Brg1介导的Nrf2/STAT3激活在恢复吸入麻醉药后处理对糖尿病鼠心脏缺血再灌注损伤保护中的作用

Inhalational anesthetics postconditioning (APO) has been found to alleviate myocardial ischemia reperfusion (IR) injury. However there are potential limitations to APO given that diabetes blocks the effectiveness of APO,and the underlying mechanism is unclear. Our preliminary studies find that the loss of diabeteic heart sensitive to isoflurane postconditioning was associated with decreased expression of cardiac Brahma-related gene 1 (Brg1) and that antioxidant mediated partial restored myocardial Brg1 expression led to partial restoration of isoflurane postconditioning cardioprotection in diabetic rats. In response to oxidantive stress, Brg1 interacts and regulates nuclear factor-erythroid 2-related factor 2 (Nrf2) activation, and Nrf2 plays the central role in cardioprotection via activation of the signal transducer and activator of transcription 3 (STAT3), a key protein in postconditioning protection. In diabetic hearts, the protein expression of Brg1, Nrf2 and phosphorylated STAT3 are conincidently diminished, which might be the reason why diabetic hearts loss sensitivity to APO. Therefore, we hypothesize that Brg1 can restore diabetic heart sensitivity to APO. In the current proposal, we aim to determine the effectiveness of Brg1 in attenuating myocardial IR injury and restoring myocardial sensitivity to APO in diabetic rodents and its impact on Nrf2/STAT3 signaling pathway. We will do so by using combination of in vivo animal model, isolated heart model, cell culture and genetic modified mice with adenovirus transfection and siRNA technologies to confirm our hypothesis. The proposed studies will improve our understanding of the pathophysiology of this phenomenon and may help facilitate the development of novel and optimal therapies in diabetic heart disease.

吸入麻醉药后处理（APO）在心肌缺血再灌注（IR）损伤中起重要保护作用。然而，APO对糖尿病心脏IR损伤保护作用消失，其机制不清楚。申请者前期研究发现糖尿病鼠对异氟烷后处理失敏感的同时伴随心肌中Brg1表达降低，而抗氧化治疗部分恢复糖尿病心肌Brg1表达同时也部分恢复异氟烷后处理对缺血心肌保护作用。鉴于氧化应激时Brg1招募是Nrf2活化所必需，Nrf2活化可激活后处理发挥心肌保护作用的关键蛋白STAT3，而糖尿病心肌Nrf2及STAT3活性降低，我们推测Brg1缺失引起的Brf2/STAT3失活是APO对糖尿病心肌失敏感的关键原因。本课题拟应用Brg1、Nrf2基因敲除鼠建立糖尿病及心肌IR损伤模型及心肌细胞缺氧复氧模型，结合腺病毒转染及siRNA技术，探讨Brg1在恢复糖尿病心脏对APO敏感性中的作用及信号转导机制，为药物治疗结合APO防治糖尿病患者心脏IR损伤提供理论及实验依据。

吸入麻醉药后处理（APO）在心肌缺血再灌注（IR）损伤中起重要保护作用。然而，APO对糖尿病心脏IR损伤保护作用消失，其分子机制不清楚。前期研究发现糖尿病鼠对异氟烷后处理失敏感的同时伴随心肌中Brg1表达降低，而抗氧化治疗部分恢复糖尿病心肌Brg1表达同时也部分恢复异氟烷后处理对缺血心肌保护作用。鉴于氧化应激时Brg1招募是Nrf2活化所必需，Nrf2活化可激活后处理发挥心肌保护作用的关键蛋白STAT3，而糖尿病心肌Nrf2及STAT3活性降低，我们推测Brg1缺失引起的Brf2/STAT3失活是APO对糖尿病心肌失敏感的关键原因。本研究中我们发现，1型糖尿病（T1DM）大鼠心肌IR损伤高于正常对照组，并伴随着氧化损伤加重、炎症因子TNF-alpha及IL-6表达升高以及心肌Brg1、HO-1及p-STAT3表达降低。而抗氧化剂治疗可以显著减轻糖尿病心肌IR损伤及炎症因子表达，并升高心肌Brg1、HO-1及p-STAT3表达水平。七氟醚后处理不能显著降低T1DM大鼠梗死面积及血浆CK-MB和cTnI水平。但是七氟醚后处理与抗氧化剂NAC合用后可以增强NAC对糖尿病大鼠的心肌保护作用。同时，我们还观察到七氟醚后处理与NAC合用可以显著增加STAT3活性，并降低Fox1的表达。T1DM小鼠心肌IR损伤加重，伴随着降低心肌核Nrf2、HO-1、Brg1蛋白表达。七氟醚后处理可减轻正常小鼠心肌IR损伤，并恢复心肌核Nrf2、HO-1、Brg1蛋白表达，但是对于糖尿病小鼠无效。在心肌细胞（H9C2细胞系）缺氧复氧模型上，我们发现七氟醚后处理可显著降低细胞损伤和氧化应激，同时伴随增加的HO-1及核Nrf2蛋白表达，而对高糖处理后的细胞无效。并且HO-1抑制剂取消了七氟醚后处理对心肌的保护作用。我们的研究结果表明：七氟醚后处理通过激活Nrf2/Brg1/HO-1信号通路发挥减轻心肌缺血再灌注损伤的作用，但是七氟醚后处理对糖尿病心肌无效，其分子机制可能与糖尿病Nrf2/Brg1/HO-1信号通路受损有关。本研究结果从Nrf2/Brg1信号通路角度阐明了七氟醚后处理对糖尿病心脏失敏感性的分子机制，并首次提出七氟醚后处理与抗氧化剂NAC合用可恢复七氟醚后处理对糖尿病心脏的敏感性，该研究为药物治疗结合APO防治糖尿病患者心脏IR损伤提供理论及实验依据。