Abstract:

Objective The study aimed to observe the expression characteristics of Let ⁃7b in the mouse model of inflammatory bowel disease（IBD），and analyze the regulation mechanism of Let⁃7b on heme oxygenase 1/ TB and CNC homology 1（Bach1/HO⁃1）axis in dendritic cells（DCs），so as to explore the mechanism of let⁃7b on MCEC wound repairing. Methods IBD mouse model was established and divided into normal group and model group. DCs cells and mouse colonic epithelial cells were cultured. DCs cells were divided into 7 groups，including Ⅰ：blank control group. Ⅱ：Negative control group of LET⁃7B simulants；Ⅲ：Let⁃7b simulant transfection group； Ⅳ：Negative control group with LET⁃7B inhibitor；Ⅴ：Let⁃7B inhibitor transfection group；Ⅵ：Let⁃7b inhibitor combined with Bach1 inhibitor epberberine hydrochloride（EC）intervention group；Ⅶ：Let⁃7b inhibitor，EC and HO⁃1 inhibitor zinc protoporphyrin（ZnPP）were co⁃treated in the intervention group. qRT⁃PCR and Western blot were used to detect the expression levels of Let ⁃ 7b，Bach1 and HO ⁃1 in colon tissues andculture supernatant of DCs. After cell culture，the supernatant of each group was co⁃cultured with MCEC and then the proliferation rate of colon epithelial cells was detected by 5⁃acetyney ⁃2′ ⁃deoxyuridine（Edu）kit method. Wound healing assay was used to detect the ability of cells to repair injury. The direct binding of Let⁃7b to the 3′UTR region of Bach1 wasanalyzed by luciferase gene reporter assay. Results Let⁃7b expression was decreased in model mice（P < 0.05）， and negatively correlated with Bach1 expression（P < 0.05），while positively correlated with HO ⁃ 1 expression （P < 0.05）. Let⁃7b was overexpressed or inhibited in DCs，and Bach1in culture supernatant was directly negatively regulated by Let⁃7b（P < 0.05），while in culture supernatant，HO⁃1 was positively regulated（P < 0.05）. Mecha⁃ nistic studies showed that Bach1 was the target gene of Let⁃7b in DCs，while HO⁃1 was the downstream target gene. In addition，after the supernatant of cell cultures in each group was co⁃cultured with colon epithelial cells，we observed that Let⁃7b mimic could significantly inhibit the extracellular expression of Bach1 and promote the expres⁃ sion of HO ⁃1 in cell culture supernatant，thereby promoting the proliferation of colonic mucosal epithelial cells and wound healing （all P < 0.05）. Conclusion Let⁃7b inhibits Bach1 of DCs and activates HO⁃1 to promote the proliferation and repair of intestinal mucosal damaged cells. These results suggest that regulation of Let⁃7b /Bach1/ HO⁃1 axis may be a potential therapeutic target for IBD.

Key words:

microRNALet ?7b, TB and CNC homology 1, heme oxygenase ?1, inflammatory bowel diseases