Zheng Ting, Zheng Chun-Ying, Zheng Xiao-Chun, Zhao Ruo-Guang, Chen Yan-Qing
Fujian Provincial Clinical College, Medical College, Fujian Medical University, Fuzhou, China.
Department of Anaesthesiology, Fujian Provincial Hospital, Fujian Medical University, Fuzhou, China.
Clin Exp Pharmacol Physiol. 2017 May;44(5):586-594. doi: 10.1111/1440-1681.12730.
Ropivacaine is one of the most common but toxic local anaesthetics, and the mechanisms underlying its neurotoxicity are still largely unknown. This study was conducted to prepare a ropivacaine-induced neuronal injury model and research the effects of ropivacaine on PARP-1 activation and nicotinamide adenine dinucleotide (NAD) depletion. The cell death and apoptosis of ropivacaine-induced SH-SY5Y cells were detected with flow cytometry. The lactate dehydrogenase cycling reaction measured the NAD level, and western blots were used to analyze the expression levels of PARP-1 and apoptosis-inducing factor (AIF) after ropivacaine treatments with different concentrations and durations. A PARP-1 inhibitor (PJ-34) was used to confirm the relationship between PARP-1 activation and NAD depletion. Hoechst 33258 nuclear staining and a mitochondrial membrane potential (Δψm) assay were used to detect the role of exogenous NAD in ropivacaine-induced neuronal injury. Ropivacaine-induced SH-SY5Y cell death and apoptosis, PARP-1 activation, and AIF increase as well as intracellular NAD depletion occurred in a time- and concentration-dependent manner (P<.05). PARP-1 activation led to NAD depletion (P<.05). Exogenous NAD impaired ropivacaine-induced nuclear injury (P<.05). Ropivacaine treatment induced PARP-1 activation and NAD depletion (P<.05). Parthanatos (PARP-1-dependent cell death) was definitely involved in ropivacaine-induced neuronal injury, and exogenous NAD may be a novel therapeutic method for parthanatos-dependent neuronal injury.
罗哌卡因是最常见但有毒性的局部麻醉剂之一，其神经毒性的潜在机制在很大程度上仍不清楚。本研究旨在制备罗哌卡因诱导的神经元损伤模型，并研究罗哌卡因对聚（ADP-核糖）聚合酶-1（PARP-1）激活和烟酰胺腺嘌呤二核苷酸（NAD）消耗的影响。采用流式细胞术检测罗哌卡因诱导的SH-SY5Y细胞的死亡和凋亡。乳酸脱氢酶循环反应测定NAD水平，并用蛋白质免疫印迹法分析不同浓度和作用时间的罗哌卡因处理后PARP-1和凋亡诱导因子（AIF）的表达水平。使用PARP-1抑制剂（PJ-34）来确认PARP-1激活与NAD消耗之间的关系。采用Hoechst 33258核染色和线粒体膜电位（Δψm）检测法来检测外源性NAD在罗哌卡因诱导的神经元损伤中的作用。罗哌卡因诱导的SH-SY5Y细胞死亡和凋亡、PARP-1激活、AIF增加以及细胞内NAD消耗呈时间和浓度依赖性（P<0.05）。PARP-1激活导致NAD消耗（P<0.05）。外源性NAD减轻了罗哌卡因诱导的核损伤（P<0.05）。罗哌卡因处理诱导PARP-1激活和NAD消耗（P<0.05）。PARP-1依赖性细胞死亡（parthanatos）肯定参与了罗哌卡因诱导的神经元损伤过程，外源性NAD可能是一种针对PARP-1依赖性神经元损伤的新型治疗方法。
