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  • Title: [Inhibition of mouse hepatocyte apoptosis by anti-caspase-12 small interfering RNA].
    Author: Liu HF, Xie Q, Jiang S, Li GM, Zhou XQ, Shi Y, Yu H, Jin YX.
    Journal: Zhonghua Gan Zang Bing Za Zhi; 2005 Dec; 13(12):923-6. PubMed ID: 16381641.
    Abstract:
    OBJECTIVES: To study the inhibition of primary mouse hepatocyte apoptosis by small interfering RNA (siRNAs) against caspase-12. METHODS: The Balb/c mouse primary hepatocytes were isolated in situ with two-step liver perfusion with 0.5 g/L collagenase type IV, and apoptosis were induced with 4 micromol/L thapsigargin (TG). The three kingds of siRNAs targeting different gene sites (130, 214, 521) were synthetized chemically. The single-stranded RNAs were annealed to produce double-stranded siRNAs, then the mouse primary hepatocytes were transfected by oligofectamine package. The inhibition of caspase-12 was analyzed with RT-PCR and Western-blot. The viable hepatocytes following the induction of apoptosis were evaluated with MTT. RESULTS: All the three kinds of siRNAs could obviously inhibit normal mouse hepatocyte caspase-12 mRNA. The siRNA (214) were more effective than the other two when the concentration was 100 nmol/L. The caspase-12 mRNA expression was inhibited by 52.08%, while that of siRNA (521) was 30.73% (t=4.30, P <0.05). However when the concentration was 200 nmol/L, the inhibitions were similar (88.07%, 86.22% and 89.41% respectively). siRNA (214) could downregulate the expression of apoptotic hepatocytes procaspase-12 by 51.43% ( t=4.30, P <0.01). Contrasted with apoptotic hepatocytes, the cell activity, which was analyzed with MTT, increased by 48.76% (t=2.23, P <0.01). CONCLUSION: siRNAs could effectively downregulate the expression of caspase-12 at mRNA and protein levels and prevent mouse primary hepatocytes from apoptosis.
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