外泌体源miR-125a调控TNF-α介导海洛因依赖者戒断期肠屏障损伤修复机制

Addiction was one of the majior characters of Heroin abuse, which also could cause nervous system damage, our previous study revealed that heroin could cause intestinal barrier injury and lead to serious infections, it also affected the withdrawal effect of heroin affection and even death. However, its mechanism was still unknown. Our previous works revealed that the increase of exsome miR-125a from immune cells during heroin withdrawal phase, decrease in TNF-α, which might assoctiated with the repair of intestinal mucosal epithelial cells. In view of the fact that TNF-a is the target gene of miR-125a, thus, we hypothesized that the miR-125a, an immune cell-derived exosome, might regulate the repair of intestinal barrier injury by TNF-α in heroin addicts. To verify this hypothesis, we intend to construct the co-culture model of human intestinal mucosa epithelial/immune cell and the heroin addiction primate model to investigate the affection of miR-125a and the recory of intestinal barrier injury during the during the stage of heroin abuse, withdrawal and drug intervention, and clarify the the relationships between exosome miR-125a and inflammatory cytokines TNF-α, and verify the potential pathyway of exosome miR-125a on regulating TNF-α during the recovery statge of intestinal barrier injury for heroin abuser and its possible molecular mechanisms. The results would provide a new perspective to repaire the intestinal barrier injury of heroin abuser.

海洛因依赖除可引起神经系统损害外，我们发现还可以引起肠屏障损伤，并导致严重感染，影响戒断效果，甚至死亡，但机制不明。TNF-α等炎症因子是引发肠道炎症的关键靶标，且受非编码RNA调控。我们预实验结果表明海洛因依赖者戒断期外泌体源miR-125a升高、TNF-α降低和肠上皮细胞损伤后的修复相关，生物信息学分析提示TNF-α是miR-125a的靶基因。我们提出假说：外泌体miR-125a调控TNF-α介导海洛因依赖者肠屏障损伤的修复。为验证上述假说，我们拟构建海洛因依赖细胞模型与灵长类动物模型，观察海洛因滥用、戒断及药物干预对细胞及动物模型肠上皮细胞损伤修复作用。在分子、细胞及动物水平探讨外泌体miR-125a和TNF-α的调控关系，进一步明确外泌体miR-125a调控TNF-α在海洛因依赖者肠屏障损伤修复的相关通路及可能的分子机制。为海洛因依赖者肠屏障损伤修复及引发的严重感染防治提供新视角。

甲基苯丙胺戒断可引起神经等多系统损害，我们发现其还可引起肠屏障损伤，导致严重感染，影响戒断效果，甚至死亡，但机制不明。我们通过戒断人群外周血多因子检测显示TNF-α与肠屏障损伤相关指标呈正相关，提示其可能是肠屏障损伤修复的重要靶标。本课题围绕TNFα甲基苯丙在胺导致的肠屏障损伤中的作用及其上下游调控机制展开探索。研究结果显示：（1）甲基苯丙胺戒断人群存在严重肠屏障损伤情况，且不同戒断期肠屏障损伤情况不一，急性戒断期最为严重，提示急性戒断期是可能干预的关键时期；（2）甲基苯丙胺戒断人群外周血多种细胞因子水平异常，主要表现为促炎因子上调，抑炎因子下调，其中TNF-α与肠屏障损伤指标呈强正相关，通过干预TNF-α表达，可缓解甲基苯丙胺导致的肠屏障损伤，提示TNF-α可能参与甲基苯丙胺导致的肠屏障损伤发生发展；（3）通过对甲基苯丙胺戒断人群血浆外泌体miRNAs检测与表达谱分析，筛选出差异表达的miRNAs。在细胞实验中发现miR-181c-5p靶向下调TNFα，并改善肠屏障损伤相关蛋白，证实miR-181c-5p通过靶向下调TNFα参与甲基苯丙胺导致的肠屏障损伤；（4）由于免疫细胞是体内细胞因子的主要来源，我们在人群及动物水平通过免疫细胞亚型分析，筛选出参与甲基苯丙胺导致肠屏障损伤发生发展的免疫细胞亚型。本课题深入的探讨了TNFα及其上下游调控分子在甲基苯丙胺导致的肠屏障损伤中的作用，为甲基苯丙胺导致的肠屏障损伤机制研究提供了新的理论依据，为其早期诊断及干预提供了潜在的分子靶点，顺利完成了研究内容，达到了预期科研目标。同时我们筛选出与甲基苯丙胺导致的肠屏障损伤相关的免疫细胞亚型，为研究的进一步深入提供了线索。