Andreas Raedler

Enhand secretion of tumour necrosis factor-alpha, IL-6, and IL-1β by isolated lamina ropria monouclear cells from patients with ulcretive cilitis and Crohn's disease

The perpetuation of inflammation in ulcerative colitis and Crohns disease may be regulated in part by an increased secretion of proinflammatory cytokines due to either an appropriate response to initial stimulating agents, and/or due to an impaired down‐regulalion of cylokine secretion. The aim of this study was to determine the secretion patterns of the proinflammatory cytokines tumour necrosis factor‐alpha (TNF‐α), IL‐6 and IL‐lβ, from isolated lamina propria mononuclear cells (LPMNC) isolated from colonic biopsies from patients with untreated ulcerative colitis or Crohns disease. LPMNC isoiated from involved inflammatory bowel disease (lBD) mucosa spontaneously produced increased amounts of TNF‐α, IL‐6, and IL‐lβ. The TNF‐α secretion from IBD LPMNC could be further enhanced by pokeweed mitogen stimulation. The secretion patterns of TNF‐α and IL‐1β by LPMNC from patients with either uicerative colitis or Crohns disease demonstrated a close correlation with the degree of tissue involvement and mucosal inflammation. LPMNC from noninvolved ulcerative colitis mucosa secreted markedly increased leveis of IL‐6 compared with noninvolved Crohns disease mucosa or control mucosa. The heightened lL‐6 secretion from LPMNC from non‐involved ulcerative colitis mucosa without visible or microscopic signs of inflammation indicates that the pathophysiologie mechanisms involved in the initiation of inflammation may differ between ulceralive colitis and Crohns disease. The determination of proinflammatory cytokine secretion by isolated LPMNC from colonoseopic biopsies may be a sensitive method for monitoring the severity of mucosal inflammation in IBD patients.

Immunoregulatory role of interleukin 10 in patients with inflammatory bowel disease

BACKGROUND/AIMS Active inflammatory bowel disease (IBD) is associated with increased proinflammatory cytokines. Deficiency of interleukin (IL) 10, a contrainflammatory cytokine, leads to the development of colitis in IL-10 knockout mice. We characterized IL-10 regulation of proinflammatory cytokine (tumor necrosis factor [TNF] alpha and IL-1 beta) expression in IBD in vitro and in vivo. METHODS IL-10 regulation of IL-1 beta, TNF-alpha, and IL-1 receptor antagonist expression by peripheral monocytes or isolated lamina propria mononuclear cells (LPMNC), respectively, was studied by enzyme-linked immunosorbent assay (cytokine secretion) and by semiquantitative reverse-transcription polymerase chain reaction. RESULTS IL-10 down-regulates IL-1 beta and TNF-alpha secretion as well as messenger RNA levels in IBD peripheral monocytes and LPMNC in a dose-dependent manner. In parallel, IL-1 receptor antagonist secretion is induced, and IL-10 can restore diminished in vitro IL-1 receptor antagonist/IL-1 beta ratios in IBD to normal levels. Equal concentrations of IL-10 are detectable in both normal and IBD intestinal lamina propria biopsy homogenates. After topical IL-10 enema treatment of three steroid therapy-refractory patients with ulcerative colitis, in vitro release of proinflammatory cytokines from IBD peripheral monocytes as well as LPMNC is dramatically down-regulated. CONCLUSIONS IL-10 down-regulates the enhanced secretion as well as messenger RNA levels of proinflammatory cytokines by IBD mononuclear phagocytes in vitro. In vivo topical application of IL-10 induces down-regulation of proinflammatory cytokine secretion both systemically and locally.

Twin Study Indicates Loss of Interaction Between Microbiota and Mucosa of Patients With Ulcerative Colitis

BACKGROUND & AIMS Interactions between genetic and environmental factors are believed to be involved in onset and initiation of inflammatory bowel disease. We analyzed the interaction between gastrointestinal mucosal microbiota and host genes in twin pairs discordant for ulcerative colitis (UC) to study the functional interaction between microbiota and mucosal epithelium. METHODS Biopsy were collected from sigmoid colon of UC patients and their healthy twins (discordant twin pairs) and from twins without UC. Microbiota profiles were determined from analysis of 16S ribosomal DNA libraries; messenger RNA profiles were determined by microarray analysis. RESULTS Patients with UC had dysbiotic microbiota, characterized by less bacterial diversity and more Actinobacteria and Proteobacteria than that of their healthy siblings; healthy siblings from discordant twins had more bacteria from the Lachnospiraceae and Ruminococcaceae families than twins who were both healthy. In twins who were both healthy, 34 mucosal transcripts correlated with bacterial genera, whereas only 25 and 11 correlated with bacteria genera in healthy individuals and their twins with UC, respectively. Transcripts related to oxidative and immune responses were differentially expressed between patients with UC and their healthy twins. CONCLUSIONS The transcriptional profile of the mucosa appears to interact with the colonic microbiota; this interaction appears to be lost in colon of patients with UC. Bacterial functions, such as butyrate production, might affect mucosal gene expression. Patients with UC had different gene expression profiles and lower levels of biodiversity than their healthy twins, as well as unusual aerobic bacteria. Patients with UC had lower percentages of potentially protective bacterial species than their healthy twins.

Tumour necrosis factor α and interleukin 1β in relapse of Crohn's disease

Summary Background Concentrations of proinflammatory cytokines are increased in the intestinal mucosa of patients with active Crohns disease. Experimental immunotherapeutic interventions with anticytokine agents in refractory Crohns disease show that tumour necrosis factor α (TNFα) may be an important mediator of inflammation. We investigated the relation between production of TNFα and interleukin 1β by mononuclear cells of the colonic lamina propria in patients with remitting Crohns disease and the risk of relapse. Methods We followed up 137 patients with Crohns disease in steroid-induced remisson for 1 year. Secretion of proinflammatory cytokines (tumour necrosis factor α [TNFα] and interleukin 1β) was assessed after short-term culture of human lamina propria mononuclear cells. Findings Increased secretion of TNFα and interleukin 1β were predictive for acute relapses within the next year. Site and extent of disease, baseline demographics, and serum acute-phase proteins had little predictive value. Interpretation TNFα is important as a target molecule for immune interventions in Crohns disease. The capacity to produce TNFα or interleukin 1β may identify patients who would benefit from anti-inflammatory remission maintenance.

Stress-Induced Gastrointestinal Secretory and Motor Responses in Rats Are Mediated by Endogenous Corticotropin-Releasing Factor

Corticotropin-releasing factor (CRF) has been implicated as a central nervous system mediator of stress. This study examined the effects of CRF and stress on gastric secretory and gastrointestinal motor functions in rats. Partial body restraint as a stress-producing stimulus significantly decreased gastric acid secretion, gastric emptying, and small bowel transit but markedly increased large bowel transit. Corticotropin-releasing factor given cerebroventricularly mimicked the gastrointestinal secretory and motor responses induced by partial body restraint. Cerebroventricular administration of a specific CRF receptor antagonist, alpha-helical CRF-(9-41), but not of the CRF fragment CRF-(1-20), prevented the gastrointestinal secretory and motor responses elicited either by partial body restraint or by exogenous administration of CRF in a dose-dependent fashion. These results suggest that the gastrointestinal secretory and motor responses in rats produced by stress (partial body restraint) are mediated by the endogenous release of CRF. They also indicate that CRF exerts its central nervous system actions on the gastrointestinal tract by a receptor-mediated event.

Autoradiographic study of early neurogenesis in rat neocortex.

SummaryThe early neurogenesis of rat neocortex was analysed by means of light and electron microscopic autoradiography. It was found that the very first preneurons originate probably as early as ED 11. They are the horizontal cells of Cajal-Retzius. The peak of their formation is on ED 13 (surface index estimated on ED 17 after injection of 3H-thymidine on ED 13:21, after injection on ED 12:4, after injection on ED 15:5), whereas no Cajal-Retzius cells could be found to have originated after ED 15. These cells are the developmentally most advanced of the neocortex. The cells second in date of origin and maturation are preneurons which presumably correspond to the presumptive neurons of Layer VII (VI b), and begin to originate on ED 12. The end of their formation could not be defined owing to a lack of ultrastructural differences to other, younger preneurons in later gestational stages. These two cell types are the first cellular components of the primordial plexiform layer (Marin-Padilla, 1978) or pallial anlage (Rickmann, 1977), demonstrating an outside-in gradient within this layer, and are separated by the formation of the cortical plate. This could be proven by their simultaneous labelling above and below the cortical plate after administration of 3H-thymidine before ED 15. These results confirm the hypothesis of a dual origin of the mammalian neocortex (Marin-Padilla, 1978).

Increased activation of isolated intestinal lamina propria mononuclear cells in inflammatory bowel disease

Normal human lamina propria lymphocytes are in a heightened state of activation compared with peripheral blood with regard to cell-surface activation antigen expression (transferrin receptor, interleukin-2 receptor, 4F2) and the increased spontaneous secretion of immunoglobulins in vitro. This study evaluates the cell-surface expression of activation-associated antigens in different subpopulations of isolated colonic lamina propria mononuclear cells in inflammatory bowel disease. In pilot studies using three-color flow cytometry, autofluorescence was observed that was emitted by unstained lamina propria mononuclear cells, which interfered with both the sensitivity and the specificity of the analyses. Because a major portion of the intestinal lymphocyte populations of interest were autofluorescent, a method to remove autofluorescence signals was developed by designing a computer program for the subtraction of autofluorescence from the emissions of each individual cell. This technique increases both the sensitivity and specificity of flow-cytometric analyses of intestinal lamina propria mononuclear cells. Using fluorescence-activated cell-sorter analyses with subtraction of autofluorescence on a single-cell basis, increased expression of lymphocyte activation antigens (interleukin-2 receptor, transferrin receptor, 4F2) was found on the cell surface of isolated intestinal B cells, T cells, CD4+ T cells, and CD8+ T cells in both Crohns disease and ulcerative colitis. Therefore, markedly increased intestinal lymphocyte activation is a major immunological alteration in inflammatory bowel disease and includes all lymphocyte subpopulations investigated in this study. In addition, 5-aminosalicylic acid, which is used for the treatment of intestinal inflammation in inflammatory bowel disease, inhibits the expression of cell-surface activation antigens on mitogen-activated peripheral blood lymphocytes in a dose-dependent manner. These observations suggest that lymphocyte activation may play an important role in underlying immune processes that lead to chronicity and perpetuation of inflammatory bowel disease and may implicate an additional mechanism for the therapeutic action of 5-aminosalicylic acid.

Genetic Analysis of Inflammatory Bowel Disease in a Large European Cohort Supports Linkage to Chromosomes 12 and 16

BACKGROUND & AIMS Inflammatory bowel disease (IBD) is a complex disorder of unknown etiology. Epidemiological investigations suggest a genetic basis for IBD. Recent genetic studies have identified several IBD linkages. The significance of these linkages will be determined by studies in large patient collections. The aim of this study was to replicate IBD linkages on chromosomes 12 and 16 in a large European cohort. METHODS Three hundred fifty-nine affected sibling pairs from 274 kindreds were genotyped using microsatellite markers spanning chromosomes 12 and 16. Affection status of the sibling pairs was defined as Crohns disease (CD) or ulcerative colitis (UC). RESULTS Nonparametric statistical analyses showed linkage for both chromosomes. Two-point results for chromosome 12 peaked at D12S303 (logarithm of odds [LOD], 2.15; P = 0.003) for CD and at D12S75 (LOD, 0.92; P = 0.03) for UC. Multipoint analyses produced a peak LOD of 1.8 for CD. Chromosome 16 showed linkage for CD at marker D16S415 (LOD, 1.52; P = 0.007). Multipoint support peaked above markers D16S409 and D16S411 (LOD, 1.7). CONCLUSIONS These data are consistent with linkage of IBD to chromosomes 12 and 16. The replication of genetic risk loci in a large independent family collection indicates important and common susceptibility genes in these regions and will facilitate identification of genes involved in IBD.

Recombinant Erythropoietin for the Treatment of Anemia in Inflammatory Bowel Disease

BACKGROUND Some patients with inflammatory bowel disease have anemia that is refractory to treatment with iron and vitamins. We examined whether administering iron and recombinant erythropoietin could raise hemoglobin levels in such patients. METHODS Thirty-four patients with inflammatory bowel disease (15 with ulcerative colitis and 19 with Crohns disease) and anemia refractory to iron therapy (hemoglobin concentrations below 10.0 g per deciliter [6.2 mmol per liter]) were randomly assigned in a prospective, double-blind, 12-week trial to receive either oral iron (100 mg per day) and subcutaneous erythropoietin (150 U per kilogram of body weight twice per week) (n=17) or oral iron and placebo (n=17). The primary measure of efficacy was an increase in hemoglobin levels of more than 1.0 g per deciliter (0.62 mmol per liter). Additional analyses were performed with other patients with inflammatory bowel disease. RESULTS The severity of anemia was related to clinical disease activity as well as to in vitro monocyte secretion of interleukin-1 beta, a proinflammatory cytokine. Serum erythropoietin concentrations were increased in 52 randomly selected outpatients with inflammatory bowel disease and anemia, but the concentrations were inadequate in relation to the degree of anemia. Twelve weeks of therapy with recombinant erythropoietin and oral iron increased mean (+/-SE) hemoglobin concentrations from 8.81+/-0.27 g per deciliter (5.47+/-0.17 micromol per liter) to 10.52+/-0.41 g per deciliter (6.5+/-0.25 micromol per liter), whereas hemoglobin concentrations in the placebo group decreased from 8.69+/-0.11 g per deciliter (5.4+/-0.068 micromol per liter) to 7.84+/- 0.33 g per deciliter (4.9+/-0.2 mmol per liter) (P<0.001). After 12 weeks, hemoglobin levels had increased by more than 1.0 g per deciliter in 82 percent of the patients in the erythropoietin group, as compared with 24 percent of those in the placebo group (P=0.002). There were five treatment failures in the placebo group and two in the erythropoietin group (P=0.18); treatment failure was defined as a decrease in hemoglobin levels of more than 2.0 g per deciliter (1.24 micromol per liter) to a value below 8.0 g per deciliter (4.96 micromol per liter) or any decrease to less than 6.5 g per deciliter (4.03 micromol per liter). CONCLUSIONS In patients with inflammatory bowel disease and anemia refractory to treatment with iron and vitamins, treatment with oral iron and recombinant erythropoietin can raise hemoglobin levels.

Epidemiology of inflammatory bowel disease in a German twin cohort: Results of a nationwide study

Background: Genetic predisposition as a cause of inflammatory bowel disease (IBD) has been proven by both family and twin studies and genetic variants associated with the disease have been identified. The aim of our study was to determine the concordance rates for IBD in German twin pairs and to evaluate clinical characteristics of concordant and discordant twin pairs. Methods: Patients with IBD were asked to participate and complete a questionnaire that contained questions about zygosity, demographic data, and medical history. Results: A total of 189 twin pairs in which at least 1 member had IBD were recruited (68 monozygotic and 121 dizygotic pairs). Within monozygotic pairs, 11 out of 31 (35%) were concordant for Crohns disease (CD) and 6 out of 37 (16%) for ulcerative colitis (UC). Two of the 58 (3%) dizygotic pairs with CD and 1 out of 63 (2%) dizygotic pairs with UC were concordant for the disease. In 14 out of 20 (70%) discordant monozygotic CD pairs and 25 out of 31 (81%) discordant monozygotic pairs with UC, the first‐born was affected by IBD. For discordant dizygotic twins, the first in birth order had IBD in 33 out of 56 (59%) pairs with CD and 40 out of 62 (64.5%) pairs with UC. Conclusions: This study confirms a stronger genetic influence in CD than in UC. The high preponderance in being affected of the first‐born twin and the fact that concordance was only 35% for CD and 16% for UC monozygotic twins highlight the important role of environmental trigger factors.