本文關(guān)鍵詞：高密度脂蛋白拮抗氧化型低密度脂蛋白誘導(dǎo)的血管平滑肌細(xì)胞中胱硫醚γ裂解酶表達(dá)的下調(diào)　出處：《福建醫(yī)科大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 高密度脂蛋白 氧化型低密度脂蛋白 胱硫醚γ-裂解酶 血紅素加氧酶-1

【摘要】：目的:高密度脂蛋白(High density lipoprotein,HDL)拮抗氧化型低密度脂蛋白(Oxidized low density lipoprotein,ox-LDL)誘導(dǎo)的氧化應(yīng)激在保護(hù)血管內(nèi)皮細(xì)胞和平滑肌細(xì)胞中發(fā)揮重要作用。本實(shí)驗(yàn)探究了ox-LDL和HDL對(duì)血管平滑肌細(xì)胞(Vascular smooth muscle cell,VSMC)中氣體信號(hào)分子硫化氫(Hydrogen sulfide,H2S)催化酶胱硫醚γ-裂解酶(Cystathionine gamma-lyase,CSE)表達(dá)的影響,以及HDL能否通過拮抗氧化應(yīng)激來抑制ox-LDL對(duì)于CSE表達(dá)的調(diào)節(jié)作用及其中參與的機(jī)制。方法:大鼠主動(dòng)脈血管平滑肌細(xì)胞在無菌環(huán)境下分離培養(yǎng)。從健康志愿者中提取的血漿經(jīng)超速離心分離HDL和LDL,ox-LDL由硫酸銅(Cu SO4)氧化而得。CSE、血紅素加氧酶1(heme oxygenase 1,HO-1)蛋白表達(dá)和ERK1/2、AKT、p38/MAPK的磷酸激活利用免疫印跡法檢測,CSE m RNA表達(dá)用實(shí)時(shí)熒光定量PCR法測定。細(xì)胞內(nèi)活性氧(Reactive oxygen species,ROS)釋放用ROS介導(dǎo)的活性氧檢測探針(DCFH-DA)檢測。用亞甲基藍(lán)顯色測吸光度法測定H2S生成。RNAi技術(shù)沉默VSMC中HO-1的表達(dá)。結(jié)果:Ox-LDL能夠呈時(shí)間和濃度依賴性地下調(diào)VSMC中CSE蛋白和m RNA表達(dá)。HDL不僅能直接促進(jìn)VSMC中CSE蛋白表達(dá)和H2S釋放,還能拮抗ox-LDL對(duì)CSE的下調(diào)作用。HDL能誘導(dǎo)HO-1生成并拮抗由ox-LDL誘導(dǎo)的胞內(nèi)ROS釋放。HDL誘導(dǎo)ERK、AKT、p38/MAPK磷酸化,ERK1/2、AKT、和p38/MAPK抑制劑能夠減弱HDL誘導(dǎo)的HO-1表達(dá)并削弱對(duì)ox-LDL誘導(dǎo)的ROS釋放的清除能力。ERK1/2、AKT、和p38/MAPK抑制劑能夠削弱HDL誘導(dǎo)的CSE蛋白表達(dá)及HDL對(duì)ox-LDL誘導(dǎo)的CSE表達(dá)的拮抗作用。干擾VSMC中HO-1的表達(dá),HDL對(duì)CSE表達(dá)的誘導(dǎo)作用減弱。結(jié)論:HDL能通過誘導(dǎo)HO-1表達(dá)拮抗ox-LDL誘導(dǎo)的VSMC中ROS釋放和CSE表達(dá)下調(diào),HDL還可直接誘導(dǎo)CSE蛋白表達(dá)和H2S的釋放,且ERK1/2、AKT、p38MAPK信號(hào)通路參與其中。
[Abstract]:Objective: to study the high density lipoprotein of high density lipoprotein. Hd) antagonized oxidized low density lipoprotein (low density lipoprotein). Oxidative stress induced by ox-LDL plays an important role in the protection of vascular endothelial cells and smooth muscle cells. In this study, we investigated the effects of ox-LDL and HDL on vascular smooth muscle cells (VSMC). Vascular smooth muscle cell. Gas signal molecule hydrogen sulfide Hydrogen sulfide. The effect of cystathionine gamma-lyase (CSE) on the expression of cystathionine 緯 -lyase catalyzed by H _ 2S was observed. And whether HDL can inhibit the regulatory effect of ox-LDL on the expression of CSE and its mechanism by antagonizing oxidative stress. Methods:. Rat aortic vascular smooth muscle cells were isolated and cultured in aseptic environment. HDL and LDL were isolated from plasma from healthy volunteers by ultracentrifugation. Ox-LDL was oxidized by CuSO4) to obtain. CSE, heme oxygenase 1 (HO-1) protein expression and ERK1/2. The phosphoric acid activation of AKT p38 / MAPK was detected by Western blotting. The expression of CSE m RNA was assayed by real-time fluorescence quantitative PCR. Intracellular reactive oxygen species was detected by reactive oxygen species. Ros) release using ROS mediated active oxygen species detection probe DCFH-DAA. Detection of HO-1 in VSMC by methylene blue colorimetric assay. Results:. Ox-LDL can down-regulate the expression of CSE protein and m RNA in VSMC in a time-and concentration-dependent manner. HDL-C can not only directly promote the expression of CSE protein and H2S release in VSMC. It could also antagonize the down-regulation of CSE by ox-LDL. HDL could induce HO-1 production and antagonize the intracellular ROS release induced by ox-LDL. HDL-induced ERKK AKT. P38 / MAPK phosphorylated ERK1 / 2 AKT. P38 / MAPK inhibitor could attenuate the expression of HO-1 induced by HDL and weaken the clearance of ROS release induced by ox-LDL. P38 / MAPK inhibitor could weaken the expression of CSE protein induced by HDL and the antagonism of HDL to CSE expression induced by ox-LDL, and interfere with the expression of HO-1 in VSMC. Conclusion HDL can antagonize the release of ROS and the down-regulation of CSE expression in VSMC induced by ox-LDL by inducing HO-1 expression. HDL can also directly induce the expression of CSE protein and the release of H2S, and ERK1 / 2 AK p38 MAPK signaling pathway is involved.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R541.75

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