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醫(yī)學(xué)論文范文:辛伐他汀防治心肌細(xì)胞肥大效應(yīng)及其與鈣通道關(guān)系的研究
【摘要】 目的：探討辛伐他汀對心肌細(xì)胞肥大的防治作用及其與鈣通道活動的關(guān)系。方法：采用血管緊張素Ⅱ誘導(dǎo)新生大鼠心肌細(xì)胞肥大模型。應(yīng)用改良Lowry法測定心肌細(xì)胞總蛋白含量。相差顯微鏡測定心肌細(xì)胞表面積。CCK-8法檢測心肌細(xì)胞活力變化。western blot方法檢測心肌細(xì)胞 L-型鈣通道亞單位Cav1.2(α1C)、T-型鈣通道亞單位Cav3.1(α1G)、Cav3.2(α1H)蛋白表達(dá)的變化。應(yīng)用激光共聚焦顯微鏡技術(shù)檢測[Ca2+]i的變化。結(jié)果：(1)辛伐他汀能明顯抑制AngⅡ誘導(dǎo)的心肌細(xì)胞總蛋白含量及細(xì)胞表面積的增加，同時(shí)提高心肌細(xì)胞活力;(2)辛伐他汀能明顯降低心肌細(xì)胞T-型鈣通道α1G、α1H蛋白表達(dá)，但對L-型鈣通道α1C蛋白表達(dá)無明顯影響;(3)辛伐他汀可呈劑量依賴性抑制心肌細(xì)胞肥大所致的鈣離子超載。結(jié)論：辛伐他汀對AngⅡ誘導(dǎo)的心肌細(xì)胞肥大具有明顯的防治作用，其作用機(jī)制可能與辛伐他汀抑制T-型鈣通道α1G、α1H蛋白的重新再表達(dá)有關(guān)，并與其抑制細(xì)胞內(nèi)鈣超載密切相關(guān)。
【關(guān)鍵詞】 心肌細(xì)胞肥大;T-型鈣通道;L-型鈣通道;辛伐他汀
The effects of simastatin on calcium channels in preventing cardiac hypertrophy
GEN Peng，WU Yang，YANG Huichao，et al (Institute of Nautical Medicine, Nantong University, Nantong 226001 )
[Abstract] Objective: To investigate the effects of simvastatin on calcium channels in preventing cardiac hypertrophy.Methods:Total protein content was measured by Lowary,s method and the cell surface area was measured by phase contrast microscope.CCK-8 method was used to observe the viability of myocardiocytes. The expression of α1C、α1G and α1H were detected by western blot. Intracellular Ca2+ were measured with Fluo-3/ AM under confocal laser microscope.Results: The total protein content and cell size of cardiomyocytes increased significantly and the cardiomyocyte viability decreased in AngⅡ treated cells and these effects could be blocked by simvastatin.The protein of α1G and α1H were significantly increased after AngⅡtreatment, which could be inhibited by simvastatin or verpamil hydrochloride. But the protein level of L-type calcium channel α1C was no significant difference between each group.Intracellular calcium overload was remarkably inhibited by simvastatin with a concentration-dependent manner.Conclusions:SIM can inhibit Ang Ⅱ-induced cardiomyocyte hypertrophy, which may be related to the inhibition of the increasing of T-type calcium channel subunit α1G、α1H, and meanwhile to the inhibition of calcium overloading醫(yī)學(xué) 全在.線提供bhskgw.cn.
[Key words] Cardiac hypertrophy; T-type calcium channel; L-type calcium channel; Simvastatin
心肌肥大是心血管疾病的一種常見的并發(fā)癥，是引起猝死、心力衰竭的獨(dú)立危險(xiǎn)因素。心肌肥大不僅是細(xì)胞體積的增大，還涉及心肌細(xì)胞離子通道、離子流及膜電位的變化。近年來隨著細(xì)胞電生理學(xué)和分子生物學(xué)技術(shù)在心血管疾病研究領(lǐng)域的廣泛應(yīng)用，關(guān)于鈣通道在心肌肥大中的作用正逐漸被人們所關(guān)注。近年來有研究發(fā)現(xiàn)[1]他汀類藥物可能具有防治心肌肥大的作用，但其作用機(jī)制的研究則少見報(bào)道。本研究采用血管緊張素Ⅱ(angiotensinⅡ, AngⅡ)誘導(dǎo)心肌細(xì)胞肥大模型，觀察他汀類藥物辛伐他汀(simvastatin,SIM)對心肌細(xì)胞肥大的防治作用，同時(shí)觀察SIM對心肌細(xì)胞L-型鈣通道α1C和T-型鈣通道α1G、α1H各亞單位蛋白表達(dá)的影響，以及心肌細(xì)胞內(nèi)Ca2+的變化，從而深入探討SIM對心肌肥大的防治作用及其機(jī)制。
1 材料和方法
1.1 主要藥物和試劑 SIM (浙江瑞邦藥業(yè)有限公司);AngⅡ(Sigma 公司);鹽酸維拉帕米(Verapamil hydrochloride, Ver)(河北醫(yī)科大學(xué)海森醫(yī)藥有限公司);甲羥戊酸(Mevalonolactone, MVA)(Sigma公司)。
1.2 動物與分組 新生1~3天SD大鼠，由南通大學(xué)實(shí)驗(yàn)動物中心提供。實(shí)驗(yàn)分組：(1)對照組(Control組);(2)AngⅡ組(10-6mol/L);(3)AngⅡ+Ver(10-7mol/L)組;(4)AngⅡ+SIM(10-6mol/L)組，(5)AngⅡ+SIM+MVA(10-4mol/L)組。
1.3 心肌細(xì)胞培養(yǎng) 新生SD大鼠，75%乙醇消毒，取心臟置預(yù)冷的D-Hanks液中，將心室肌切碎約1 mm3、加入10 ml 0.125%胰酶，攪拌消化10 min，棄上清，吹打分散細(xì)胞，孵育8 min。經(jīng)濾網(wǎng)過濾后，接種于100 mm規(guī)格的無菌組織培養(yǎng)瓶中，于5%CO2、高濕度的37 ℃培養(yǎng)箱中放置2 h。孵育后，收集所有培養(yǎng)液，臺盼藍(lán)染色觀察細(xì)胞存活率，細(xì)胞計(jì)數(shù)后將細(xì)胞密度調(diào)整至(5~6)×105 cells/L接種于6孔培養(yǎng)板，每孔2 ml。每孔加入Brdu使其終濃度為0.1 mmol/L。培養(yǎng)48 h后，換無血清培養(yǎng)基，24 h后加入干預(yù)因素。
