【摘要】：目的探討Rho相關(guān)卷曲蛋白激酶1(Rho-associated coiled-coil containing protein kinase 1,ROCK1)及其相關(guān)信號(hào)分子在感受張應(yīng)變機(jī)械刺激、調(diào)控血管平滑肌細(xì)胞(vascular smooth muscle cells,VSMCs)增殖功能中的作用。方法應(yīng)用張應(yīng)變加載系統(tǒng)對(duì)體外培養(yǎng)VSMCs施加牽張幅度10%、頻率1.25 Hz生理性周向張應(yīng)變;Brdu檢測(cè)VSMCs增殖水平;Western blotting檢測(cè)力學(xué)加載后VSMCs的ROCK1表達(dá)水平以及蛋白激酶C(protein kinase C,PKC)α/βII、蛋白激酶D(protein kinase D,PKD)、胞外信號(hào)調(diào)節(jié)激酶(extracellular regulated protein kinase,ERK)磷酸化水平;采用RNA干擾技術(shù)(RNA interference,RNAi)檢測(cè)ROCK1對(duì)VSMC增殖和PKCα/βII、PKD、ERK磷酸化的調(diào)控作用。結(jié)果 10%生理性張應(yīng)變加載12、24 h顯著抑制VSMCs的ROCK1表達(dá),并顯著抑制PKD和ERK的磷酸化;10%生理性張應(yīng)變加載12 h顯著抑制PKCα/βⅡ的磷酸化,但加載24 h PKCα/βⅡ的磷酸化與靜止對(duì)照組相比無顯著差異。RNAi抑制VSMCs的ROCK1表達(dá)后,VSMCs增殖水平顯著降低,同時(shí)PKCα/βⅡ和PKD磷酸化水平顯著降低,但ERK磷酸化無明顯變化。結(jié)論 10%生理性張應(yīng)變可能通過抑制ROCK1表達(dá)調(diào)控PKCα/βⅡ和PKD的磷酸化水平,從而影響VSMCs增殖,維持血管穩(wěn)定性。探討張應(yīng)變力學(xué)刺激調(diào)控血管細(xì)胞功能的細(xì)胞內(nèi)信號(hào)轉(zhuǎn)導(dǎo)網(wǎng)絡(luò),對(duì)心血管生理和疾病病理機(jī)制研究具有一定意義。
[Abstract]:Objective to investigate the role of Rho associated crimp protein kinase 1 (Rho-associated coiled-coil containing protein kinase 1 / ROCK1) and its associated signaling molecules in the proliferation of vascular smooth muscle cells (VSMCs) by sensing tensile strain mechanical stimulation and regulating the proliferation of VSMCs. Methods tension strain loading system was applied to VSMCs cultured in vitro with strain amplitude of 10 and frequency 1.25 Hz physiologically circumferential tensile strain. Brdu was used to detect VSMCs proliferation level; Western blotting was used to detect ROCK1 expression of VSMCs after mechanical loading and protein kinase C (protein kinase CpPKC) 偽 / 尾 II, eggs were detected by Brdu. The phosphorylation level of white kinase (D (protein kinase) and extracellular signal-regulated kinase (extracellular regulated protein kinase,ERK); The effects of ROCK1 on VSMC proliferation and PKC 偽 / 尾 II,PKD,ERK phosphorylation were detected by RNA interference technique (RNA interference,RNAi). Results the ROCK1 expression of VSMCs and the phosphorylation of PKD and ERK were significantly inhibited by 10% physiological strain loading for 12h, and the phosphorylation of PKC 偽 / 尾 鈪，

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