胆道闭锁T细胞中ERK通路调控干扰素-γ基因甲基化的机制

Biliary atresia (BA) is the most common cause of obstructive jaundice in infants and accounts for over half of children who undergo liver transplantation. The etiology of BA remains unknown. Recent evidence indicates that BA is a virus-induced autoimmune disease characterized by impaired T cells. DNA methylation levels regulate autoimmunity-related gene in T cells. ERK signaling pathway in T cells plays an important role in DNA methylation. In our previous research, we found that autoimmunity-related gene interferon gamma (IFN-γ) mRNA expression levels were significantly increased in BA, and the IFN-γ gene promoter region was hypomethylated and negatively correlated with DNA methylation. The protein levels of ERK 1/2, p-ERK 1/2 and DNA methyltransferase (DNMT) 1 in BA were significantly lower. Therefore, this study intends to investigate the role of ERK signaling pathway and the regulation of methylation, and further study the role of "ERK signaling pathway - methylation regulation" in BA. First, we will detect the expression of ERK signaling pathway and DNA methyltransferase in BA, and validate the role of ERK signaling pathway to DNA methyltransferase. Meanwhile, we will investigate genomic and gene specific DNA methylation levels in BA, specially interferon gamma (IFN-γ) gene, and further examine the role of "ERK signaling pathway induces the methylation regulation of IFN-γ". Thus, this study suggestes that the role of "ERK signaling pathway - methylation regulation" involve in the molecular mechanism of BA. Finally, the cell model and animal model will be used to evaluate the likelihood of preventing the occurrence of BA by blocking the ERK signaling pathway.

胆道闭锁是儿童最常见的严重肝脏疾病，目前认为是自身免疫性疾病，与T淋巴细胞功能异常有关。DNA低甲基化可以引起甲基化敏感免疫相关基因表达升高，与T细胞功能异常密切相关，但是机制尚不明确。胞外信号调节激酶（ERK）信号通路在DNA甲基化中起重要作用。前期研究中，我们发现胆道闭锁中甲基化敏感免疫相关基因IFN-γ启动子区低甲基化，ERK通路关键分子表达明显下调。因此，拟进一步以ERK通路与DNA甲基化的作用为出发点，研究"ERK通路调控DNA甲基化"在胆道闭锁中的作用。首先，检测胆道闭锁中ERK通路、DNA甲基转移酶的表达情况；接着，验证前者对后者的正向调控作用；进一步检测IFN-γ基因启动子区甲基化状态，并考察"ERK通路介导IFN-γ基因启动子区甲基化"的作用；从而揭示ERK通路通过表观遗传学调控影响胆道闭锁发生的分子机制；最后，应用细胞、动物模型评价ERK通路在胆道闭锁发生中的作用。

胆道闭锁是严重危害患者生命健康的肝脏疾患，也是儿童肝移植的首位原因，目前认为是自身免疫性疾病，与T淋巴细胞功能异常有关。既往研究指出DNA低甲基化可以引起甲基化敏感免疫相关基因表达升高，与T细胞功能异常密切相关，但是机制尚不明确，而胞外信号调节激酶（ERK）信号通路在DNA甲基化过程中起着重要作用。本研究在此基础上，研究发现胆道闭锁中甲基化敏感免疫相关基因IFN-γ启动子区发生低甲基化、ERK通路关键分子表达明显下调。进一步研究发现胆道闭锁中ERK通路、DNA甲基转移酶1的表达降低，且前者对后者具有正向调控作用；而DNA甲基转移酶1又负向调控IFN-γ基因，并导致IFN-γ基因启动子区发生低甲基化，转录水平表达升高；以上研究明确了ERK通路对IFN-γ基因DNA甲基化的调控作用。从而揭示ERK通路通过表观遗传学调控影响胆道闭锁发生的分子机制，对现有的理论体系进行了新的深入和补充，为疾病的防治，特别是胆道闭锁的表观遗传学调控机制提供了新的理论和现实依据。