线粒体移植抑制异烟肼肝毒性的机制研究

Isoniazide (INH) is the first-line anti-tuberculosis drug, but its clinical effect is severely affected by hepatotoxicity. In spite of widespread attention for mitochondrial toxicity of INH-induced hepatotoxicity in the recent years, the mechanisms by which INH impairs the balance of mitochondrial homeostasis and how to treat INH-induced hepatotoxicity effectively are still unclear. Mitochondrial transplantation is a method that normal mitochondria are delivered to cells with damaged mitochondria to recover mitochondrial homeostasis. Aiming at the toxic effects of INH on liver mitochondrial homeostasis, we choose the signaling pathways that play importance roles in mitochondrial homeostasis, mitochondrial biogenesis and degradation PGC1α and PINK1/Parkin signaling pathways as well as mitochondrial fission and fusion DRP1 and MFN1/2-OPA1 signaling pathways. On the base of interpretation of inhibitory effect of mitochondrial transplantation on INH-induced hepatotoxicity, we analyze the differences and internal connections between PGC1α and PINK1/Parkin signaling pathways as well as DRP1 and MFN1/2-OPA1 signaling pathways before and after mitochondrial transplantation to clarify the relationship between the balance of mitochondrial biogenesis and degradation and the balance of mitochondrial fission and fusion in protection against INH-induced hepatotoxicity. The findings will provide experimental evidence for the clinical application of mitochondrial transplantation for treatment of INH-induced hepatotoxicity.

异烟肼（INH）是一线抗结核药物，但在治疗中出现的肝毒性严重影响了其临床治疗效果。近年来，虽然INH线粒体毒性诱发的肝毒性受到广泛关注，但对其破坏肝线粒体稳态的机制和如何有效治疗肝毒性仍有待阐明。线粒体移植是通过向线粒体受损的细胞中传递正常线粒体来维持线粒体稳态的方法。本课题针对INH对肝线粒体稳态的毒性作用，选取在维持线粒体稳态中起关键作用的线粒体新生降解平衡PGC1α和PINK1/Parkin调控通路以及线粒体分裂融合平衡DRP1和MFN1/2-OPA1调控通路，在证实线粒体移植抑制INH肝毒性的基础上，通过分析线粒体移植前后INH作用下肝细胞PGC1α和PINK1/Parkin通路以及DRP1和MFN1/2-OPA1通路的变化差异和内在联系，揭示线粒体新生降解平衡与线粒体分裂融合平衡在线粒体移植发挥抑制INH肝毒性作用中的相互关系，为线粒体移植用于临床治疗INH肝毒性提供实验依据。

异烟肼（INH）的肝毒性是临床上亟待解决的问题。INH可通过破坏线粒体诱发肝毒性，而线粒体移植可通过外源性补充线粒体修复细胞内受损线粒体。本部分研究聚焦维持线粒体稳态的线粒体新生降解平衡和线粒体分裂融合平衡的关键调控通路，探究线粒体移植对INH肝毒性的保护作用。体外研究发现INH通过下调PGC1α通路，抑制线粒体新生；通过促进自噬，促进线粒体降解，诱发线粒体新生降解失衡。同时，INH通过影响DRP1和MFN2通路，促进线粒体碎裂，诱发线粒体分裂融合失衡。这导致线粒体三羧酸循环抑制、线粒体呼吸功能下降、线粒体膜电位下调、ATP产生减少以及对细胞基础水平ROS抑制，最终诱发线粒体稳态失衡，激活线粒体凋亡通路诱发肝细胞凋亡。线粒体移植虽然减弱INH对肝细胞三羧酸循环的抑制，但是对INH所致线粒体膜电位下调和基础水平ROS抑制并没有恢复作用，同时线粒体移植也没有抑制INH所致小鼠血浆AST上升、肝细胞浊肿和肝纤维化，这表明线粒体移植对INH所致肝毒性没有抑制作用。因此，在聚焦线粒体稳态的前提下，对基金内容做了如下两部分调整。. 氧化铁纳米粒子是生物医学领域最具研发前景的纳米材料之一，然而其免疫系统毒性限制了其临床应用。研究发现PEG-COOH包裹的Fe3O4（PEG-Fe3O4）纳米粒子通过激活PGC1α，促进线粒体新生；通过下调自噬，抑制线粒体降解；通过促进DRP1和MFN2表达，促进线粒体碎裂这三方面破坏树突状细胞（DC）线粒体稳态。并导致葡聚糖摄取以及CD80、CD86和CCR7表达下调，表明PEG-Fe3O4纳米粒子破坏了DC功能性不成熟状态。自噬抑制剂3-MA通过促进PGC1α/MFN2介导的线粒体新生和融合之间的协同作用，抑制PEG-Fe3O4对DC毒性作用。本研究为减弱PEG-Fe3O4纳米粒子的免疫毒性，促进其临床应用提供了研究基础。. Nampt是线粒体复合物I作用底物NADH氧化形式NAD+的限速酶，同时Nampt也是线粒体氧化磷酸化中连接三羧酸循环和呼吸链的关键辅酶，在维持线粒体功能中发挥重要作用。研究发现肝特敲Nampt能够加重高脂饮食所致小鼠肝毒性，同时伴随血脂升高。鉴于前面研究发现异烟肼通过破坏线粒体稳态诱发肝损伤，本部分研究为今后以肝Nampt作为靶点，探究其可能通过维持线粒体稳态，防治异烟肼肝毒性提供了新思路。