Inflammation-induced Occludin Downregulation Limits Epithelial Apoptosis by Suppressing Caspase-3 Expression

Abstract

Epithelial tight junctions are compromised in patients with gastrointestinal disease. Occludin is a protein component of tight junctions; its removal from tight junctions or reduced expression results in barrier loss. Knockout (KO) of occludin in mice, however, does not appear to affect intestinal in tight junction structure or function. We investigated whether mucosal injury or repair are compromised in occludin-KO mice. We performed studies with occludin-KO mice, mice with intestinal epithelial cell-specific KO of occludin, and B6 (control) mice, as well as mice that overexpress occludin from a transgene in the intestinal epithelium. Colitis was induced by administration of dextran sulfate sodium or 2,4,6-trinitrobenzene sulphonic acid. Intestinal tissues were collected from mice, patients with Crohn’s disease or ulcerative colitis, or subject without these diseases (controls) and analyzed by histology, immunohistochemistry, quantitative real-time PCR, and immunoblots. Occludin was knocked down in Caco-2 BBe cells. Mice with intestinal epithelial cell-specific KO of occludin developed less-severe colitis than control mice. This protection was due to reduced activation of intrinsic and extrinsic apoptotic pathways. Promoter analysis revealed that occludin increased transcription of the caspase 3 gene (CASP3). Mucosal biopsies from patients with Crohn’s disease or ulcerative colitis had lower levels of occludin than controls; reduced occludin correlated with lower levels of CASP3. Incubation of Caco-2 BBe monolayers with tumor necrosis factor caused occludin downregulation, which reduced CASP3 expression and prevented induction of apoptosis via the intrinsic pathway (stimulated by 5-fluorouracil) or extrinsic pathway (stimulated by tumor necrosis factor). In intestinal epithelial cells, the tight junction protein occludin increases expression of CASP3. Occludin loss reduces expression of CASP3 and susceptibility of these cells to apoptosis. Reduced levels of occludin and CASP3 in intestinal epithelial cells of patients with inflammatory bowel diseases might promote restoration of mucosal homeostasis in response to inflammatory conditions. Intestinal tissues from patients with inflammatory bowel diseases have reduced expression of the barrier protein occludin. Occludin loss protects cells from programmed cell death. Inflammation-induced downregulation of occludin might prevent damage in intestinal tissues.