Helicobacter hepaticus CdtB Triggers Colonic Mucosal Barrier Disruption in Mice via Epithelial Tight Junction...

Background:Helicobacter hepaticus (H. hepaticus) has been demonstrated to have clinical relevance to the development of colitis in rodents. H. hepaticus produces cytolethal distending toxins (CDTs), which are identified as the most important virulence factors to the pathogenicity of CDT-producing bacteria in animals. However, the precise relationship between CDTs of H. hepaticus and intestinal barrier dysfunction remains unclear. The objective of the present study was to ascertain the impact of CdtB, the active subunit of CDTs, on the colonic mucosal barrier during H. hepaticus infection. Materials and Methods: We investigated the infection of male BALB/c mice, intestinal organoids, and IEC-6 cell monolayers by H. hepaticus or CdtB-deficient H. hepaticus (ΔCdtB). A comprehensive histopathological examination was conducted, encompassing the assessment of H. hepaticus colonization, the levels of mRNA expression for inflammatory cytokines, the expression levels of tight junction proteins, and the related signaling pathways. Results: The results demonstrate that the presence of ΔCdtB led to a mitigation of the symptoms associated with H. hepaticus-induced colitis, as evidenced by colon length shortening and the colon histological inflammation score. In addition, the levels of pro-inflammatory cytokines were reduced in the ΔCdtB group. Moreover, a downward trend was observed in the phosphorylation levels of STAT3 and nuclear factor-κB (p65). In vitro, the presence of H. hepaticus resulted in a reduction in the expression of tight junction (TJ) markers (ZO-1 and occludin) and an impairment of the F-actin structure in either the intestinal epithelium or intestinal organoids. However, these effects were reversed by CdtB deletion. Concurrently, both ROS levels and apoptosis levels were found to be significantly reduced in cells treated with the ΔCdtB strain. Mechanistically, myosin light chain kinase (MLCK) activation was observed in the H. hepaticus-infected group in vivo, whereas the MLCK inhibitor ML-7 was found to reverse the CdtB-induced alterations in TJ proteins in IEC6 cells. Conclusions: The collective findings demonstrate that CdtB plays a pivotal role in the H. hepaticus-induced colonic mucosal barrier. This is achieved through the regulation of TJs via the MLCK/pMLC2 signaling pathway, which is linked to elevations in oxidative stress and inflammation within intestinal epithelial cells.