基于Brg1/Nrf2信号通路探讨移植肝缺血再灌注损伤和丙泊酚后处理的保护机制

Reactive oxygen species induced oxidative damage plays a critical role in postoperative hepatic ischemia reperfusion injury(IRI) induced by orthotopic liver transplantation (OLT), however, the mechanism governing endogenous antioxidant regulation is unclear. In recent years, antioxidant enzymes expression induced by Nrf2 is considered to be a crucial event in protecting against oxidative damage. Of note, our preliminary study reveals that Nrf2 increases significantly after OLT but this does not result in enhanced function and expression of antioxidant enzymes. This unexpected disassociation between Nrf2 and endogenous antioxidant enzyme expression during Post-OLT liver IRI prompts us to postulate that there exists restricting factor(s) that affects Nrf2 normal functioning. Recent study indicates that the participation of Brg1, an ATPase subunit of the SWI/SNF chromatin remodelling complex, is mandatory for Nrf2 to induce endogenous antioxidant enzyme expression. Our preliminary experiment demonstrated that liver Brg1 was downregulated which was coincident with increased hepatic oxidative damage despite of the increase in liver Nrf2 expression, while propofol postcondition can upregulate Brg1 and enhance Nrf2 function and attenuated Post-transplantational liver IRI. Accordingly, we hypothesized that Brg1/Nrf2 signaling plays an important role in postoperative liver IRI induced by OLT and that activation of Brg1/Nrf2 signaling may represent a key mechanism of propofol postcondition protection. In our studies,we will use various models including in vivo rat model of OLT, hepatic IRI model respectively in Brg1 and Nrf2 knockout mice and in isolated cultured cells, in combination with Brg1 adenovirus transfection,and incorperate clinic reseach,in order to elucidate the role of Brg1, and in particular the role of Brg1/Nrf2 signaling in post-transplantational liver IRI and in propofol postcondition mediated protection.The proposed study will not only provide new mechanistic insight in post-transplantational liver IRI protection, but also provide experimental and practical guidance for the development of effective therapeutic strategies combating agaist liver IRI.

活性氧在移植肝缺血再灌注损伤（IRI）起关键作用，而相关的内源性抗氧化调控机制尚不清楚，近年发现Nrf2诱导抗氧化酶表达在IRI保护中起重要作用。而我们研究发现移植肝再灌注后Nrf2表达增强而抗氧化酶类表达和活性降低的分离现象，提示存在制约Nrf2作用的因素。新近发现Nrf2调节抗氧化酶表达需染色质重组复合物亚基Brg1参与；而我们预实验发现移植肝Brg1下调，尽管Nrf2表达增强却仍伴随严重的氧化损伤，丙泊酚后处理上调Brg1，促进Nrf2功能并减轻肝IRI。据此，我们设想Brg1/Nrf2通路激活在移植肝IRI和丙泊酚后处理保护中起关键作用。本课题拟用大鼠肝移植模型、供肝Brg1补充后移植模型、结合Brg1和Nrf2基因敲除鼠在体和细胞Brg1腺病毒转染后IRI模型等，及临床研究，阐明Brg1/Nrf2通路在移植肝IRI和丙泊酚后处理的作用及机制，为防治移植肝IRI提供理论及实验依据。

肝缺血再灌注损伤（HIRI）是临床常见的危重症，在肝移植术、创伤、肝门阻断的肝叶切除术、心跳骤停和失血性休克等过程中均不可避免地发生HIRI。本研究在明确HIR后肝脏损伤以及Brg1、Nrf2和其靶基因（HO-1和NQO1）表达动态变化的基础上，建立肝细胞缺氧复氧模型研究Brg1是否依赖Nrf2对肝细胞Nrf2靶基因HO-1和NQO1转录调控的作用和机制，进而构建Brg1过表达的转基因小鼠，在小鼠HIR模型中进一步探讨Brg1在HIRI中的作用和机制；在上述基础上，通过在体、体外和临床研究探讨丙泊酚后处理能否通过激活Brg1/Nrf2通路促进Nrf2靶基因表达从而对HIRI起保护作用。为干预或预防HIRI提供新的思路和有效靶点。本研究阐明Brg1是Nrf2介导的抗氧化酶类诱导性表达所必需，Brg1缺失是导致移植肝缺血再灌注损伤重要机制；证明Brg1补充可提高Nrf2的抗氧化能力及减轻移植肝缺血再灌注损伤，证明Brg1补充成为一种新的防治移植肝损伤方法；在临床肝移植手术中，观察丙泊酚后处理对移植术后肝功能恢复和预后的影响；并且阐述Brg1与丙泊酚后处理的关系，证明Brg1表达增强是丙泊酚后处理保护作用主要药理机制。本项目首次将染色质重塑因子Brg1与Nrf2、抗氧化酶信号通路结合，探讨这一信号通路在移植肝缺血再灌注损伤的作用和机制，将为开发新的多环节干预措施提供理论依据。