Search

In about 70% of patients Crohn's disease (CD) affects the small intestine. This disease location is stable over time and associated with a genetic background different from isolated colonic disease. A characteristic feature of small intestinal host defense is the presence of Paneth cells at the bottom of the crypts of Lieberkühn. These cells produce different broad spectrum antimicrobial peptides (AMPs) most abundantly the α-defensins HD-5 and -6 (DEFA5 und DEFA6). In small intestinal Crohn's disease both these PC products are specifically reduced. As a functional consequence, ileal extracts from Crohn's disease patients are compromised in clearing bacteria and enteroadherent E. coli colonize the mucosa. Mechanisms for defective antimicrobial Paneth cell function are complex and include an association with a NOD2 loss of function mutation, a disturbance of the Wnt pathway transcription factor TCF7L2 (also known as TCF4), the autophagy factor ATG16L1, the endosomal stress protein XBP1, the toll-like receptor TLR9, the calcium mediated potassium channel KCNN4 as well as mutations or inactivation of HD5. Thus we conclude that small intestinal Crohn's disease is most likely a complex disease of the Paneth cell: Paneth's disease., (Copyright © 2010 European Crohn's and Colitis Organisation. Published by Elsevier B.V. All rights reserved.)

Inflammatory bowel diseases are characterized by chronic intestinal inflammation at different sites. Data from animal models as well as human patients including gene-association studies suggest that different components of the innate barrier function are primarily defective. These recent advances support the evolving hypothesis that intestinal bacteria induce inflammation predominantly as a result of a weakened innate mucosal barrier in genetically predisposed individuals. This article discusses our current understanding of the primary events of disease. Together, these findings should result in new therapeutic avenues aimed at restoring antimicrobial barrier function to prevent a bacterial-triggered inflammatory response.

Crohn's disease (CD), a major form of human inflammatory bowel disease, is characterized by primary immunodeficiencies. The nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) is essential for intestinal homeostasis in response to both dietary- and microbiota-derived signals. Its role in host defense remains unknown, however. We show that PPARgamma functions as an antimicrobial factor by maintaining constitutive epithelial expression of a subset of beta-defensin in the colon, which includes mDefB10 in mice and DEFB1 in humans. Colonic mucosa of Ppargamma mutant animals shows defective killing of several major components of the intestinal microbiota, including Candida albicans, Bacteroides fragilis, Enterococcus faecalis, and Escherichia coli. Neutralization of the colicidal activity using an anti-mDefB10 blocking antibody was effective in a PPARgamma-dependent manner. A functional promoter variant that is required for DEFB1 expression confers strong protection against Crohn's colitis and ileocolitis (odds ratio, 0.559; P = 0.018). Consistently, colonic involvement in CD is specifically linked to reduced expression of DEFB1 independent of inflammation. These findings support the development of PPARgamma-targeting therapeutic and/or nutritional approaches to prevent colonic inflammation by restoring antimicrobial immunity in CD.

Objectives: Adult ileal Crohn's disease (CD) is characterized by a specific decrease in ileal Paneth cell alpha-defensins. In addition to NOD2, we previously identified a disturbance of the Wnt-signaling transcription factor TCF-4 as a major mechanism for this deficiency. The aim of this study was to evaluate human alpha-defensin-5 (HD-5) and TCF-4 in an independent cohort of pediatric CD patients., Methods: Expression levels of HD-5 and TCF-4 mRNA were quantified by real-time PCR in biopsies from newly diagnosed untreated pediatric CD patients (<18 years, n=36) and age-matched symptomatic non-inflammatory bowel disease controls with a histologically normal gut (n=29). To assess the influence of current inflammation, mucosal interleukin-8 (IL-8) and fecal calprotectin levels were determined., Results: Small intestinal HD-5 and TCF-4 mRNA were significantly reduced in pediatric ileal CD (L1+L3) (P=0.022 and P=0.0005, respectively) and were significantly correlated (r=0.499; P=0.0001). In ileal but not colonic CD, TCF-4 was also reduced in the colon (P=0.005). Importantly, both HD-5 and TCF-4 were independent of inflammation, as measured by IL-8 expression or fecal calprotectin. In contrast to the small intestine, colonic Paneth cell HD-5 mRNA was significantly elevated in colonic CD (L2) (P=0.026) and was correlated with fecal calprotectin levels (r=0.481; P=0.020)., Conclusions: In this study, we describe a specific decrease in HD-5 and TCF-4 mRNA expression levels in children with ileal CD. In the small intestine, this decrease was independent of current inflammation, whereas inflammation seems to induce Paneth cell metaplasia in the colon. Our data extend the hypothesis of an important role of antimicrobial host defense in pediatric CD patients.

Paneth cells (PCs) are specialized epithelial cells predominantly found in the small intestinal crypts of Lieberkuehn. They produce different broad spectrum antimicrobial peptides most abundantly the alpha-defensins HD-5 and -6 (DEFA5 und DEFA6). Both these PC products show a specific reduction in small intestinal Crohn's disease (CD) - a form of inflammatory bowel disease (IBD). Their decrease is independent of current inflammation and an association with a NOD2 frameshift mutation has been demonstrated. More recently, another independent and even more frequent mechanism has been found which is linked to diminished levels of the Wnt pathway transcription factor TCF7L2 (also known as TCF4). Besides regulating the expression of HD-5 and HD-6 as TCF4 target genes, the Wnt pathway also orchestrates Paneth cell differentiation and maturation and controls stem cell maintenance in the small intestine. Besides NOD2 (which is predominantly expressed in PC) and ATG16L1 (inter alia important in the exocytosis of PC products), TCF4 is the third gene which is associated with small intestinal CD and Paneth cell antimicrobial function. Thus, Paneth cells seem to be key player emphazising a paramount importance of antimicrobial host defense in small intestinal CD pathogenesis., (Copyright 2009. Published by Elsevier GmbH.)

Defensins are natural mucosal antimicrobial peptides and their broad spectrum activity against aerobic or facultative anaerobic bacteria has been well investigated. The aim of this study was to systematically examine the antibacterial activity of the small intestinal Paneth cell derived alpha-defensin HD5 and the major colonic beta-defensins HBD-1-3 against strict anaerobic intestinal bacteria. The antibacterial activity was assessed with a flow cytometric assay employing a membrane potential sensitive dye as marker for loss of cell viability. The majority of the tested strains belonging to the dominant anaerobe genera of the gut, Bacteroides and Parabacteroides, were only minimally affected by the constitutively expressed defensins HD5 and HBD-1. The inducible defensin HBD-2 had a limited antibacterial effect, whereas the inducible HBD-3 exhibited potent activity against most strains. The effect of HBD-3 on Bacteroides sp. appeared to be dependent on the presence of oxygen. Bacteroides fragilis strains isolated from blood during bacteremia or from extraintestinal infections were more resistant to HBD-3 than strains from the physiological gut flora. Thus, defensin resistance is not only species- but also strain-specific and may be clinically relevant in the host-bacteria interaction influencing mucosal translocation and systemic infection.

Background: Crohn's disease (CD) of the pouch can occur in patients with restorative proctocolectomy and ileal pouch-anal anastomosis originally performed for a preoperative diagnosis of ulcerative colitis (UC). CD of the pouch was often observed in patients with a family history of CD. The purpose was to determine whether the family history of CD increased the risk for CD of the pouch in patients who underwent restorative proctocolectomy., Methods: A total of 558 eligible patients seen in the Pouchitis Clinic were enrolled, including 116 patients with CD of the pouch and 442 patients with a normal pouch or other pouch disorders. Demographic and clinical variables were included in the study. Multivariable logistic regression analyses were performed., Results: The adjusted multivariate logistic analyses revealed that the risk for CD of the pouch was increased in patients with a family history of CD, with an odds ratio (OR) of 3.22 (95% confidence interval [CI] 1.56-6.67), or with a first-degree relative with CD (OR = 4.18, 95% CI, 1.48-11.8), or with a greater number of family members with CD (OR = 2.00 per family member, 95% CI, 1.19-3.37), adjusting for age, gender, smoking status, duration of IBD, duration of having a pouch, and a preoperation diagnosis of indeterminate colitis or CD. In addition, patients of younger age and longer duration of having a pouch had a higher risk for CD of the pouch. A diagnosis of CD of the pouch was associated with a poor outcome, with a greater than 5-fold estimated increased odds of pouch failure (OR = 5.58, 95% CI, 2.74-11.4)., Conclusions: The presence of a family history of CD is associated with an increased risk for CD of the pouch, which in turn has a high risk for pouch failure.

Objectives: Irritable bowel syndrome (IBS) is a highly prevalent functional disorder. According to the Rome criteria, macroscopic and histological inflammation is a crucial exclusion criterion for IBS. Human defensins appear to be part of the innate immune system in the gastrointestinal tract. Human beta-defensin-2 (HBD-2) was the first inducible human antimicrobial protein discovered. The expression is induced by probiotic microorganisms and proinflammatory cytokines. Recent results imply that HBD-2 is expressed in active intestinal inflammation, especially in ulcerative colitis (UC). Our aim was to evaluate fecal measurements of HBD-2 in patients with active UC and IBS, and in healthy controls (HCs)., Methods: Fecal specimens were collected from a total of 100 participants (30 with active UC, 46 IBS, and 24 HCs). Exclusion criteria were the current use of probiotics and antibiotics. Furthermore, IBS patients with elevated C-reactive protein or leukocytes, a history of bacterial overgrowth or infectious gastrointestinal disease over the last 6 month were excluded. Disease status was addressed in all participating subjects by medical history and current symptoms. In addition, each IBS and UC patient underwent ileocolonoscopy with histopathology. Fecal inflammation markers lactoferrin (Lf) and calprotectin (Cal) were measured by enzyme-linked immunosorbent assay (ELISA) and reported as microg/g. Fecal HBD-2 was measured by ELISA and reported as ng/g feces. In addition, immunoblots were performed for fecal HBD-2. Paraffin-embedded tissue from colonic biopsies was tested for HBD-2 peptides by immunohistochemistry., Results: Lf as well as Cal was elevated in active UC (mean: 152.1+/-s.d. 374.7 microg/g; 103.5+/-87.1 microg/g), compared with IBS (8.3+/-19.4 microg/g; 18.6+/-23.3 microg/g), and HCs (0.4+/-0.5 microg/g; 7.1+/-7.9 microg/g). Scheffe post hoc tests revealed significant differences (P=0.006; P<0.001) between active UC vs. IBS and HC. In contrast, HBD-2 levels were highest in active UC (mean: 106.9+/-s.d. 91.5 ng/g), almost as high in IBS (pts 76.0+/-67.9 ng/g), and lowest for HCs (29.9+/-16.1 ng/g). Scheffe post hoc tests revealed significant differences (P<0.001) between the groups of patients (UC and IBS) vs. HCs. Immunohistochemical investigation was consistent with fecal secretion data and demonstrated the presence of beta-defensin 2 peptides in colonic epithelial enterocytes in UC as well as IBS patients with elevated fecal HBD-2., Conclusions: The results indicate significantly elevated levels of HBD-2 in patients with IBS compared with HCs and similar to those with active UC. The results support an activation of the mucosal innate defense system toward a proinflammatory response in IBS patients in the absence of macroscopic signs of inflammation.

Goblet cells are mucin-secreting intestinal cells forming the mucus layer that protects the mucosal surface. Ulcerative colitis (UC) has been associated with a defective colonic mucus layer and a reduced number of goblet cells. In experimental animals, colonic goblet cell differentiation is regulated by interacting transcription factors Hath1, KLF4 and the Notch, as well as Wnt pathways, whereas data in humans are limited. We investigated goblet cell differentiation factors and mucins in controls and in inflammatory bowel diseases (IBDs). We performed real-time PCR for Hath1, KLF4, several ligands, receptors and target genes of the Notch and Wnt pathways, as well as several mucins in biopsies from the sigmoid colon of controls (n=21), Crohn's disease (CD, n=48) and UC (n=40). In addition, Hath1 protein was quantitated with Western blot and localized with immunohistochemistry. Notably, the degree of inflammation as measured by IL-8 and histology was similar in both disease entities. The proportion of goblet cells was lowered in both IBDs, but specifically diminished in the upper third of the crypt in UC. Comparable levels of inflammation induced both Hath1 (2.0-fold, p<0.001) and KLF4 (1.8-fold for KLF4, p=0.031) mRNA expression in CD but not in UC (0.8-0.9-fold, ns). The differential induction was confirmed for Hath1 protein using Western blot. Hath1 immunostaining was found mostly in the lower half of the colonic crypts. Hath1, KLF4 and the Notch target gene Hes1 were significantly (p<0.001) and positively correlated. Moreover, both Hath1 and KLF4 were correlated (p<0.001) with MUC1, MUC2 as well as MUC4 in all control and IBD cohorts. The results indicate that both transcription factors are key regulators of goblet cell differentiation and mucin formation in the human colon. Conspicuously, inflammation is associated with an enhanced goblet cell differentiation in CD but not in UC, a defect possibly of pathogenic importance.

Reduced expression of Paneth cell antimicrobial alpha-defensins, human defensin (HD)-5 and -6, characterizes Crohn's disease (CD) of the ileum. TCF-4 (also named TCF7L2), a Wnt signalling pathway transcription factor, orchestrates Paneth cell differentiation, directly regulates the expression of HD-5 and -6, and was previously associated with the decrease of these antimicrobial peptides in a subset of ileal CD. To investigate a potential genetic association of TCF-4 with ileal CD, we sequenced 2.1 kb of the 5' flanking region of TCF-4 in a small group of ileal CD patients and controls (n = 10 each). We identified eight single nucleotide polymorphisms (SNPs), of which three (rs3814570, rs10885394, rs10885395) were in linkage disequilibrium and found more frequently in patients; one (rs3814570) was thereby located in a predicted regulatory region. We carried out high-throughput analysis of this SNP in three cohorts of inflammatory bowel disease (IBD) patients and controls. Overall 1399 healthy individuals, 785 ulcerative colitis (UC) patients, 225 CD patients with colonic disease only and 784 CD patients with ileal involvement were used to determine frequency distributions. We found an association of rs3814570 with ileal CD but neither with colonic CD or UC, in a combined analysis (allele positivity: OR 1.27, 95% CI 1.07 to 1.52, p = 0.00737), which was the strongest in ileal CD patients with stricturing behaviour (allele frequency: OR 1.32, 95% CI 1.08 to1.62, p = 0.00686) or an additional involvement of the upper GIT (allele frequency: OR 1.38, 95% CI 1.03 to1.84, p = 0.02882). The newly identified genetic association of TCF-4 with ileal CD provides evidence that the decrease in Paneth cell alpha-defensins is a primary factor in disease pathogenesis.

Crohn's disease may principally involve the whole gastrointestinal tract. Most commonly, the inflammation occurs in the small intestine and/or in the colon with stable disease location over the years. The pathogenesis of both disease phenotypes is complex, the likely primary defect lies in the innate rather than adaptive immunity, particularly in the chemical antimicrobial barrier of the mucosa. Crohn's ileitis is associated with a reduced expression of the Wnt signalling pathway transcription factor T-cell factor 4 (TCF4), which is regulating Paneth cell differentiation. As a result, the alpha-defensins and principal Paneth cell products HD5 and HD6 are deficiently expressed in ileal disease, independent of current inflammation. In contrast, Crohn's colitis is typically associated with an impaired induction of the beta-defensins HBD2 and HBD3 caused by fewer gene copy numbers in the gene locus of the beta-defensins on chromosome 8. This ileal and colonic defect in innate defence mediated by a deficiency of the protective alpha- and beta-defensins may enable the luminal microbes to invade the mucosa and trigger the inflammation. A better understanding of the exact molecular mechanisms behind ileal and colonic Crohn's disease may give rise to new therapeutic strategies based on a stimulation of the protective innate immune system.

Objectives: The Crohn's disease (CD) susceptibility gene, nucleotide-binding oligomerizetion domain 2 (NOD2)/caspase recruitment domain 15 (CARD15), is linked to the innate immune response associated with altered epithelial bacterial defense. Its relevance in antibiotic therapy of perianal fistulating CD remains elusive. The aim of the study was to explore systematically the association between NOD2/CARD15 variants and clinical response of perianal fistulas in patients using antibiotic therapy., Methods: Fifty-two patients (median age 36 yr) with draining perianal fistulas were treated with ciprofloxacin (N = 49) or metronidazole (N = 3) for a median duration of 7 wk. Complete response was defined as the absence of any draining fistula despite gentle finger compression. Genotyping for NOD2/CARD15 variants and human beta (beta)-defensin 2 (HBD-2) copies was performed by 5' nuclease assays (Applied Biosystems, Foster City, CA). The examiners and laboratory investigators were blinded. The Fisher exact test and Wilcoxon signed rank test were used for statistical analysis., Results: Ciprofloxacin was discontinued in one patient due to diarrhea after 2 wk. Complete fistula response was observed in 13 of 39 patients with NOD2/CARD15 wild-type (33.3%) compared with none in patients carrying NOD2/CARD15 variants (0%, P= 0.02). The median number of HBD-2 gene copies between responders and partial/nonresponders was similar., Conclusions: The study result suggests a substantial contribution of NOD2/CARD15 to the antibiotic treatment outcome of perianal fistulating CD. NOD2/CARD15 variants may predispose to an altered intestinal microflora in perianal fistulas that is less responsive to antibiotic treatment.

Background: Non-obese gallstone patients exhibit a diminished expression of apical sodium-dependent bile acid transporter (ASBT) in terminal ileum. Crohn's ileitis demonstrates a significant downregulation of this transporter., Aim: To test whether subclinical ileal inflammation contributes to gallstone disease., Methods: Biopsies from terminal ileum of female subjects with gallstone disease (n = 7), active Crohn's disease (n = 17) and controls (n = 22) were investigated. mRNA expression of ASBT, tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-8, c-jun and c-fos was measured. c-jun and c-fos protein levels were determined and hematoxylin and eosin staining was applied for ileal histology., Results: ASBT expression was comparably low both in gallstone (47% of controls, p = 0.0093) and Crohn's disease (42% of controls, p = 0.0008). In gallstone disease there was a non-significant trend towards elevated TNF-alpha and IL-1beta, but all cytokines were increased in active Crohn's disease. c-jun and c-fos were slightly diminished in patients with gallstones. Neither cytokines nor transcription factors correlated significantly with ASBT. The gallstone-associated ileal biopsies exhibited no histological inflammation., Conclusion: Although the expression of ASBT was similarly diminished in both gallstone and Crohn's disease, subclinical ileal inflammation does not appear to be relevant in gallstone patients. The mechanisms of transcriptional repression of ASBT in both diseases are apparently different., (2008 S. Karger AG, Basel.)

Defensins are endogenous antibiotics with microbicidal activity against Gram-negative and -positive bacteria, fungi, enveloped viruses and protozoa. A disturbed antimicrobial defense, as provided by Paneth and other epithelial cell defensins, seems to be a critical factor in the pathogenesis of Crohn's disease, an inflammatory disease of the intestinal tract. Different direct and indirect mechanisms leading to a breakdown of antimicrobial barrier function include direct changes in defensin gene numbers (e.g., copy number polymorphism), genetic mutations in pattern-recognition receptors (e.g., nucleotide-binding oligomerization domain 2) and, as described recently, a differentiation problem of epithelial stem cells mediated by the wingless type (Wnt) pathway. Knowledge regarding the regulation and biology of defensins provides an attractive target to open up new therapeutic avenues.

Ileal Crohn's disease (CD), a chronic mucosal inflammation, is characterized by two pertinent features: a specific decrease of Paneth cell-produced antimicrobial alpha-defensins and the presence of mucosal-adherent bacteria. A mutation in NOD2, the muramyl dipeptide recognition receptor, is found in some patients, which leads to an even more pronounced alpha-defensin decrease. However, the underlying mechanism remains unclear for the majority of patients. In this study, we report a reduced expression in ileal CD of the Wnt-signaling pathway transcription factor Tcf-4, a known regulator of Paneth cell differentiation and alpha-defensin expression. Within specimens, the levels of Tcf-4 mRNA showed a high degree of correlation with both HD5 and HD6 mRNA. The levels of Tcf-4 mRNA were decreased in patients with ileal disease irrespective of degree of inflammation, but were not decreased in colonic CD or ulcerative colitis. As a functional indicator of Tcf-4 protein, quantitative binding analysis with nuclear extracts from small intestine biopsies to a Tcf-4 high-affinity binding site in the HD-5 and HD-6 promoters showed significantly reduced activity in ileal CD. Furthermore, a causal link was shown in a murine Tcf-4 knockout model, where the comparably reduced expression of Tcf-4 in heterozygous (+/-) mice was sufficient to cause a significant decrease of both Paneth cell alpha-defensin levels and bacterial killing activity. Finally, the association between Paneth cell alpha-defensins and Tcf-4 was found to be independent of the NOD2 genotype. This new link established between a human inflammatory bowel disease and the Wnt pathway/Tcf-4 provides a novel mechanism for pathogenesis in patients with ileal CD.

Human beta-defensin 2 (hBD-2) is an inducible antimicrobial peptide synthesized by the epithelium to counteract bacterial adherence and invasion. Proinflammatory cytokines, as well as certain bacterial strains, have been identified as potent endogenous inducers. Recently, we have found that hBD-2 induction by probiotic Escherichia coli Nissle 1917 was mediated through NF-kappaB- and AP-1-dependent pathways. The aim of the present study was to identify the responsible bacterial factor. E. coli Nissle 1917 culture supernatant was found to be more potent than the pellet, indicating a soluble or shed factor. Chemical analysis demonstrated the factor to be heat resistant and proteinase digestible. Several E. coli Nissle 1917 deletion mutants were constructed and tested for their ability to induce hBD-2 expression in Caco-2 cells. Deletion mutants for flagellin specifically exhibited an impaired immunostimulatory capacity. Reinsertion of the flagellin gene restored the induction capacity to normal levels. Isolated flagellin from E. coli Nissle 1917 and from Salmonella enterica serovar Enteritidis induced hBD-2 mRNA significantly in contrast to the flagellin of the apathogenic E. coli strain ATCC 25922. H1 flagellin antiserum abrogated hBD-2 expression induced by flagellin as well as E. coli Nissle 1917 supernatant, confirming that flagellin is the major stimulatory factor of E. coli Nissle 1917.

Elafin (or skin-derived antileukoprotease) and secretory leukocyte protease inhibitor (SLPI) are serine antiproteases antagonizing human neutrophil elastase (HNE), thereby preventing tissue injury from excessive release of proteolytic enzymes by inflammatory cells. Furthermore, elafin and SLPI are "defensin-like" molecules with broad antimicrobial activity. The balance between proteases and antagonists may critically determine inflammatory processes in Crohn's disease (CD) and ulcerative colitis (UC). Real-time PCR was performed to quantitate colonic, proinflammatory cytokine IL-8, protease (HNE), and antiprotease mRNA (elafin and SLPI) in a total of 340 biopsies from 117 patients (47 CD, 45 UC, 25 controls). Histological inflammation was scored, and HNE, elafin, and SLPI were localized and semiquantified by immunostaining in 51 colonic paraffin sections (23 CD, 11 UC, 17 controls). Proinflammatory IL-8, degree of histological inflammation, and granulocyte content were similar in UC and CD. Elafin stained predominantly in the epithelium and SLPI in mucosal inflammatory cells. HNE mRNA levels and immunostaining were increased equally in both forms of inflammatory bowel disease. Levels of mRNA and immunostaining of the antiproteases elafin and SLPI were enhanced strongly in inflamed versus noninflamed UC. It is surprising that comparing inflamed versus noninflamed CD, this increase was significantly less pronounced for elafin and even lacking for SLPI. Despite comparable degrees of inflammation and protease levels, the induction of both antiproteases was attenuated in CD. This could contribute to the transmural depth of tissue destruction in CD. Elafin and SLPI may be added to the list of defensin-like peptides with diminished induction in CD versus UC.

Antimicrobial peptides and proteins are key effectors of innate immunity, expressed both by circulating phagocytic cells and by epithelial cells of mucosal tissues. In the human small intestine, Paneth cells are secretory epithelial cells that express the antimicrobials human alpha-defensin-5 (HD5), HD6, lysozyme and secretory phospholipase A(2) (sPLA(2)), and recent studies have implicated reduced HD5 and HD6 expression levels in the pathogenesis of ileal Crohn's disease. However, expression levels of these molecules have not been determined routinely by techniques that readily permit quantitative comparisons of their distribution between tissues and samples. Using quantitative real-time PCR with external standards and Northern blot analysis, we compared expression levels of mRNA encoding these four Paneth cell antimicrobial peptides, as well as circulating human neutrophil defensins in several different gastrointestinal tissues and the bone marrow. HD5 and HD6 were the most abundant antimicrobials expressed in the small intestine. The concentration of HD5 mRNA is approximately 5 x 10(5) copies per 10ng RNA in the jejunum and ileum; HD6 mRNA levels were about six times lower than those of HD5. With the exception of low levels in the pancreas (10(3) copies/10 ng RNA), the expression of HD5 and HD6 in tissues other than small intestine was at or below detectable limits. The expression of sPLA2 and lysozyme mRNA was observed in the small intestine (approximately, 3 x 10(3) and 9 x 10(3) copies/10 ng RNA, respectively), but also in several other tissues. Lysozyme expression was high in the duodenum (10(5) copies/10 ng RNA), and the protein localized to both Brunner's glands in the lamina propria and Paneth cells. By comparison, the hematopoietic alpha-defensins HNP1-3 mRNA were detected at 6 x 10(5) copies per 10 ng RNA in the bone marrow. These quantitative RT-PCR data from healthy tissues represents the first quantitative topographical assessment of antimicrobial expression in the gastrointestinal tract and provides a means to directly compare expression levels between healthy tissues and disease specimens for multiple antimicrobial peptides.

Defensins are endogenous antimicrobial peptides that protect the intestinal mucosa against bacterial invasion. It has been suggested that deficient defensin expression may underlie the chronic inflammation of Crohn disease (CD). The DNA copy number of the beta-defensin gene cluster on chromosome 8p23.1 is highly polymorphic within the healthy population, which suggests that the defective beta-defensin induction in colonic CD could be due to low beta-defensin-gene copy number. Here, we tested this hypothesis, using genomewide DNA copy number profiling by array-based comparative genomic hybridization and quantitative polymerase-chain-reaction analysis of the human beta-defensin 2 (HBD-2) gene. We showed that healthy individuals, as well as patients with ulcerative colitis, have a median of 4 (range 2-10) HBD-2 gene copies per genome. In a surgical cohort with ileal or colonic CD and in a second large cohort with inflammatory bowel diseases, those with ileal resections/disease exhibited a normal median HBD-2 copy number of 4, whereas those with colonic CD had a median of only 3 copies per genome (P=.008 for the surgical cohort; P=.032 for the second cohort). Overall, the copy number distribution in colonic CD was shifted to lower numbers compared with controls (P=.002 for both the surgical cohort and the cohort with inflammatory bowel diseases). Individuals with < or = 3 copies have a significantly higher risk of developing colonic CD than did individuals with > or = 4 copies (odds ratio 3.06; 95% confidence interval 1.46-6.45). An HBD-2 gene copy number of < 4 was associated with diminished mucosal HBD-2 mRNA expression (P=.033). In conclusion, a lower HBD-2 gene copy number in the beta-defensin locus predisposes to colonic CD, most likely through diminished beta-defensin expression.

Crohn's disease (CD), a chronic inflammatory disease of the intestinal mucosa is usually located in the small intestine (ileum) and or in the colon. Ileal CD has been linked to a mutation in the NOD2 gene, a bacterial recognition protein. A disturbed antimicrobial defense as provided by an arsenal of different epithelial defensins seems to be a critical factor in disease pathogenesis. Defensins are antimicrobial peptides and in the ileum are mainly expressed in Paneth cells (PCs), epithelial cells that also express NOD2. In the colon, defensins are expressed by enterocytes or metaplastic PCs. Ileal CD patients are characterized by a reduced antibacterial activity and a specific reduction of ileal PC defensins. In ileal Crohn's patients displaying a NOD2 mutation, this decrease is even more pronounced. In contrast, CD of the colon is characterized by an impaired induction of beta defensins in enterocytes. In conclusion, the regional localizations of CD, either ileal or colonic disease, can be linked to different defects in defensin expression. In line with these new findings, we predict that future therapeutic strategies aimed at restoring the host-microbe balance at the intestinal mucosa may prove superior to those that broadly suppress inflammation and adaptive immunity.

To determine the antibacterial activity of defensins and other antimicrobial peptides in biopsy extracts, we evaluated a flow cytometric method with the membrane potential sensitive dye bis-(1,3-dibutylbarbituric acid) trimethine oxonol [DiBAC4(3)]. This assay enables us to discriminate intact non-fluorescent and depolarized fluorescent bacteria after exposure to antimicrobial peptides by measurement at the direct target, the cytoplasmic membrane and the membrane potential. The feasibility of the flow cytometric assay was evaluated with recombinant human beta-defensin 3 (HBD-3) against 25 bacterial strains representing 12 species. HBD-3 showed a broad-spectrum dose dependent activity and the minimal dose to cause depolarization ranged from 1.25 to >15 microg/ml HBD-3, depending on the species tested. The antibacterial effect was diminished with sodium chloride or dithiothreitol and could be abrogated with a HBD-3 antibody. Additionally, isolated cationic extracts from human intestinal biopsies showed a strong bactericidal effect against Escherichia coli K12, E. coli ATCC 25922 and Staphylococcus aureus ATCC 25923, which was diminished towards E. coli at 150 mM NaCl, whereas the activity towards S. aureus ATCC 25923 remained unaffected at physiological salt concentrations. DTT blocked the bactericidal effect of biopsy extracts completely.

Production of inducible antimicrobial peptides offers a first and rapid defense response of epithelial cells against invading microbes. Human beta-defensin-2 (hBD-2) is an antimicrobial peptide induced in various epithelia upon extracellular as well as intracellular bacterial challenge. Nucleotide-binding oligomerization domain protein 2 (NOD2/CARD15) is a cytosolic protein involved in intracellular recognition of microbes by sensing peptidoglycan fragments (e.g. muramyl dipeptide). We used luciferase as a reporter gene for a 2.3-kb hBD-2 promoter to test the hypothesis that NOD2 mediates the induction of hBD-2. Activation of NOD2 in NOD2-overexpressing human embryonic kidney 293 cells through its ligand muramyl dipeptide (MDP) induced hBD-2 expression. In contrast, overexpression of NOD2 containing the 3020insC frame-shift mutation, the most frequent NOD2 variant associated with Crohn disease, resulted in defective induction of hBD-2 through MDP. Luciferase gene reporter analyses and site-directed mutagenesis experiments demonstrated that functional binding sites for NF-kappaB and AP-1 in the hBD-2 promoter are required for NOD2-mediated induction of hBD-2 through MDP. Moreover, the NF-kappaB inhibitor Helenalin as well as a super-repressor form of the NF-kappaB inhibitor IkappaB strongly inhibited NOD2-mediated hBD-2 promoter activation. Expression of NOD2 was detected in primary keratinocytes, and stimulation of these cells with MDP induced hBD-2 peptide release. In contrast, small interference RNA-mediated down-regulation of NOD2 expression in primary keratinocytes resulted in a defective induction of hBD-2 upon MDP treatment. Together, these data suggest that NOD2 serves as an intracellular pattern recognition receptor to enhance host defense by inducing the production of antimicrobial peptides such as hBD-2.

The pathogenesis of Crohn's disease (CD), an idiopathic inflammatory bowel disease, is attributed, in part, to intestinal bacteria that may initiate and perpetuate mucosal inflammation in genetically susceptible individuals. Paneth cells (PC) are the major source of antimicrobial peptides in the small intestine, including human alpha-defensins HD5 and HD6. We tested the hypothesis that reduced expression of PC alpha-defensins compromises mucosal host defenses and predisposes patients to CD of the ileum. We report that patients with CD of the ileum have reduced antibacterial activity in their intestinal mucosal extracts. These specimens also showed decreased expression of PC alpha-defensins, whereas the expression of eight other PC products either remained unchanged or increased when compared with controls. The specific decrease of alpha-defensins was independent of the degree of inflammation in the specimens and was not observed in either CD of the colon, ulcerative colitis, or pouchitis. The functional consequence of alpha-defensin expression levels was examined by using a transgenic mouse model, where we found changes in HD5 expression levels, comparable to those observed in CD, had a pronounced impact on the luminal microbiota. Thus, the specific deficiency of PC defensins that characterizes ileal CD may compromise innate immune defenses of the ileal mucosa and initiate and/or perpetuate this disease.

Despite many years of intensive research the pathogenesis of inflammatory bowel diseases is still enigmatic. All efforts to identify the cause of Crohn's disease and ulcerative colitis in a dysregulation of specific immune mechanisms have failed. This review presents a novel pathogenetic concept, based on the expression of natural mucosal antibiotic peptides. These so called defensins are part of innate immunity and defend the mucosa as antimicrobial peptides against intraluminal potentially pathogenic and invasive bacteria. In contrast to ulcerative colitis, Crohn's disease is characterised by a diminished defensin expression. This defect may represent the molecular pathogenesis of this disease.

Defensins are endogenous antibiotics with microbicidal activity against Gram-negative and Gram-positive bacteria, fungi, viruses and protozoa. In the gastrointestinal tract, defensins help regulate the composition and number of colonizing microbes, and protect the host from food-borne and water-borne pathogens. In health, the normal host relationship with the commensal luminal microbiota is beneficial, but the same commensal bacteria could have a pathogenic role in inflammatory diseases. A disturbance in antimicrobial defense, as provided by Paneth cells of the small intestine, seems to be a critical factor in the pathogenesis of ileal Crohn's disease, an inflammatory disease of the intestinal tract. The disruption of the critical balance between antimicrobial peptides and luminal bacteria might also explain other gastrointestinal infections and diseases. Elucidating the underlying mechanisms involved in the regulation and biology of defensins could open up new therapeutic avenues.

Many patients with ileal Crohn's disease, a chronic intestinal inflammation, carry mutations in the gene encoding NOD2 (CARD15), but the mechanistic details of how this mutation leads to disease are not fully understood. NOD2 is expressed in antigen-presenting cells and Paneth cells, which are secretory epithelial cells of the small intestine. Two complementary studies using genetically engineered murine models help to explain the association of NOD2 malfunction and mucosal disease. One study observes a dysregulation of proinflammatory responses, suggesting that the most common NOD2 mutation in humans results in a gain of function. The other study determined that NOD2-null mutations impair the Paneth-cell antimicrobial response, which is consistent with recent findings in humans. Together, these studies fuel optimism that new therapeutic directions might emerge to better treat this severe mucosal disease.

Crohn's disease is a chronic, inflammatory disease of the intestinal mucosa. Although intestinal bacteria are implicated in disease pathogenesis, the etiology is still unclear. The main location of disease is the small intestine (ileum) and the colon. Ileal disease has been linked to a mutation in the NOD2 gene. Defensins are antimicrobial peptides and in the ileum, are mainly expressed in Paneth cells, epithelial cells that also express NOD2. In the colon, defensins are expressed by enterocytes or metaplastic Paneth cells. Crohn's disease patients with ileal involvement, compared with controls or Crohn's patients without ileal involvement, have diminished expression of ileal Paneth cell defensins. This decrease is even more pronounced in Crohn's patients displaying a NOD2 mutation. In contrast, Crohn's disease of the colon is characterized by an impaired induction of beta-defensins in enterocytes. The colonic expression of the constitutive beta-defensin 1 is also decreased in the inflamed colonic mucosa, but this decrease is less specific to Crohn's disease, as it can also be found in ulcerative colitis patients. In conclusion, the regional localizations of Crohn's disease, ileal or colonic disease, can be linked to different defensin profiles. Crohn's disease of the ileum is associated with diminished defensin expression in Paneth cells. Crohn's disease of the colon is associated with diminished beta-defensin expression in enterocytes. Thus, it can be speculated that decreased defensin levels lead to a weakened intestinal barrier function to intestinal microbes and might be crucial in the pathophysiology of Crohn's disease.

Crohn's disease, a transmural inflammation of the gut, has been linked to good childhood hygiene, frequent use of antibiotics before diagnosis, adherent or invasive mucosal bacteria and a break in the tolerance of luminal bacteria. A decrease or lack of mucosal peptide antibiotics may play a central role in the etiopathogenesis of Crohn's disease. The dysregulated adaptive immune system may reflect only the primary break of the mucosal defence since the immune response is mostly directed against luminal bacteria. Crohn's disease patients with ileal involvement, as compared to controls and Crohn's disease patients without ileal disease, are characterized by a diminished expression of the ileal Paneth cell defensins. This decrease is even more pronounced in Crohn's disease patients with a mutation in the NOD2 gene, which is associated with Crohn's disease and ileal involvement. NOD2 is an intracellular peptidoglycan receptor and is expressed in Paneth cells. In contrast to ulcerative colitis, Crohn's disease of the colon is characterized by an impaired induction of human beta defensins 2 and 3. The hypothesis of an impaired mucosal antibacterial activity is also consistent with the benefit from antibiotic or probiotic treatment in certain disease states.

Background: Mutations in NOD2, a putative intracellular receptor for bacterial peptidoglycans, are associated with a subset of Crohn's disease but the molecular mechanism linking this protein with the disease pathogenesis remains unclear. Human alpha defensins (HD-5 and HD-6) are antibiotic effector molecules predominantly expressed in Paneth cells of the ileum. Paneth cells also express NOD2. To address the hypothesis that the function of NOD2 may affect expression of Paneth cell defensins, we compared their expression levels with respect to NOD2 mutations in Crohn's disease., Methods: Forty five Crohn's disease patients (24 with NOD2 mutations, 21 with wild-type NOD2) and 12 controls were studied. Real time reverse transcription-polymerase chain reaction was performed with mucosal mRNA for HD-5, HD-6, lysozyme, secretory phospholipase A2 (sPLA2), tumour necrosis factor alpha, interleukin 8, and human hypoxanthine phosphoribosyltransferase (housekeeping gene). Immunohistochemistry with anti-HD-5 and histological Paneth cell staining were performed in 10 patients with NOD2 mutations or wild-type genotypes., Results: Ileal expression of HD-5 and HD-6, but not sPLA2 or lysozyme, were diminished in affected ileum, and the decrease was significantly more pronounced in patients with NOD2 mutations. In the colon, HD-5, HD-6, and sPLA2 were increased during inflammation in wild-type but not in NOD2 mutated patients. In both the colon and ileum, proinflammatory cytokines and lysozyme were unaffected by NOD2 status. Immunohistochemistry identified Paneth cells as the sole source of HD-5., Conclusion: As alpha defensins are important in the mucosal antibacterial barrier, their diminished expression may explain, in part, the bacterial induced mucosal inflammation and ileal involvement of Crohn's disease, especially in the case of NOD2 mutations.

Little is known about the defensive mechanisms induced in epithelial cells by pathogenic versus probiotic bacteria. The aim of our study was to compare probiotic bacterial strains such as Escherichia coli Nissle 1917 with nonprobiotic, pathogenic and nonpathogenic bacteria with respect to innate defense mechanisms in the intestinal mucosal cell. Here we report that E. coli strain Nissle 1917 and a variety of other probiotic bacteria, including lactobacilli--in contrast to more than 40 different E. coli strains tested--strongly induce the expression of the antimicrobial peptide human beta-defensin-2 (hBD-2) in Caco-2 intestinal epithelial cells in a time- and dose-dependent manner. Induction of hBD-2 through E. coli Nissle 1917 was further confirmed by activation of the hBD-2 promoter and detection of the hBD-2 peptide in the culture supernatants of E. coli Nissle 1917-treated Caco-2 cells. Luciferase gene reporter analyses and site-directed mutagenesis experiments demonstrated that functional binding sites for NF-kappaB and AP-1 in the hBD-2 promoter are required for induction of hBD-2 through E. coli Nissle 1917. Treatment with the NF-kappaB inhibitor Helenalin, as well as with SP600125, a selective inhibitor of c-Jun N-terminal kinase, blocked hBD-2 induction by E. coli Nissle 1917 in Caco-2 cells. SB 202190, a specific p38 mitogen-activated protein kinase inhibitor, and PD 98059, a selective inhibitor of extracellular signal-regulated kinase 1/2, were ineffective. This report demonstrates that probiotic bacteria may stimulate the intestinal innate defense through the upregulation of inducible antimicrobial peptides such as hBD-2. The induction of hBD-2 may contribute to an enhanced mucosal barrier to the luminal bacteria.

Background: The distinction between idiopathic inflammatory bowel disease (IBD) and infectious, usually self-limited enterocolitis is still a diagnostic dilemma. Procalcitonin (PCT) is the prohormone of calcitonin and is considered a specific marker of bacterial infection. The aim of this prospective study was to determine the value of PCT in differentiating flares of IBD from self-limited colitis. In addition, because standard laboratory inflammatory parameters are poorly correlated with disease activity in IBD, the relation between PCT levels and disease activity was investigated., Methods: A total of 76 patients (26 Crohn's disease, CD; 25 ulcerative colitis, UC; and 25 patients with self-limited enterocolitis) were enrolled. Serum levels of PCT were measured by a sandwich immunoluminometric assay. C-reactive protein (CRP) levels, white blood cell counts, and stool cultures were obtained from all patients. Disease activity was assessed by the Crohn's disease activity index (CDAI) and the Truelove index for CD and UC, respectively., Results: Patients with self-limited enterocolitis showed significantly higher PCT levels when compared with IBD patients (0.36 ng/mL, range 0.18-1.7 vs 0.10 ng/mL, range 0.08 0.5, p < 0.001). For a PCT value of > or =0.4, the sensitivity for self-limited colitis was 92% and specifity 96%. The positive predictive value (PPV) for self-limited colitis was 96%, whereas the negative predictive value (NPV) was 93%. In IBD patients, PCT levels were in the normal range although significantly higher in active disease when compared with inactive disease (0.13 ng/mL, range 0.08-0.5 vs 0.09 ng/mL, range 0.08-0.15, p < 0.001). This difference was less pronounced for CD (0.11 ng/mL, range 0.08-0.2 vs 0.09 ng/mL, range 0.08-0.15, p < 0.05) than for UC (0.14 ng/mL, range 0.08-0.5 vs 0.09 ng/mL, range 0.08-0.11, p < 0.01). In CD, PCT levels correlated significantly 0.5, p < 0.01). with the CDAI (r =0.05, p <0.01)., Conclusions: The measurement of PCT offers two diagnostic options in IBD. Supranormal levels indicate self-limited enterocolitis. Furthermore, although within the normal range in IBD, PCT levels may serve as a new serological marker of disease activity.

Defensins are endogenous antimicrobial peptides with a broad activity spectrum. Even at micromolar concentrations gramnegative and grampositive bacteria, but also mycobacteria, as well as fungi (candida), viruses (herpes) and protozoa (giardia lamblia) are destroyed. As part of the innate immune system defensins are expressed by the intestinal epithelium and contribute to the maintenance of the mucosal barrier. This barrier appears to be defective in inflammatory bowel diseases since on one hand, the immune response is directed against the "normal" luminal bacterial flora and on the other hand, mucosal adherent and invasive bacteria have been observed in these diseases. A defective defensin expression may well explain these phenomena. Indeed, Crohn's disease of the terminal ileum, especially if associated with a NOD2 mutation, is characterised by a diminished alpha-defensin (human defensin 5 and 6) expression, and in inflamed Crohn's colitis, in contrast to ulcerative colitis, the beta-defensin (human beta-defensins 2 and 3) response is reduced. Through a deficient chemical mucosal barrier this defect could lead to increased bacterial invasion into the intestinal mucosa and might well explain an adequate inflammatory response. Although the final proof that this deficient defensin response leads to a reduced antibacterial activity of the intestinal mucosa is still lacking, the most plausible concept of pathogenesis of Crohn's disease is a defensin deficiency syndrome.

Antimicrobial peptides such as defensins provide nonspecific mucosal defense against a multitude of microorganisms. Recently, it has been shown that luminal bacteria may invade the mucosa in inflammatory bowel diseases, suggesting a defect in innate mucosal immunity. The aim of this study was to investigate the expression of human beta-defensins (HBD) in controls, Crohn's disease (CD), ulcerative colitis (UC), and unspecific inflammation. Up to 4 biopsies were taken from 103 patients (33 controls, 24 with Crohn's disease, 36 with ulcerative colitis, 10 with unspecific colitis). Mucosal mRNA was measured using real-time fluorescence temperature cycler reverse-transcription polymerase chain reaction with primers for HBD-1, HBD-2, HBD-3, tumor necrosis factor alpha, and interleukin 8. Mucosal HBD-1 expression was marginally decreased in both CD and UC. HBD-2 was increased exclusively in UC but not in CD. The expression of the novel defensin HBD-3 was strongly correlated with HBD-2 and also raised predominantly in UC. The expression of both inducible beta-defensins was enhanced in the state of inflammation. Expression of HBD-2 showed a weak correlation with interleukin 8 only in inflamed CD biopsies but not with tumor necrosis factor alpha. The missing induction of both inducible beta-defensins in CD as compared with UC may cause a defect in barrier function that predisposes to bacterial invasion.

This comprehensive review promotes the novel concept that a defensin deficiency, i.e. lack of mucosal peptide antibiotics, may play a pivotal role in the aetiopathogenesis of Crohn's disease. Such an impaired function of this chemical barrier is consistent with the epidemiological relationship of good domestic hygiene with the incidence of inflammatory bowel diseases. The disregulated adaptive immune system, formerly believed to be the major cause in the development of Crohn's disease, may reflect only the primary break of the mucosal defence since the immune response is mostly directed against lumenal bacteria. Recent work has identified five different defensins expressed in colonic mucosa. In contrast to ulcerative colitis, Crohn's disease is characterised by an impaired induction of human beta defensins 2 and 3. This deficient induction may be due to changes in the intracellular transcription by NFkappaB and the intracellular peptidoglycan receptor NOD2, mutated in Crohn's disease. These findings are consistent with the mucosal attachment of lumenal bacteria in inflammatory bowel diseases and the frequent occurrence of other infectious agents. The hypothesis of an impaired mucosal antibacterial activity is also consistent with the benefit from antibiotic or probiotic treatment in certain inflammatory bowel disease states.

Background/aims: Recent reports have suggested that Helicobacter pylori infection induces the mucosal antibiotic peptide human beta defensin 2 (HBD-2). Therefore, the present study investigated mRNA and peptide expression of four different defensins in the upper gastrointestinal tract in patients with H pylori positive and negative chronic gastritis., Materials/methods: Biopsies from the oesophagus to the duodenum were taken during routine gastroscopy in 71 individuals. Total RNA was extracted and the reverse transcription polymerase chain reaction was performed with primers for human defensins 5 and 6 (HD-5/6) or HBD-1 and HBD-2. Paraffin wax embedded tissue from gastric resections was tested for HD-5, HBD-1, and HBD-2 by immunohistochemistry., Results: Helicobacter pylori colonisation was associated with an increased percentage of positive biopsies with respect to HBD-2 in the corpus (p < 0.05). Helicobacter pylori had no impact on the gastric expression of HD-5 and HBD-1, whereas HD-6 was increased in the fundus. The abundant expression of alpha defensins in the duodenum and beta defensins in the oesophagus served as a positive control in each individual. Immunohistochemical analysis confirmed the presence of the HD-5, HBD-1, and HBD-2 peptides in gastric resection specimens., Conclusions: The recently described induction of HBD-2 upon H pylori infection was confirmed in a clinical setting of chronic gastritis. This phenomenon may be mediated by components of the pathogen itself or may occur secondary to immune events in chronic inflammation.

Objective: Various antimicrobial peptides such as defensins are part of innate immunity and contribute to the intestinal barrier that may be defective in inflammatory bowel disease (IBD). This study investigated beta-defensin mRNA and peptide expression in the colon from controls and patients with Crohn's disease, ulcerative colitis or unspecific colitis as inflammatory controls., Methods: Mucosal mRNA expression was measured by multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) with primers for human beta-defensin 1 (HBD-1) and human beta-defensin 2 (HBD-2) in CaCo-2 cells and in biopsies from 103 patients (33 controls, 24 Crohn's disease patients, 36 ulcerative colitis patients, 10 unspecific colitis patients). Paraffin-embedded tissue from colonic resections was tested for HBD-1 and HBD-2 peptides by immunohistochemistry., Results: HBD-1 mRNA was expressed constitutively whereas HBD-2 was induced by pro-inflammatory cytokines in CaCo-2 cells. HBD-1 mRNA was detectable in 61% of control and Crohn's disease biopsies and 53% of ulcerative colitis biopsies. HBD-2 transcript was expressed differentially, with 18% of control biopsies positive as opposed to 34% in Crohn's disease and 53% in ulcerative colitis. HBD-2 mRNA but not HBD-1 mRNA was expressed preferentially in inflamed areas. Immunohistochemical investigation demonstrated the presence of defensin peptides in colonic epithelium as well as the differential induction in IBD., Conclusions: HBD-1 is expressed constitutively in colonic tissue irrespective of inflammation. HBD-2 is barely present in uninflamed colon but it is induced in inflammation. The lower expression of HBD-2 in Crohn's disease compared with ulcerative colitis indicates different responses of the mucosal innate defence. Defensins may play a crucial role in controlling pathogen invasion in IBD, although the functional significance remains to be established.

Human alpha-defensins contribute to local intestinal host defense as part of innate immunity and may be of major relevance in microbial infection and chronic inflammatory bowel disease. The objective was to investigate human defensin 5 and 6 (HD-5, HD-6) mRNA expression by RT-PCR from colonic biopsies and peptide expression by immunohistochemistry from colonic resections under basal and inflammatory conditions. Both alpha-defensins were induced by proinflammatory cytokines in cell culture. HD-5 mRNA expression was enhanced in both idiopathic and nonidiopathic inflammatory states of the large bowel [32% of control vs 73% of unspecific colitis, 69% of Crohn's disease (CD) and 73% of ulcerative colitis (UC)], whereas HD-6 was specifically related to CD and UC (14% in controls vs 20% in unspecific coltis, 49% in CD and 42% in UC). Immunohistochemistry confirmed the presence of HD-5 in colonic epithelium. Antimicrobial peptides in the colon may be of importance in maintaining the mucosal barrier and controlling microbial invasion in inflammatory bowel diseases.