgA (sIgA) deficiency could promote invasion of relatively non-pathogenic microorganisms. Absorption of some macromolecules caused by decreased sIgA levels could serve as sensitizing antigens. Patients with Crohn's disease (CD) and their healthy relatives have.been found to have increased intestinal permeability to macromolecular probes. Children with chronic enterocolitis associated with Hirschsprung's disease have been reported to have defective salivary IgA secretion. Salivary slgA has also been found to be decreased in a small number of adult patients with active inflammatory bowel disease (IBD). Other investigators reported elevated IgA levels in saliva of patients with ulcerative colitis (OC) and CD. Salivary slgA has not been measured in pediatric IBD patients. METHOD: Salivary stgA levels were measured by a modified radial immunodiffusion technique of Maneini on low level plates (Kent Labs), using slgA from human colostrum (ICN Biomedicals) as standards. Patients studied are summarized in the table: NUMBER AGE SEX Salivary slgA (years) (M/F) (mg/dl)(mean+SD) CD Patients 27 (10 active) 9-18.3 20/7 32.4 + 23,1 UC Patients 20 (8 active) 5-17 10/10 35.2 + 22.3 CD 1st Degree Family Members 36 6-46 9/27 32.8 + 20.8 UC 1st Degree Family Members 31 5-51 7/24 31.5 -I- 20.8 Healthy Controls 73 6-49 39/34 29.9 4- 19.6 RESULTS: No significant differences (analysis of variance) in salivary sIgA concentrations were found among the groups studied. CONCLUSIONS: These data do not suggest a primary defect in salivary sIgA synthesis or secretion in patients with IBD. The decreased and increased concentrations of salivary slgA previously reported in adults with IBD may represent a secondary phenomenon.
• DISTURBED mRNA-EXPRESSIONOF IMMUNOREGULATORYc~rrOKINES IN " MUCOSA OF PATIENTS WITH IBD. M.Niessner and B.A.Volk.
Department of Medicine, University of Freiburg, GERMANY. Introduction: Cytokines play a pivotal role in the regulation of the intestinal immune response and in the mediation of tissue damage in inflammatory bowel diease (IBD). Abnormalities in the expression of T-cell derived cytokines such as IL-2, IL-4, IL10 and IFN-y may indicate a dysregulation of intestinal immunity probably associated with pathogenic events: Therefore, cy~ok!ne mRNA expression was examined in the intestinal mucosa of patients with IBD and controls. Methods: The concentration of t h e immunoregulatory cytokin--'~wa's determined in biopsy specimens derived from patients with active (n=13) and inactive (n=12) ulcerative colitis (UC), active (n=l t) and inactive (n=11) Crohn's disease (CD) and control patients (n=14) using competitive RT-PCR with synthetic cytokine mRNAs. To assess a potential relation of cytokine mRNA levels within individual biopsy specimens regression analysis was performed. Results: Quantitation o f cytokine mRNAs in the mucosa revealed a significantly increased expression of IL,10 mRNA in patients with active UC (Median: 0.5pgi~g total RNA; p<0.01) and active CD (0.5pg/l~g; p<0.02) compared to normal mucosa (0.06pg/~g) and to inactive sites from the same patient (UC: 0.2pg/~g, CD: 0.1pg/pg). IFN-y mRNA levels were increased in active UC (0.25pg/pg) and CD (0.2pg/mT) compared t o normal mucosa (0.05pg/~g) and inactive disease (UC: 0.15pg/pg, CD: 0.05pg/~g). Furthermore IL-2 mRNA levels were significantly (p<0.01) increased in patients with active CD. The IL-4 mRNA expression did not differ between patients with IBD and controls. In normal tissue cytokine mRNA levels were Significantly correlated, whereas no such relation was observed in patients with active UC and CD. Conclusions: Our results implicate, that IL-10 plays an ~ole in the .regulation of the intestinal immune response and might act as a key element in the regulation of anti-inflammatory events. In addition, data obtained from regression analysis revealed, that there is a balanced cytokine production under normal conditions which is disturbed during inflammation.
AGA ABSTRACTS
GASTROENTEROLOGY, Vol. 1 0 8 , No. 4
LYMPHOCYTE HOMING TO THE GUT IN INFLAMMATORY BOWEL DISEASE (IBD) K Nakamuta, S Bloom, DP Jewell. Gastroenterology Unit, Radcliffe Infirmary, Oxford, UK Background: The role of homing receptors potentially relevant for the migration of naive and memory (CD45RA/RO) lymphocytes into the intestine in IBD has been investigated. Patients and Methods: Peripheral blood (PBL), lamina propria (LPL) and intraepithelial (IEL) lymphocytes were isolated from patients undergoing colonic resection: 12 ulcerative colitis (UC), 7 Crohn's disease (CD), and 10 carcinoma (control). Using two eolour FACS analysis, cells were characterised as naive (CD45 RA) or memory (CD45RO) and each sub-population anaiysed for expression of CD31, L-Selectin, c~4 integrin, LFA-1, and the integrin OqELI37. Data are shown as mean +/- SE. Results OtlELI37
(XlELI37
naive memory naive memory
Control IEL 63+/-8.0 71+/-7.0 Control LPL 14+/-4.0 17+/-3.0
UC IEL 9+/-3.0* 48+/-8.0 UC LPL 5+/-1.0 10+/-1.5
CD IEL 11+/-2.0" 70+/-6.0 CD LPL 12+/-8.5 14+/-5.0
No differences were seen between groups for PBL. The CD45RA/45RO ratio was similar for PBL, LPL and IEL in UC and CD compared with controls. However, there was a marked reduction in oriEL[37 expression (p<0.01*) in naive IEL in the disease groups. There were no differences in expression of LFA-1 or ct4 integrin on naive compared with memory cells, although LFA-1 generally reduced on IEL. CD31 was not increased.on ]EL or LPL CD45RO cells, suggesting that it is not involved in m*gration of lymphocytes across endothelium. Conclusions: Expression of CqELI37 is higher on antigen-primed intraepithelial lymphocytes in IBD than naive lymphocytes. This molecule may play a role in homing of memory cells to intestinal mucosa following antigenic re-challenge.
INFLUENCE OF H E ~ I C O B A C T E R PYLORI ON GASTRIC INTERLEUKIN-8 SYNTHESIS IN A DANISH AND AN ALBANIAN PATIENT POPULATION. O.H. Nielsen, S. Prifti, P. Peichl, I. Lindley, T. Horn. Depts. of Gastroenterology C & Pathology, Herlev Hospital, Univ. of Copenhagen, Denmark, Dept. of Gastroenterology, Tirana Univ. Hospital No. l, Albania, and Sandoz Research Inst., Vienna, Austria. There is growing evidence that Helicobacter pylori (H. pylQri) may initiate an inflammatory cascade. One important mediator for phagocyte chemotaxis and activation is interleukin-8 (IL-8). The aim was to study the association be ~ tween H. Dylori and IL-8, together with anti-IL-8-1gA antibodies, and f u r t h e r t o compare the results in two distinct patient populations. Totally 154 consecutive patients, who underwent an upper endoscopic examination were enrolled. In the Danish material (n=65) 28 were controls (C) (i.e. irritable bowel syndrome), 6 suffered from gastric ulcer disease (GU), 15 fro m duodenal ulcer (DU), 9 from gastritis (G), and 7 from duodenitis (D). In t h e A l b a n i a n material (n=89) 19 were C, 13 suffered from GU, 36 from DU, 12 from G, and 9 from D. IL-8 and anti-~L-8 IgA antibodies were analyzed in homogenized biopsy specimens by ELISA techniques. Significantly more Albanians, including C and dyspeptic patients (93.3%; 95% confidence limits 85.9-97.5%) were H. pvlori positive (histology and CLOtest) as c o m p a r e d t o the Danish subgroup (40.0%; 28.0-52.9%) (p<0.008). Three of 26 Danes (i1.5%) with a positive'CLOtest had IL-8 concentrations below the detection limit (DL) {18 pg/ml) as compared t o 2 9 of 39 (74.4%) with a negative CLotest (p<0.0O05). The IL-8 concentrations in gastric biopsies were significantly higher in C and D in the Albanian than in the Danish sub-populations (p<0.O01 and p
SALIVARY SECRETORY IMMUNOGLOBULIN A IN PEDIATRIC PATIENTS WITH CHRONIC IDIOPATHIC INFLAMMATORY BOWEL DISEASE. H. Nann, K. McGilligan, R. Reifen*, P. Sherman*, F. Sinatra, D. Thomas, Childrens Hospital Los Angeles, University of Southern California School of Medicine and *The Hospital for Sick Children, Toronto. Secretory immunoglobulinsplay an important role in mucosal defense by inhibition of bacterial adherence to mucosal surfaces, inducing antigenic shift in microorganisms, and controlling the permeability of the intestine to macromolecules. Secretory IgA (sIgA) deficiency could promote invasion of relatively non-pathogenic microorganisms. Absorption of some macromolecules caused by decreased sIgA levels could serve as sensitizing antigens. Patients with Crohn's disease (CD) and their healthy relatives have.been found to have increased intestinal permeability to macromolecular probes. Children with chronic enterocolitis associated with Hirschsprung's disease have been reported to have defective salivary IgA secretion. Salivary slgA has also been found to be decreased in a small number of adult patients with active inflammatory bowel disease (IBD). Other investigators reported elevated IgA levels in saliva of patients with ulcerative colitis (OC) and CD. Salivary slgA has not been measured in pediatric IBD patients. METHOD: Salivary stgA levels were measured by a modified radial immunodiffusion technique of Maneini on low level plates (Kent Labs), using slgA from human colostrum (ICN Biomedicals) as standards. Patients studied are summarized in the table: NUMBER AGE SEX Salivary slgA (years) (M/F) (mg/dl)(mean+SD) CD Patients 27 (10 active) 9-18.3 20/7 32.4 + 23,1 UC Patients 20 (8 active) 5-17 10/10 35.2 + 22.3 CD 1st Degree Family Members 36 6-46 9/27 32.8 + 20.8 UC 1st Degree Family Members 31 5-51 7/24 31.5 -I- 20.8 Healthy Controls 73 6-49 39/34 29.9 4- 19.6 RESULTS: No significant differences (analysis of variance) in salivary sIgA concentrations were found among the groups studied. CONCLUSIONS: These data do not suggest a primary defect in salivary sIgA synthesis or secretion in patients with IBD. The decreased and increased concentrations of salivary slgA previously reported in adults with IBD may represent a secondary phenomenon.
• DISTURBED mRNA-EXPRESSIONOF IMMUNOREGULATORYc~rrOKINES IN " MUCOSA OF PATIENTS WITH IBD. M.Niessner and B.A.Volk.
Department of Medicine, University of Freiburg, GERMANY. Introduction: Cytokines play a pivotal role in the regulation of the intestinal immune response and in the mediation of tissue damage in inflammatory bowel diease (IBD). Abnormalities in the expression of T-cell derived cytokines such as IL-2, IL-4, IL10 and IFN-y may indicate a dysregulation of intestinal immunity probably associated with pathogenic events: Therefore, cy~ok!ne mRNA expression was examined in the intestinal mucosa of patients with IBD and controls. Methods: The concentration of t h e immunoregulatory cytokin--'~wa's determined in biopsy specimens derived from patients with active (n=13) and inactive (n=12) ulcerative colitis (UC), active (n=l t) and inactive (n=11) Crohn's disease (CD) and control patients (n=14) using competitive RT-PCR with synthetic cytokine mRNAs. To assess a potential relation of cytokine mRNA levels within individual biopsy specimens regression analysis was performed. Results: Quantitation o f cytokine mRNAs in the mucosa revealed a significantly increased expression of IL,10 mRNA in patients with active UC (Median: 0.5pgi~g total RNA; p<0.01) and active CD (0.5pg/l~g; p<0.02) compared to normal mucosa (0.06pg/~g) and to inactive sites from the same patient (UC: 0.2pg/~g, CD: 0.1pg/pg). IFN-y mRNA levels were increased in active UC (0.25pg/pg) and CD (0.2pg/mT) compared t o normal mucosa (0.05pg/~g) and inactive disease (UC: 0.15pg/pg, CD: 0.05pg/~g). Furthermore IL-2 mRNA levels were significantly (p<0.01) increased in patients with active CD. The IL-4 mRNA expression did not differ between patients with IBD and controls. In normal tissue cytokine mRNA levels were Significantly correlated, whereas no such relation was observed in patients with active UC and CD. Conclusions: Our results implicate, that IL-10 plays an ~ole in the .regulation of the intestinal immune response and might act as a key element in the regulation of anti-inflammatory events. In addition, data obtained from regression analysis revealed, that there is a balanced cytokine production under normal conditions which is disturbed during inflammation.