Title: Clinical Improvement in IBS After Disodium Cromoglycate Involves Mast Cell-Mediated Toll-Like Receptor Signaling Downregulation
Abstract: in ileal and colonic inflammatory bowel diseases (IBD).Materials and Methods: The expression of OLFM4, Muc1, Muc2 and IL8 was measured by quantitative RT-PCR in 200 ileal and colonic biopsies of patients with active/inactive IBD and controls.OLFM4 immunostaining was performed in formalin fixed or carnoy-fixed and paraffin-embedded tissue.The mRNA levels of the major antimicrobial peptides in mice (Cryptidin-1, mBD1, mBD2), as well as mMuc1 and mMuc2 were measured in OLFM4 knock-out mice compared to wild type controls.The effect of bacterial stimulation with different apathogenic bacteria on OLFM4 expression was determined using the mucin producing colon carcinoma cell line LS174T in the presence and absence of DBZ, a gamma-secretase inhibitor blocking the Notch pathway.Results: As expected, IL8 expression was significantly augmented in inflamed biopsies with comparable levels in the active IBD subgroups.OLFM4 mRNA was significantly induced in colonic inflamed vs. noninflamed Crohn′s disease (CD, 3.6-fold, p<0.0001) and even more pronounced in UC (8.2-fold, p<0.0001).The colonic increase of OLFM4 under inflammatory conditions was confirmed on protein level by immunostaining.In contrast to controls during inflammation the staining expanded from the lower crypts up to the epithelial surface.Moreover, OLFM4 was found in the crypt lumen/mucus and thus seems to be secreted.Muc1 expression also showed a significant induction in ileal (5.8-fold, p>0.0001) and colonic (2.3-fold, p<0.001) inflamed vs. noninflamed CD, whereas this induction was less pronounced in inflamed vs. noninflamed UC (1.9-fold, p=0.002).Colonic OLFM4 and Muc1 expression were correlated (r=0.59).Muc2 mRNA was significantly induced in inflamed vs. noninflamed ileal CD (1.5-fold, p=0.02), but not in the colon.LS174T experiments showed a significant induction of OLFM4 expression after incubation with E. coli K12, E. coli Nissle and Bacteroides vulgatus.DBZ treatment resulted in a downregulation and a block of this induction.OLFM4 knock-out mice showed similar expression profiles of Cryptidin-1, mBD1, mBD2 and mMuc2 as compared to wild types.In contrast, mMuc1 expression seems to be significantly decreased in the colon of OLFM4 knock-out mice vs. wild-types.Conclusion: It is plausible that OLFM4 is involved in the epithelial defense in IBD upon bacterial induction, apparently secreted into mucus.Its precise function is still unclear.