摘要
目的观察RNA干扰技术（RNAi）是否能有效抑制兔OA模型膝关节软骨中基质金属蛋白酶13（MMP13）的表达。方法体外化学合成MMP13序列特异性双链RNA与脂质体混合后转染OA模型中软骨细胞，实验分四组，特异性RNAi组：特异性RNAi＋Lipofectamine2000，非特异RNAi组：非特异RNAi＋Lipofectamine2000，脂质体组：Lipofectamine2000，对照组：无血清DMEM。用逆转录．聚合酶链反应（RT—PCR）和免疫组织化学法检测软骨细胞中的MMP13的表达。噻唑蓝（MTT）比色法检测各组的软骨细胞生长情况。结果RT—PCR显示RNAi组较对照组、非特异性组和脂质体组的MMP13基因的PCR扩增条带明显减弱，免疫组织化学法显示RNAi组中MMP13阳性细胞数比对照组明显减少。与对照组比较，非特异性组、RNAi组、脂质体组软骨细胞的存活率显著降低（P〈0．05），但在3组间两两差异无统计学意义。结论使用RNA干扰技术可有效抑制免OA模型中膝关节软骨细胞MMP13的表达，RNA干扰技术为基因治疗OA提供了新策略。
Objective To explore whether RNA interference （RNAi） could suppress the expression of MMP13 in the chondrocytes of rats with osteoarthritis （OA）. Methods Chondrocytes in OA models were transfected using in vitro chemically-synthesized MMP13 sequence-specific double-stranded RNA formulated with liposome. The experiments were conducted in 4 groups ： group A （ control group）, serumfree DMEM ; group B （ RNAi group ）, RNAi and Lipfectamine2000 ; group C （ non-distinctive group ） , non- specificity dsRNA and Lipfectamine2000; group D （liposome group）, Lipofectamine2000. The expression of MMP13 in chondrocytes was detected by RT-PCR and immunohistochemistry. The proliferation of chrondrocytes was assessed by MTT assay. Results The expression of MMP13 mRNA was decreased markedly in group B as compared with group A, group C and group D. Immunohistochemistry showed the significant decreases of MMP13 in group B as compared with group. MTT results revealed that the growth of chrondrocytes was suppressed in all 4 groups, but there was no significant difference among the groups A, B and C （P 〉 0.05）. Conclusion RNAi was able to effectively suppress the expression of MMP13 in chrondrocytes of rabbit OA models.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2008年第11期1502-1504,共3页
Chinese Journal of Experimental Surgery
