异丙酚对布比卡因致PC12细胞毒性时细胞Ca^2＋浓度和一氧化氮合酶活性的影响

Effects of propofol on cytotoxicity of bupivacaine： intracellular Ca^2＋ concentration and nitric oxide synthase activity in PC12 cells exposed to bupivacaine

目的评价异丙酚对布比卡因致PCI2细胞毒性时细胞Ca^2＋浓度和一氧化氮合酶（NOS）活性的影响。方法PC12细胞悬液（10^5／ml）随机分成4组：对照组（C组）、异丙酚组（P组）、布比卡因组（B组）和异丙酚＋布比卡因组（PB组），每组PC12细胞分别接种于36孔板（每孔1ml，每组9孔）、激光共聚焦显微镜专用培养皿（每皿1ml，每组6皿）和24孔板（每孔1ml，每组6孔）。C组加入D．Hank液500μl；P组加入异丙酚至终浓度为2mmol／L；B组加入布比卡因至终浓度为0．09mmol／L；PB组同时加入异丙酚和布比卡因，终浓度分别为2mmol／L和0．09mmol／L。于36孔板中孵育24h后测定PC12细胞凋亡率；于激光共聚焦显微镜专用培养皿中孵育6、24h时，测定PCI2细胞游离Ca^2＋浓度；于24孔板中孵育6、24h时测定细胞NOS活性。结果与C组相比，B组和PB组PC12细胞游离Ca^2＋浓度、NOS活性和凋亡率均升高（P〈0．01），P组上述指标差异无统计学意义（P〉0．05）；与B组相比，PB组PC12细胞游离Ca^2＋浓度、NOS活性和凋亡率均降低（P〈0．05）。结论在细胞水平，异丙酚可能通过抑制NOS活性和钙超载，减轻布比卡因诱导的神经毒性。

Objective To investigate the effects of propofol on intracellular free Ca^2＋ concentration and nitric oxide synthase （NOS） activity in PC12 cells exposed to bupivacaine. Methods The PC12 cells were provided by Shanghai Cell Biology Research Institute, Chinese Academy of Sciences and cultured in DMEM liquid culture medium. The cultured PC12 cells were seeded in 36 well plates and randomly assigned to one of 4 groups （ n = 9 wells each） ： group Ⅰ control （ C ） ; group Ⅱ propofol （P） ; group Ⅲ bupivacaine （ B ） and group iV propofol ＋ bupivacainc （PB）. In control group D-Hank solution was added. The cells were exposed to propofol 2 mmol/L and bupivacaine 0.09 mmol/L in group P and B respectively. In group PB the cells were incubated with propofol 2 mmol/L and bupivacaine 0.09 mmol/L simultaneously. After being incubated for 6 and 24 h the apoptosis in BC12 cells was assessed by flow cytometry. Apoptotic rate was calculated. NOS activity and intracellular free Ca^2＋ concentration in PC12 cells were determined. Results Bupivacaine significantly increased the apoptotic rate of PC12 cells, the intracellnlar free Ca^2＋ concentration and NOS activity in PC12 cells in group B as compared with control group. Propofol significantly decreased the toxic effects of bupivacaine on PC12 cells in group PB compared with group B. Conclusion Bupivacaine is toxic to PC12 cells by increasing apoptosis, intracellular Ca^2＋ concentration and NOS activity in the cells. The toxic effects can be prevented to some extent by concomitant administration of propofol.