CAJ | 학술논문

目的：验证构建成功的携基质金属蛋白酶组织抑制因子-1（TIMP-1）基因特异性小干扰RNA的慢病毒载体对目的基因表达的抑制效应。方法以慢病毒质粒感染HSC-T6细胞，在感染6天和8天后，观察慢病毒感染效率；采用定量PCR法检测TIMP-1 mRNA水平变化；采用免疫印迹法检测TIMP-1蛋白表达变化。结果在慢病毒载体感染6天时，TIMP-1 mRNA水平较正常对照组下降约0.668倍[2-ΔΔCt为（0.668±0.046）]，下降程度明显高于阴性对照组[2-ΔΔCt为（1.001±0.041），（P<0.05）]；在慢病毒感染6天和8天时，RNAi组对TIMP-1蛋白表达具有明显的抑制作用，且以感染第6天为显著。结论构建成功的携TIMP-1基因特异性RNAi慢病毒载体能有效感染HSC-T6细胞，并抑制目的基因的表达。
목적：험증구건성공적휴기질금속단백매조직억제인자-1（TIMP-1）기인특이성소간우RNA적만병독재체대목적기인표체적억제효응。방법이만병독질립감염HSC-T6세포，재감염6천화8천후，관찰만병독감염효솔；채용정량PCR법검측TIMP-1 mRNA수평변화；채용면역인적법검측TIMP-1단백표체변화。결과재만병독재체감염6천시，TIMP-1 mRNA수평교정상대조조하강약0.668배[2-ΔΔCt위（0.668±0.046）]，하강정도명현고우음성대조조[2-ΔΔCt위（1.001±0.041），（P<0.05）]；재만병독감염6천화8천시，RNAi조대TIMP-1단백표체구유명현적억제작용，차이감염제6천위현저。결론구건성공적휴TIMP-1기인특이성RNAi만병독재체능유효감염HSC-T6세포，병억제목적기인적표체。
Objective To explore the inhibiting effect of constructed lentiviral vector containing tissue in-hibitor of metalloproteinase-1 (TIMP-1)gene-specific small interfering RNA on target gene expression in hepatic stellate cells-T6 (HSC-T6). Methods HSC-T6 cells were infected with lentiviral virus plasmid for 6 to 8 days. Real-time PCR was used to detect the infection efficiency at 6 days;Western blot was performed to detect the protein levels of TIMP-1 at 6 and 8 days. Results TIMP-1 mRNA was reduced to 0.668 time that of the nor-mal control at 6 days[2-△△Ct was (0.668±0.046)],and this decrease was significantly higher than that in the nega-tive control [2-△△Ct was (1.001±0.041),(P<0.05)];Similarly,TIMP-1 protein was significantly reduced at 6 and 8 days after viral infection. Conclusion Constructed lentiviral vector containing TIMP-1 gene-specific small inter-fering RNA can effectively infect rat HSC-T6 cells and reduce the target gene expression.