大鼠肺動脈平滑肌細胞Rho激酶對這p-Smad1核遷移的作用及機制
發(fā)布時間：2018-10-24 17:26
【摘要】：目的 揭示Rho激酶對p-Smad1核遷移的作用及信號轉(zhuǎn)導(dǎo)途徑,探討它們在肺血管重構(gòu)中的作用機制。 材料與方法 分離培養(yǎng)大鼠遠端肺動脈平滑肌細胞,應(yīng)用PDGF-BB啟動肺動脈平滑肌細胞Rho激酶信號通路,應(yīng)用BMP-2啟動BMP信號通路,并用Rho激酶抑制劑Y-27632、MEK抑制劑-U0126進行干預(yù)。培養(yǎng)細胞分成五組：(1)對照組；(2)BMP-2組；(3)BMP-2+PDGF-BB組；(4)BMP-2+PDGF-BB+Y-27632組；(5)BMP-2+PDGF-BB+U0126組。免疫熒光染色標(biāo)記p-Smad1在細胞核內(nèi)外的分布并計算p-Smad1核遷移陽性細胞百分數(shù)(即核遷移率)。分離平滑肌細胞核蛋白和胞漿蛋白,western blotting分析p-Smad1在細胞內(nèi)的總量和細胞核內(nèi)外相對含量的變化。Cell Counting Kit-WST-8試劑盒檢測細胞增殖。 結(jié)果 BMP-2組p-Smad1在細胞內(nèi)的總量以及在細胞核中的相對含量和核遷移率明顯高于對照組(p0.05); BMP-2+PDGF-BB組p-Smad1的核遷移率和在細胞核中的相對含量明顯低于BMP-2組(p0.05),但是細胞內(nèi)p-Smad1總量無明顯改變(p0.05); BMP-2+PDGF-BB+ Y-27632組和BMP-2+PDGF-BB+U0126組細胞內(nèi)p-Smad1的總量以及在細胞核中的相對含量和核遷移率與BMP-2組基本相似(p0.05)。BMP-2組,OD值明顯小于對照組(p0.05); PDGF-BB+BMP-2組,OD值明顯大于BMP-2組(p0.05)且大于對照組(p0.05); BMP-2+PDGF-BB+Y-27632組和BMP-2+PDGF-BB+U0126組OD值均小于PDGF-BB+BMP-2組(P0.05)。 結(jié)論 在大鼠肺動脈平滑肌細胞,PDGF-BB激活的Rho激酶通過MEK/ERK1/2抑制BMP-2引起的p-Smad1的核遷移,促進平滑肌細胞增殖,進而引起肺血管重構(gòu)。
[Abstract]:Aim to investigate the effect of Rho kinase on nuclear migration of p-Smad1 and its signal transduction pathway, and to explore the mechanism of its role in pulmonary vascular remodeling. Materials and methods Rat pulmonary artery smooth muscle cells were isolated and cultured. PDGF-BB was used to activate the Rho kinase signaling pathway in pulmonary smooth muscle cells, and BMP-2 was used to initiate the BMP signal pathway. Rho kinase inhibitor Y-27632 was used to intervene with U0126. The cultured cells were divided into five groups: (1) control group; (2) BMP-2 group; (3) BMP-2 PDGF-BB group; (4) BMP-2 PDGF-BB Y-27632 group; (5) BMP-2 PDGF-BB U0126 group. The distribution of p-Smad1 in and out of the nucleus was labeled with immunofluorescence staining and the percentage of positive cells (i.e. nuclear mobility) of p-Smad1 nuclear migration was calculated. Nuclear protein and Cytoplasmic protein, western blotting were isolated from smooth muscle cells (SMC). The total amount of p-Smad1 in the cell and the relative content of p-Smad1 in and out of the cell nucleus were analyzed by. Cell Counting Kit-WST-8 kit to detect cell proliferation. Results the total amount of p-Smad1 in the cells, the relative content and nuclear mobility in the nucleus of BMP-2 group were significantly higher than those in the control group (p0.05), the nuclear mobility and relative content of p-Smad1 in the BMP-2 PDGF-BB group were significantly lower than those in the BMP-2 group (p0.05), but the relative content of p-Smad1 in the nucleus in the BMP-2 PDGF-BB group was significantly lower than that in the BMP-2 group (p0.05). The total amount of intracellular p-Smad1 in BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was similar to that in BMP-2 group (p0.05). The OD value of BMP-2 group was significantly lower than that of control group (p0.05), and that of PDGF-BB BMP-2 group was significantly lower than that of PDGF-BB BMP-2 group, while that of BMP-2 PDGF-BB U0126 group was significantly lower than that of BMP-2 group (p0.05), while that of BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was similar to BMP-2 group (p0.05). The OD value of BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was lower than that of PDGF-BB BMP-2 group (P0.05). Conclusion in rat pulmonary artery smooth muscle cells, Rho kinase activated by PDGF-BB inhibits the nuclear migration of p-Smad1 induced by BMP-2 through MEK/ERK1/2, promotes the proliferation of smooth muscle cells and leads to pulmonary vascular remodeling.
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R363
本文編號：2292052
[Abstract]:Aim to investigate the effect of Rho kinase on nuclear migration of p-Smad1 and its signal transduction pathway, and to explore the mechanism of its role in pulmonary vascular remodeling. Materials and methods Rat pulmonary artery smooth muscle cells were isolated and cultured. PDGF-BB was used to activate the Rho kinase signaling pathway in pulmonary smooth muscle cells, and BMP-2 was used to initiate the BMP signal pathway. Rho kinase inhibitor Y-27632 was used to intervene with U0126. The cultured cells were divided into five groups: (1) control group; (2) BMP-2 group; (3) BMP-2 PDGF-BB group; (4) BMP-2 PDGF-BB Y-27632 group; (5) BMP-2 PDGF-BB U0126 group. The distribution of p-Smad1 in and out of the nucleus was labeled with immunofluorescence staining and the percentage of positive cells (i.e. nuclear mobility) of p-Smad1 nuclear migration was calculated. Nuclear protein and Cytoplasmic protein, western blotting were isolated from smooth muscle cells (SMC). The total amount of p-Smad1 in the cell and the relative content of p-Smad1 in and out of the cell nucleus were analyzed by. Cell Counting Kit-WST-8 kit to detect cell proliferation. Results the total amount of p-Smad1 in the cells, the relative content and nuclear mobility in the nucleus of BMP-2 group were significantly higher than those in the control group (p0.05), the nuclear mobility and relative content of p-Smad1 in the BMP-2 PDGF-BB group were significantly lower than those in the BMP-2 group (p0.05), but the relative content of p-Smad1 in the nucleus in the BMP-2 PDGF-BB group was significantly lower than that in the BMP-2 group (p0.05). The total amount of intracellular p-Smad1 in BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was similar to that in BMP-2 group (p0.05). The OD value of BMP-2 group was significantly lower than that of control group (p0.05), and that of PDGF-BB BMP-2 group was significantly lower than that of PDGF-BB BMP-2 group, while that of BMP-2 PDGF-BB U0126 group was significantly lower than that of BMP-2 group (p0.05), while that of BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was similar to BMP-2 group (p0.05). The OD value of BMP-2 PDGF-BB Y-27632 group and BMP-2 PDGF-BB U0126 group was lower than that of PDGF-BB BMP-2 group (P0.05). Conclusion in rat pulmonary artery smooth muscle cells, Rho kinase activated by PDGF-BB inhibits the nuclear migration of p-Smad1 induced by BMP-2 through MEK/ERK1/2, promotes the proliferation of smooth muscle cells and leads to pulmonary vascular remodeling.
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2011
【分類號】：R363
【共引文獻】
相關(guān)期刊論文 前6條
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相關(guān)博士學(xué)位論文 前2條
1 李建安;肺動脈灌注低溫HTK液對未成熟肺體外循環(huán)損傷作用的臨床研究[D];北京協(xié)和醫(yī)學(xué)院;2010年
2 靳建軍;辛伐他汀對大鼠急性肺栓塞的保護作用及其機制的研究[D];北京協(xié)和醫(yī)學(xué)院;2011年
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