本文選題：miR-c　切入點(diǎn)：Ras基因家族成員A　出處：《中國醫(yī)科大學(xué)學(xué)報(bào)》2016年12期

【摘要】：目的研究miR-200c調(diào)控Ras基因家族成員A(RhoA)表達(dá)介導(dǎo)RMP7增加血腫瘤屏障(BTB)通透性的機(jī)制。方法應(yīng)用real-time PCR檢測RMP7作用BTB后人腦微血管內(nèi)皮細(xì)(ECs)miR-200c的表達(dá);應(yīng)用miR-200c模擬物和miR-200c抑制物分別轉(zhuǎn)染GECs(ECs和U87腦膠質(zhì)瘤細(xì)胞共培養(yǎng)的細(xì)胞),測量跨內(nèi)皮阻抗值(TEER)、辣根過氧化物酶(HRP)滲漏量,分析BTB通透性;應(yīng)用Western blotting檢測RhoA的表達(dá);應(yīng)用免疫熒光方法觀察GECs中RhoA表達(dá)和分布;應(yīng)用雙熒光素酶報(bào)告基因檢測miR-200c轉(zhuǎn)錄后水平調(diào)控RhoA機(jī)制。結(jié)果 RMP7作用GECs后,使GECs內(nèi)源性miR-200c表達(dá)顯著降低;miR-200c模擬物和miR-200c抑制物成功轉(zhuǎn)染到GECs中;miR-200c模擬物顯著抑制RMP7誘導(dǎo)TEER值的降低、HRP的升高;miR-200c模擬物顯著減少RhoA的表達(dá),促使RhoA在GECs細(xì)胞質(zhì)和細(xì)胞核分布減少;miR-200c模擬物轉(zhuǎn)錄后水平負(fù)性調(diào)控RhoA基因表達(dá)。miR-200c抑制物與miR-200c模擬物實(shí)驗(yàn)結(jié)果相反。結(jié)論 miR-200c轉(zhuǎn)錄后水平負(fù)性調(diào)控RhoA表達(dá),介導(dǎo)RMP7增加血腫瘤屏障通透性機(jī)制。
[Abstract]:Objective to investigate the mechanism of miR-200c regulating the expression of Ras gene family member, Agnia RhoA, and mediate the increase of RMP7 permeability of BBB. Methods real-time PCR was used to detect the expression of ECsmmiR-200c in human brain microvascular endothelial cells after RMP7 treatment with BTB. MiR-200c mimics and miR-200c inhibitors were used to transfect GECs(ECs and U87 glioma cells, respectively, to measure the transendothelial impedance and horseradish peroxidase (HRP) leakage, to analyze the permeability of BTB, and to detect the expression of RhoA by Western blotting. The expression and distribution of RhoA in GECs were observed by immunofluorescence method, and the RhoA mechanism was detected by double luciferase reporter gene. The expression of endogenous miR-200c in GECs was significantly decreased after transfection of miR-200c inhibitor and mimic of miR-200c into GECs. MiR-200c mimics significantly inhibited the increase of TEER value induced by RMP7. The mimic of RMP7 R-200c significantly reduced the expression of RhoA. The expression of RhoA gene was negatively regulated by RhoA at the posttranscriptional level of GECs mimics. MiR-200c inhibitor was contrary to that of miR-200c mimics. Conclusion miR-200c posttranscriptional level negatively regulates RhoA expression. Mediates RMP7 to increase the permeability of blood tumor barrier.
【作者單位】： 中國醫(yī)科大學(xué)基礎(chǔ)醫(yī)學(xué)院神經(jīng)生物學(xué)教研室;
【基金】：國家自然科學(xué)基金(81101918;81673028)
【分類號】：R363

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