【摘要】：目的:探討絲裂霉素C(MMC)調(diào)控mi R-200b表達(dá)誘導(dǎo)人成纖維細(xì)胞凋亡的潛在機(jī)制。方法:體外培養(yǎng)人硬膜外瘢痕組織中提取的成纖維細(xì)胞,用不同濃度的MMC處理后,采用Real-time PCR檢測(cè)mi R-200b表達(dá)變化,TUNEL染色分析成纖維細(xì)胞凋亡情況,Western blot檢測(cè)凋亡相關(guān)蛋白cleaved-PARP、Bax、Bcl-2表達(dá)的變化,亞硫酸氫鹽(BSP)法測(cè)定mi R-200b啟動(dòng)子區(qū)域甲基化程度的改變。結(jié)果:Real-time PCR結(jié)果顯示MMC處理后,成纖維細(xì)胞中mi R-200b表達(dá)量顯著下降,差異具有統(tǒng)計(jì)學(xué)意義(P0.05),TUNEL染色結(jié)果表明MMC能夠顯著地誘導(dǎo)人成纖維細(xì)胞的凋亡,差異具有統(tǒng)計(jì)學(xué)意義(P0.05)。Western blot結(jié)果顯示MMC能夠升高cleaved-PARP、Bax表達(dá),降低Bcl-2表達(dá)。MMC處理后mi R-200b啟動(dòng)子區(qū)域甲基化程度發(fā)生了變化,隨著藥物濃度的增加,甲基化程度不斷增加。結(jié)論:MMC通過下調(diào)mi R-200b誘導(dǎo)人成纖維細(xì)胞凋亡,可能是通過促進(jìn)其啟動(dòng)子區(qū)甲基化實(shí)現(xiàn)的。
[Abstract]:Aim: to investigate the potential mechanism of mitomycin C (MMC) regulating the expression of mi R-200b in inducing apoptosis of human fibroblasts. Methods: fibroblasts extracted from human epidural scar tissue were cultured in vitro. After treated with different concentrations of MMC, the expression of mi R-200b was detected by Real-time PCR, and the apoptosis of fibroblasts was analyzed by TUNEL staining. The expression of apoptosis-related protein (cleaved-PARP,Bax,Bcl-2) was detected by Western blot, and the methylation degree of mi R-200b promoter was determined by (BSP) method. Results: Real-time PCR results showed that the expression of mi R-200b in fibroblasts decreased significantly after MMC treatment, and the difference was statistically significant (P0.05), TUNEL staining showed that MMC could significantly induce apoptosis of human fibroblasts. The difference was statistically significant (P0.05). Western blot results showed that MMC could increase the expression of cleaved-PARP,Bax and decrease the expression of Bcl-2. The methylation degree of mi R-200b promoter region changed with the increase of drug concentration after MMC treatment. The degree of methylation is increasing. Conclusion: MMC induces apoptosis of human fibroblasts by down-regulating mi R-200b, which may be achieved by promoting methylation of its promoter region.
【作者單位】： 揚(yáng)州大學(xué)臨床醫(yī)學(xué)院(蘇北人民醫(yī)院)骨科;
【基金】：國家自然科學(xué)基金青年項(xiàng)目(編號(hào):81301550) 江蘇省六大人才高峰資助項(xiàng)目(編號(hào):2015-WSN-108)
【分類號(hào)】：R68

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本文編號(hào)：2303796