BAHD1-TNFR1-RIPK1/3轴调控结肠上皮细胞坏死性凋亡及在溃疡性结肠炎中作用与机制探讨

Ulcerative colitis (UC) is an idiopathic colonic mucosal disease, and its pathogenesis has not been fully understood. Recent studies have found that necroptosis can be mediated by TNFR1 in colonic epithelia. Our previous study observed that BAHD1 might play an essential role in UC. Necroptosis was significantly activated in UC patients while BAHD1 level was down-regulated. Inhibition of BAHD1 expression could upregulate TNFR1 expression, inducing severe inflammation and injury in Caco-2 cells. However, the underlying mechanism remains unclear. In this research project, we aim to explore and determine the role of BAHD1-TNFR1-RIPK1/3 axis in regulating necroptosis in UC by using western blot, Co-IP, dual luciferase reporter assay system, flow cytometry, target gene overexpression and siRNA-dependent silence as well as UC cell model and DSS-induced mouse model. The likely molecular mechanisms are involved with evoking high level of DAMPs and ROS production, activating endoplasmic reticulum stress (ERS), lysosomal membrane permeabilization, mitochondrial oxidative stress (MOS) and inducing severe inflammation as well as damage in intestinal epithelia. The findings of this project will assist in the eventual development of new therapeutic targets for the disease and possess high originality and clinical significance.

溃疡性结肠炎（UC）是慢性非特异粘膜性疾病，其发病机制尚未明确。近年报道TNFR1介导肠道上皮细胞坏死性凋亡；我们前期研究发现，BAHD1可能与UC密切相关，UC病人中BAHD1表达下调且坏死性凋亡途径激活，抑制Caco-2细胞BAHD1可增加TNFR1表达并加重结肠上皮细胞凋亡坏死，但相关机制不明。本项目在已有研究基础上，拟采用western blot、Co-IP、双荧光素酶报告基因系统、电镜、流式细胞术、靶基因过表达与干扰等技术以及UC细胞模型和小鼠模型，探讨并确定BAHD1-TNFR1-RIPK1/3轴通过调控坏死性凋亡的发生，促进DAMPs释放、ROS激活，并激发内质网应激（ERS），溶酶体释放和线粒体氧化应激反应（MOS），导致结肠上皮细胞炎症损伤及凋亡坏死加重，从而参与UC发病的作用及分子机制，为发现潜在的药物靶标奠定基础，具有较高创新性与临床价值。

背景：溃疡性结肠炎（UC）是一种免疫介导的慢性非特异肠道疾病，在我国发病呈升高趋势，且其发病机制尚不明确，近年报道TNFR1介导肠道上皮细胞坏死性凋亡。主要研究内容：通过转录谱印记法筛选出可能与UC发病密切相关的三个蛋白分子，阐明其在UC发生发展中的分子作用机理。重要结果：通过转录谱印记关联分析的方法，预测出106 种可能与UC发病密切相关蛋白分子，并在葡聚糖硫酸钠（dextran sodium sulfate, DSS）诱导的小鼠UC 模型验证表达，发现BAHD1表达明显最明显，故成为主要的研究蛋白。并发现UC病人病变肠黏膜中BAHD1表达下调、TNFR1表达上升且坏死性凋亡途径激活，抑制Caco-2细胞BAHD1可增加TNFR1表达并加重结肠上皮细胞凋亡坏死，采用western blot、双荧光素酶报告基因系统、流式细胞术、靶基因过表达与干扰等技术以及UC细胞模型和小鼠模型，确定BAHD1-TNFR1-RIPK1/3轴通过调控坏死性凋亡的发生，促进DAMPs释放、ROS激活，导致结肠上皮细胞炎症损伤及凋亡坏死加重，从而参与UC发病的作用及分子机制。意义：为基于BAHD1为潜在靶点的UC新药研发提供了新的理论依据。