Conclusion
In cultured primary rat hippocampal neurons, caspase-1 activation plays a role in bilirubin-induced neurotoxicity, and VX-765 treatment provides protection against bilirubin-induced neuronal injury by inhibiting caspase-1 activation. 目的: 研究半胱氨酸天冬氨酸蛋白酶-1 (caspase-1)活化及VX-765在胆红素诱导大鼠海马神经元损伤中的作用。 方法: 原代培养大鼠海马神经元分为胆红素组、对照组、VX-765组; 胆红素组给予胆红素(50 μmol/L), 对照组给予同体积药物溶剂二甲基亚砜(DMSO), VX-765组在胆红素干预前1 h给予VX-765 (50 μmol/L); Western blot检测NLRP3、活化caspase-1表达, 改良MTT、乳酸脱氢酶(LDH)释放率及台盼蓝染色检测细胞相对存活率及死亡率, ELISA法检测原代培养上清IL-18水平。 结果: 胆红素诱导原代培养大鼠海马神经元3、6 h, NLRP3、活化caspase-1蛋白表达明显高于对照组(P < 0.05)。VX-765干预后, 活化caspase-1表达与胆红素组相比明显降低(P < 0.05)。分组干预细胞24 h, VX-765组的相对存活率为(84.020±2.311) %, 明显高于胆红素组(P < 0.05), 低于对照组(P < 0.05); VX-765组LDH释放率为(10.780±1.577) %, 明显低于胆红素组(P < 0.05), 高于对照组(P < 0.05); VX-765组台盼蓝染色阳性率为(5.580±1.234) %, 明显低于胆红素组(P < 0.05), 高于对照组(P < 0.05)。 结论: 在原代培养的大鼠海马神经元中, caspase-1活化参与胆红素神经毒性的发生, VX-765抑制其活化发挥神经保护作用。
Methods
Cultured primary rat hippocampal neurons were exposed to DMSO (control group), 50 µmol/L bilirubin, or 50 µmol/L bilirubin 1 h after 50 µmol/L VX-765 treatment. The expressions of NLRP3 and caspase-1 in the neurons were detected by Western blotting, and the relative cell survival and death rates were assessed with a modified MTT assay, lactate dehydrogenase assay and Typan blue staining. Interleukin-18 (IL-18) concentration in the culture supernatant was measured using enzyme-linked immunosorbent assay (ELISA).
Objective
To investigate the role of caspase-1 activation in bilirubin-induced neuronal injury and the protective effect of VX-765 against bilirubin-induced neurotoxicity in cultured primary rat hippocampal neurons.
Results
In cultured primary rat hippocampal neurons, bilirubin exposure for 3 and 6 h caused significant increases in the expressions of NLRP3 and activated caspase-1 compared with those in the control group (P<0.05). Pretreatment of the cells with VX-765 obviously suppressed bilirubin-induced activation of caspase-1 (P<0.05). The relative survival rate of the neurons was (84.02∓2.31)% in VX-765 intervention group, significantly higher than that in bilirubin group (P<0.05) but lower than that in the control group (P<0.05); LDH release rate in VX-765 intervention group was (10.78∓1.58)%, significantly lower than that in bilirubin group (P<0.05) but higher than that in the control group (P<0.05). The cell death rate in VX-765 intervention group was (5.58∓1.23)%, significantly lower than that in bilirubin group (P<0.05) but higher than that in the control group (P<0.05).