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目的 观察抗小鼠 Caspase-12小干扰 RNA(siRNA)对小鼠原代肝细胞凋亡的抑制作用。方 法 原位二步灌流法分离并培养 Balb/c 小鼠原代肝细胞，化学合成制备三种抗 Caspase-12不同位点(130， 214，521)的 siRNA，脂质体包裹转染小鼠原代肝细胞。毒胡萝卜素4μmol/L 诱导肝细胞凋亡。逆转录聚 合酶链反应、western blot 检测抑制效果；四甲基偶氮唑盐法检测对细胞凋亡的影响。结果 在浓度为 100 nmol/L 时，siRNA(130)对小鼠原代肝细胞 Caspase-12 mRNA 的抑制率为0，siRNA(214)为52．08%， siRNA(521)为30．73%(t=4．30，P＜0．05)；在浓度为200 nmol/L 时，三种 siRNA 的抑制率分别为88．07%、 86．22%，89．41%。200 nmol/L 的 siRNA(214)对凋亡细胞 procaspase-12的抑制率为51．43%(t=4．30， P＜0．01)；MTT 比色实验发现，凋亡细胞活力增高48．76%(t=2．23，P＜0．01)。结论 抗 Caspase-12 siRNA 对小鼠原代肝细胞凋亡有一定的抑制作用。
Objective To observe the inhibitory effect of anti-mouse Caspase-12 small interfering RNA (siRNA) on mouse primary hepatocyte apoptosis. METHODS: Primary Balb / c mouse hepatocytes were isolated and cultured by in situ two-step perfusion. Three siRNAs against different sites of Caspase-12 (130,214,521) were prepared by chemical synthesis. The liposomes were transfected into mice Primary hepatocytes. Induction of hepatocyte apoptosis by thapsigargin at 4 μmol / L. Reverse transcription polymerase chain reaction, Western blot detection of inhibition; tetramethylzirconium salt assay on apoptosis. Results The inhibitory rate of siRNA (130) on Caspase-12 mRNA of mouse primary hepatocytes was 0 at the concentration of 100 nmol / L, that of siRNA (214) was 52.08% and that of siRNA (521) was 30.73% (t = 4.30, P <0.05). The inhibitory rates of the three siRNAs were 88.07%, 86.22% and 89.41% at the concentration of 200 nmol / L, respectively. The inhibitory ratio of 200 nmol / L siRNA (214) to procaspase-12 was 51.43% (t = 4.30, P <0.01). MTT colorimetric assay showed that the viability of apoptotic cells increased 48 .76% (t = 2.23, P <0.01). Conclusion Anti-Caspase-12 siRNA can inhibit the primary hepatocyte apoptosis in mice.