HSPA5/HSPA1A泛素化蛋白降解途径在溃疡性结肠炎中的作用机制及复方苦参汤的干预作用研究

The balance of proliferation and apoptosis of intestinal epithelial cells is essential for maintaining the function of the intestinal mucosal barrier. Continuous endoplasmic reticulum stress in intestinal epithelial cells activates NF-κB signaling, whereas uncleared apoptotic cells inhibit Treg proliferation and promote the development and progression of ulcerative colitis (UC). Heat shock protein A5 (HSPA5) is a key regulator in endoplasmic reticulum stress. Our previous study found that HSPA5 was highly expressed in UC tissue samples. Our further overexpression of HSPA5 in HeLa cells was found to upregulate HSPA1A in the ubiquitination protein degradation pathway. Further, it is necessary to further determine the molecular mechanism by which HSPA5 regulates HSPA1A gene expression in intestinal epithelial cells, as well as HSPA5 downstream target genes. Therefore, we propose the hypothesis that HSPA5/HSPA1A regulates FOXP3 ubiquitination protein degradation in Treg which may participate in UC. This study investigated the effects of HSPA5 on the biological characteristics of intestinal epithelial cells; and analyzed the downstream pathways and important genes regulated by transcriptome sequencing; and intervened in the UC mouse model by compound Kushen Decoction, suggesting that Fufang Kushen Decoction may work by this pathway, which will may provide new strategies for clinical treatment.

肠上皮细胞的增殖与凋亡的平衡对于维持肠黏膜屏障的功能至关重要。肠上皮细胞持续性内质网应激会激活NF-κB信号，而未清除的凋亡细胞会抑制Treg增殖促进溃疡性结肠炎（UC）发生和发展。热休克蛋白A5(HSPA5)是内质网应激中的关键调节器，我们前期研究发现UC组织样本中HSPA5显著高表达。我们进一步在HeLa细胞中过表达HSPA5发现会上调泛素化蛋白降解途径中的HSPA1A。还需进一步在肠上皮细胞中确定HSPA5调控HSPA1A基因表达的分子机制，以及HSPA5下游靶基因。因此，我们提出HSPA5/HSPA1A调控Treg中FOXP3泛素化蛋白降解参与UC形成的假说。本课题研究HSPA5对肠上皮细胞生物学特性的影响；通过转录组测序分析其调控的下游通路和重要基因，以及可变剪接事件；在UC小鼠模型中予复方苦参汤干预，推测复方苦参汤可能通过该途径发挥作用，为临床治疗提供新的策略。

肠上皮细胞持续性内质网应激会激活NF-κB信号，而未清除的凋亡细胞会抑制Treg增殖促进溃疡性结肠炎（UC）发生和发展。热休克蛋白A5(HSPA5)是内质网应激中的关键调节器，我们前期研究发现UC组织样本中HSPA5显著高表达。我们进一步在HeLa细胞中过表达HSPA5发现会上调泛素化蛋白降解途径中的HSPA1A。还需进一步在肠道中确定HSPA5调控HSPA1A基因表达的分子机制，以及HSPA5下游靶基因。因此，本课题探讨了HSPA5/HSPA1A在UC中的调节机制，以HSPA5的抑制及与复方苦参汤协同对UC的治疗作用研究，调节Th17/Treg平衡，控制炎症进程，从而提高治疗UC的疗效。动物实验结果表明，HSPA5可以靶向抑制HSPA1A；抑制HSPA5的表达，可以缓解DSS诱导的老鼠结肠炎，从而抑制Th17分化及功能，促进Treg分化及功能，脾脏及肠系膜淋巴结 Th17 细胞的比例、结肠组织中促炎细胞因子 IL-17A、IFN-γ、IL-6 的含量及 IL-17A、IFN-γmRNA水平明显降低，Treg 细胞的比例、FOXP3 mRNA 水平、抑炎因子 TGF- β1、IL-10 的含量及 TGF-β1mRNA 水平明显升高，结肠组织中 HSPA5、HSPA1A 及 CHIP 蛋白表达及 mRNA 相对含量明显降低。HSPA5可通过调节DSS诱导的结肠炎小鼠的Th17 / Treg细胞平衡，协同复方苦参汤会增强治疗效果，显着改善结肠炎小鼠的症状和病理损害，并影响免疫功能。由此说明，抑制HSPA5联合协同复方苦参汤可能是治疗UC的新方法。

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