本文關(guān)鍵詞： 心肌 缺血再灌注損傷 鈉泵 內(nèi)質(zhì)網(wǎng)應激 細胞凋亡　出處：《皖南醫(yī)學院》2017年碩士論文　論文類型：學位論文

【摘要】：目的:在細胞水平上,探討鈉泵α2亞基對心肌缺血再灌注(ischemia reperfusion,IR)損傷的影響并進一步探討鈉泵信號通路是否通過內(nèi)質(zhì)網(wǎng)應激反應參與心肌缺血再灌注損傷。方法:原代心肌細胞培養(yǎng)成功后,在細胞水平上構(gòu)建缺氧復氧模型。缺氧復氧過程中,對各組采用不同的干預處理。實驗分四組:(1)鈉鉀泵α2亞基siRNA+缺氧復氧組(Na+-K+pumpα2siRNA+I/R)(2)陰性病毒+缺氧復氧組(control+I/R),(3)正常對照組(4)缺氧復氧組(I/R)。缺氧復氧完成后通過熒光顯微鏡觀察細胞生長情況,熒光定量PCR檢測各組鈉鉀泵α2亞基m RNA的表達;流式細胞儀,采用Annexin V-FITC/PI雙染法檢測細胞凋亡;采用Western blot檢測內(nèi)質(zhì)網(wǎng)應激指標及信號分子GRP-78以及促凋亡蛋白CHOP表達水平。結(jié)果:1.熒光定量PCR檢測鈉鉀泵α2亞基mRNA:與對照組相比,I/R組與control-I/R組細胞α2亞基mRNA含量降低(P0.05);與I/R組相比,鈉鉀泵α2si RNA-I/R組細胞m RNA明顯含量降低(P0.05)2.流式細胞儀檢測細胞凋亡率:與對照組相比,I/R組與control-I/R組細胞的凋亡率明顯增高(P0.05),與I/R組相比,鈉鉀泵α2siRNA-I/R組細胞凋亡率亦明顯增高(P0.05)3.western-bolt結(jié)果:與對照組相比,control-I/R組細胞內(nèi)CHOP及GRP-78的蛋白表達量明顯增高(P0.05);與I/R組相比,鈉鉀泵α2siRNA-I/R組細胞CHOP及GRP-78的蛋白表達量亦明顯增高(P0.05)。結(jié)論:1、抑制鈉泵α2亞基的活性會加重缺血再灌注細胞內(nèi)質(zhì)網(wǎng)應激、加重心肌細胞凋亡。2、鈉泵α2亞基可能通過誘導內(nèi)質(zhì)網(wǎng)應激參與心肌細胞凋亡并參與心肌缺血再灌注損傷。
[Abstract]:Objective: at the cellular level, To investigate the effect of sodium pump 偽 2 subunit on myocardial ischemia reperfusion IRs and whether the sodium pump signaling pathway is involved in myocardial ischemia reperfusion injury through endoplasmic reticulum stress. Methods: primary myocardial cells were cultured successfully. Build a model of anoxia reoxygenation at the cellular level. Each group was treated with different intervention. The experiment was divided into four groups: 1) sodium and potassium pump 偽 2 subunit siRNA anoxic reoxygenation group (Na K pump 偽 2 siRNA I / R 2) negative virus hypoxia reoxygenation group control I / R (3) normal control group 4) hypoxia reoxygenation group (4) hypoxia reoxygenation group (I / R). Microscopic observation of cell growth, Fluorescence quantitative PCR was used to detect the expression of 偽 2 subunit m RNA, flow cytometry was used to detect apoptosis by Annexin V-FITC / Pi double staining. Western blot was used to detect endoplasmic reticulum stress, signal molecule GRP-78 and CHOP expression of apoptotic protein. Results: 1. Fluorescence quantitative PCR was used to detect 偽 2 subunit mRNAs of sodium and potassium pump: compared with control group, mRNA content of 偽 2 subunit in IPRR group and control-I/R group was decreased (P 0.05), compared with that in IR / R group. The content of m RNA in 偽 2si RNA-I/R group decreased significantly. Flow cytometry was used to detect the apoptosis rate. Compared with the control group, the apoptosis rate of the control-I/R group and the I / R group was significantly higher than that of the I / R group, and that of the I / R group was significantly higher than that of the I / R group, and that of the I / R group was significantly higher than that of the control group. The apoptosis rate of 偽 2siRNA-I / R group was also significantly higher than that of the control group. The results showed that the expression of CHOP and GRP-78 protein in the control group was significantly higher than that in the control group (P 0.05), and that in the I / R group was significantly higher than that in the I / R group. In sodium potassium pump 偽 2siRNA-I / R group, the protein expression of CHOP and GRP-78 was also significantly increased (P 0.05). Conclusion the inhibition of the activity of sodium pump 偽 2 subunit may aggravate the endoplasmic reticulum stress in ischemia-reperfusion cells. By inducing endoplasmic reticulum stress, sodium pump 偽 2 subunit may participate in myocardial apoptosis and myocardial ischemia-reperfusion injury.
【學位授予單位】：皖南醫(yī)學院
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R54

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