【摘要】：目的:觀察成骨細(xì)胞不同生長階段caveolin-1表達(dá)的時程變化,并研究caveolin-1對成骨細(xì)胞MC3T3-E1增殖和分化功能的影響。 方法:MC3T3-E1細(xì)胞在含β-甘油磷酸鈉,抗壞血酸的培養(yǎng)基中培養(yǎng)18d,在細(xì)胞培養(yǎng)的不同時間(3,6,9,12,15,18 d) ,采用western blotting方法觀察caveolin-1蛋白表達(dá)變化。采用經(jīng)western blotting法篩選并確證有效的RNA干擾質(zhì)粒pGenesil-1.1- cav-1質(zhì)粒下調(diào)caveolin-1表達(dá)后,觀察其對成骨細(xì)胞MC3T3-E1增殖和分化作用的影響:轉(zhuǎn)染后第3天測定細(xì)胞存活率(MTT法),分別在轉(zhuǎn)染后第3,6,9,12,15d測定細(xì)胞裂解液ALP活性(PNPP法)、培養(yǎng)上清骨鈣素濃度(ELISA)。 結(jié)果:caveolin-1蛋白隨MC3T3-E1細(xì)胞的分化,表達(dá)量逐漸降低(P 0. 01)各處理組對增殖期MC3T3-E1細(xì)胞存活率無明顯影響(P 0. 05)。轉(zhuǎn)染RNA干擾質(zhì)粒pGenesil-1.1- cav-1組,ALP活性和骨鈣素濃度與對照組相比峰值明顯增大,且峰值前移。 結(jié)論:caveolin-1對成骨細(xì)胞MC3T3-E1分化具有負(fù)性調(diào)節(jié)作用,不影響增殖。
[Abstract]:Aim: to observe the temporal changes of caveolin-1 expression in osteoblasts at different growth stages and to study the effects of caveolin-1 on the proliferation and differentiation of MC3T3-E1 in osteoblasts. Methods MC3T3-E1 cells were cultured in a medium containing 尾 -glycerophosphate and ascorbic acid for 18 days, and the expression of caveolin-1 protein was observed by western blotting method at different time of cell culture. The RNA interference plasmid pGenesil-1.1-cav-1 was screened by western blotting method and confirmed to down-regulate the expression of caveolin-1. To observe its effect on the proliferation and differentiation of osteoblasts MC3T3-E1: the cell survival rate (MTT method) was measured on the 3rd day after transfection, the ALP activity of the cell lysate (PNPP method) was measured on the 3rd day after transfection and the osteocalcin concentration (ELISA).) in the culture supernatant was measured respectively. Results with the differentiation of MC3T3-E1 cells, the expression of 1: caveolin-1 protein decreased gradually (P 0. 0. 01) the survival rate of MC3T3-E1 cells in proliferative phase was not significantly affected by each treatment group (P 0. 0. ) The peak value of RNA activity and osteocalcin concentration in RNA interference plasmid pGenesil-1.1 cav-1 group was significantly higher than that in control group, and the peak value moved forward. Conclusion: 1: caveolin-1 has a negative regulatory effect on MC3T3-E1 differentiation of osteoblasts, and does not affect proliferation of osteoblasts.
【學(xué)位授予單位】：南昌大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：R329

【共引文獻(xiàn)】

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