miR-34a/Sirt1/FoxO1信号通路介导的自噬在脂肪乳剂解救布比卡因心脏毒性中的机制研究

Local anesthetics, represented by bupivacaine, are widely applied in clinical, with its severe complication of cardiac arrest if inadvertently used. Previous studies demonstrated that lipid emulsion effectively rescue bupivacaine-induced cardiac arrest, but the mechanism of survival condition improved by lipid emulsion after heart re-beat was not clear. Autophagy plays an important role in cardiac stress. And the miR-34a/Sirt1/FoxO1 signaling pathway also plays a key role in the mediation of autophagy, that has not reported in bupivacaine-induced cardiotoxiciy at home and abroad. In pilot experiments, we found that bupivacaine significantly inhibits the fusion of autophagosomes and lysosomes in cardiomyocytes, resulting in injury of cardiomyocyte, whereas lipid emulsion significantly improved the inhibiting effect. In order to explore the mechanism, we will research the mechanism of autophagy mediated by miR-34a/Sirt1/FoxO1 signaling pathway in lipid emulsion rescuing bupivacaine cardiotoxicity, and the effect of resveratrol that is Sirt1 agonist in rescuing bupivacaine cardiotoxicity, through in vitro and in vivo experiments, in animal, cellular and molecular level. In this project, we will elucidate the new mechanism of lipid emulsion in the rescuing of bupivacaine cardiotoxicity, that will provide evidence for the treatment of local anesthetic cardiotoxicity.

布比卡因为代表的局麻药在临床应用广泛，使用不慎容易导致心脏骤停。前期研究证实脂肪乳剂有效复苏布比卡因导致的心脏骤停，但复苏后生存情况改善的机制不明。自噬在心脏应激中扮演重要的角色，miR-34a/Sirt1/FoxO1信号通路对自噬调控起关键的作用，然而目前国内外有关自噬在布比卡因心脏毒性中的研究未见报道。我们通过预实验发现，布比卡因明显抑制心肌细胞的自噬体与溶酶体的融合，造成心肌细胞损伤，而脂肪乳剂显著改善其抑制作用。为探讨其机制，本项目通过体内实验和体外实验，从整体、细胞、分子水平研究miR-34a/Sirt1/FoxO1信号通路介导的自噬在脂肪乳剂解救布比卡因心脏毒性中的作用机制，以及Sirt1激动剂白藜芦醇在布比卡因心脏毒性解救中的作用，阐明脂肪乳剂解救布比卡因心脏毒性的新机制，为局麻药心脏毒性的救治提供依据。

局麻药布比卡因可引起心脏毒性，我们的前期研究证明应用脂肪乳剂可以加快心肌细胞自噬流，解救布比卡因中毒，但具体机制并未阐明。此项研究的重要结果:(1)脂肪乳剂通过促进自噬流，减少自噬体堆积，从而解救布比卡因的心肌毒性。(2)在心肌细胞中，miR-34a与SIRT1 3’-UTR结合可下调SIRT1的表达 (3)脂肪乳剂可下调布比卡因处理后心肌细胞中miR-34a的表达，使SIRT1的表达增加，从而促进自噬体的清除；在心肌细胞中过表达miR-34a则会逆转脂肪乳剂的作用。（4）使用SIRT1抑制剂以后，脂肪乳剂促进心肌细胞自噬的作用也被逆转了。综上所述，脂肪乳剂能改善布比卡因引起的心肌细胞自噬体堆积，miR-34a/SIRT1通路参与其中。本研究项目基本阐明了脂肪乳剂解救布比卡因心脏毒性的作用机制，其中miR-34a/SIRT1/自噬通路机制为解决局麻药中毒的心脏毒性提供了新的研究方向，有利于进一步增加局麻药中毒的解救成功率，以及最终提高临床区域麻醉的安全性。