Hongwei Zhao

BACKGROUND: Intestinal hyper-permeability plays a critical role in the etiopathogenesis of inflammatory bowel disease (IBD) by affecting the penetration of pathogens, toxic compounds and macromolecules. 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], the active form of vitamin D, has been shown to be an important regulator of IBD and recent epidemiology suggests that patients with IBD have an impaired vitamin D status. The purpose of this study is to investigate the possible protective effects of 1,25(OH)2D3 on mucosal injury and epithelial barrier disruption on dextran sulfate sodium (DSS)-induced acute colitis model. METHODS: We used DSS-induced acute colitis model to investigate the protective effects of 1,25(OH)2D3 on mucosal injury and epithelial barrier integrity. Severity of colitis was evaluated by disease activity index (DAI), body weight (BW) change, colon length, histology, myeloperoxidase (MPO) activity, and proinflammatory cytokine production including tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). In vitro the protective role of 1,25(OH)2D3 was assessed by incubating Caco-2 cells with or without DSS and measuring transepithelial electrical resistance (TEER) and fluorescein isothiocyanate dextran (FITC-D). The intestinal permeability was analyzed by FITC-D, bacterial translocation and measurement of lipopolysaccharide (LPS). Ultrastructural features of the colon tissue and Caco-2 cell monolayer were observed by electron microscopy. Expressions of tight junction (TJ) proteins in the colon mucosa and Caco-2 cells were detected by immunohistochemistry, immunofluorescence, Western blot and real-time fluorescent quantitative PCR, respectively. RESULTS: DSS-induced acute colitis model was characterized by a reduced BW, AUC of BW, serum calcium, higher DAI, AUC of DAI, shortened colon length, elevated MPO activity, worsened histologic inflammation, increased mononuclear cell numbers in mesenteric lymph nodes (MLNs) and colonic lamina propria (LP), and enhanced proteins and mRNA levels of TNF-α and IFN-γ. 1,25(OH)2D3 markedly increased expressions of TJ proteins and mRNA and decreased the FITC-D permeability and the level of LPS. Furthermore, 1,25(OH)2D3 abrogated bacterial translocation to MLNs and ameliorated ultrastructural features of the colon epithelium by scanning electron microscopy (SEM). In vitro, 1,25(OH)2D3 increased TEER, TJ proteins and mRNA expressions, decreased the FITC-D permeability, and preserved structural integrity of the TJ in Caco-2 cells. CONCLUSIONS: 1,25(OH)2D3 may play a protective role in mucosal barrier homeostasis by maintaining the integrity of junction complexes and in healing capacity of the colon epithelium. 1,25(OH)2D3 may represent an attractive and novel therapeutic agent for the adjuvant therapy of IBD.

Binding of the lipid A portion of bacterial lipopolysaccharide (LPS) to leukocyte CD14 activates phagocytes and initiates the septic shock syndrome. Two lipid A analogs, lipid IVA and Rhodobacter sphaeroides lipid A (RSLA), have been described as LPS-receptor antagonists when tested with human phagocytes. In contrast, lipid IVA activated murine phagocytes, whereas RSLA was an LPS antagonist. Thus, these compounds displayed a species-specific pharmacology. To determine whether the species specificity of these LPS antagonists occurred as a result of interactions with CD14, the effects of lipid IVA and RSLA were examined by using human, mouse, and hamster cell lines transfected with murine or human CD14 cDNA expression vectors. These transfectants displayed sensitivities to lipid IVA and RSLA that reflected the sensitivities of macrophages of similar genotype (species) and were independent of the source of CD14 cDNA. For example, hamster macrophages and hamster fibroblasts transfected with either mouse or human-derived CD14 cDNA responded to lipid IVA and RSLA as LPS mimetics. Similarly, lipid IVA and RSLA acted as LPS antagonists in human phagocytes and human fibrosarcoma cells transfected with either mouse or human-derived CD14 cDNA. Therefore, the target of these LPS antagonists, which is encoded in the genomes of these cells, is distinct from CD14. Although the expression of CD14 is required for macrophage-like sensitivity to LPS, CD14 cannot discriminate between the lipid A moieties of these agents. We hypothesize that the target of the LPS antagonists is a lipid A recognition protein which functions as a signaling receptor that is triggered after interaction with CD14-bound LPS.