黏菌素诱导细胞凋亡和自噬分子机制及靶向调控研究

发布时间：2018-05-30 12:58

  本文选题：黏菌素 + 凋亡　； 参考：《中国农业大学》2017年博士论文

【摘要】：多黏菌素(包括多黏菌素B及黏菌素)是临床治疗多重耐药革兰氏阴性细菌感染的最为重要抗生素之一。多黏菌素肾毒性及神经毒性是其临床使用过程中主要毒副作用及剂量限制性因素,但其分子毒性机制尚不清楚。本文通过建立小鼠及体外细胞模型系统研究了黏菌素诱导细胞凋亡及自噬的分子机制,并进一步探究了靶向调控细胞凋亡及自噬信号通路对黏菌素肾毒性及神经毒性的影响。主要发现如下:(1)使用小鼠及体外细胞模型(HEK293及N2a细胞系)研究证实,黏菌素可通过剂量依赖性诱导氧化应激介导的线粒体功能失调,激活线粒体、死亡受体及内质网凋亡通路;同时激活NF-κB、MAPKs、p53、Nrf2/ARE、Akt等信号通路。其中,ERK、JNK信号通路的激活在黏菌素诱导的细胞凋亡中发挥保护性作用,P53的激活及Akt信号通路的抑制发挥促凋亡的作用。(2)黏菌素暴露可上调组织及细胞内Beclin1表达及LC3Ⅱ/Ⅰ比率,伴随明显的自噬-溶酶体及自噬体结构,表明黏菌素处理激活细胞自噬,通过使用GFP-LC3质粒转染后进一步证实。进一步使用N2a细胞模型研究证实,自噬抑制剂氯喹(CQ)依赖于ROS介导的氧化应激损伤及casapse激活,加剧黏菌素诱导的细胞凋亡。雷帕霉素,可特异性的抑制mTOR/p70s6k信号通路,上调Akt/CREB、Nrf2/ARE等信号通路信号通路及ULK1依赖性的细胞自噬及其降解过程,改善黏菌素诱导的氧化应激损伤、线粒体功能失调及caspase激活介导的细胞凋亡;小鼠原代神经元及动物模型进一步验证了自噬的保护性作用。(3)姜黄素及黄芩素补充能够提高动物体内及细胞抗氧化、抗炎能力,上调Nrf2/HO-1信号通路及ULK1介导的细胞自噬,下调黏菌素受体蛋白Megalin表达、p53蛋白表达,显著改善黏菌素对组织/细胞造成的的氧化应激损伤、caspase依赖性凋亡及NF-κB信号通路介导的炎症反应,从而改善黏菌素诱导的神经毒性及肾毒性。(4)体外及动物模型研究发现,抗生素药物米诺环素能够通过清除细胞内ROS,抑制氧化应激、caspase激活及细胞凋亡,有效降低黏菌素诱导的神经毒性及肾毒性,具有重要的临床应用价值。综上所述,黏菌素诱导的神经毒性及肾毒性涉及外源性及内源性细胞凋亡通、ULK1介导的细胞自噬及NF-κB介导的炎症反应。药理性激活Akt/CREB、Nrf2/HO-1及自噬可通过抑制氧化损伤及细胞凋亡能够有效降低黏菌素肾毒性及神经毒性。本研究为新一代多黏菌素脂肽类抗生素的开发、多黏菌素联合用药、减毒策略提供了重要的理论参考依据。
[Abstract]:Polymyxin (including polymyxin B and myxin) is one of the most important antibiotics for the treatment of multidrug resistant gram-negative bacterial infections. The nephrotoxicity and neurotoxicity of polymyxin are the main side effects and dose limiting factors in clinical use, but the molecular toxicity mechanism is not clear. The molecular mechanism of myxin induced apoptosis and autophagy was studied by establishing mouse and in vitro cell models, and the effects of targeted regulation of apoptosis and autophagy signaling pathway on myxin nephrotoxicity and neurotoxicity were further explored. The main findings were as follows: (1) using mouse and in vitro cell models (HEK293 and N2a cell lines), it was demonstrated that myxin could induce mitochondrial dysfunction mediated by oxidative stress in a dose-dependent manner and activate mitochondria, death receptors and endoplasmic reticulum (ER) apoptosis pathway. At the same time, the signal pathway of NF- 魏 B mapksmapk p53 Nrf 2 / AREN Akt was activated. The activation of JNK signaling pathway in ERK plays a protective role in myxin-induced apoptosis. The activation of p53 and the inhibition of Akt signaling pathway play a role in promoting apoptosis.) Myxin exposure can up-regulate the expression of Beclin1 and the ratio of LC3 鈪，

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