本文選題：促紅細胞生成素衍生肽 + 心肌微血管內(nèi)皮細胞��； 參考：《中國循環(huán)雜志》2015年03期

【摘要】：目的:探討促紅細胞生成素衍生肽(HBSP)抑制大鼠心肌微血管內(nèi)皮細胞(CMECs)缺血/再灌注損傷的作用及其機制。方法:分離培養(yǎng)大鼠CMECs,建立缺血/再灌注損傷模型,隨機分為對照組、模擬缺血/再灌注組(SI/R組)、SI/R+重組人促紅細胞生成素組(SI/R+rh EPO組)、SI/R+HBSP組;HBSP分別選擇2.5 ng/ml、25 ng/ml、50 ng/ml、100 ng/ml四個濃度,通過噻唑藍(MTT)比色實驗檢測CMECs增殖能力以確定藥物作用的最適濃度。而后采用細胞劃痕實驗檢測細胞遷移能力,末端脫氧核苷酸轉(zhuǎn)移酶介導的d UTP缺口末端標記測定(TUNEL)法檢測細胞凋亡率來進一步驗證其保護作用。研究保護機制的實驗分組為:對照組、SI/R組、SI/R+HBSP(HBSP最適濃度為25 ng/ml)組、SI/R+HBSP+磷脂酰肌醇-3激酶(PI3K)抑制劑(LY294002)組、SI/R+HBSP+雷帕霉素組,采用蛋白免疫印跡(Western blot)法檢測蛋白激酶B(AKT)、哺乳動物雷帕霉素靶蛋白(m TOR)、核糖體S6蛋白激酶(p70S6K)等蛋白的表達及其磷酸化水平。結(jié)果:與SI/R組比,SI/R+rh EPO組和SI/R+HBSP組CMECs增殖能力明顯上升(P0.05),凋亡率顯著下降(P0.01),遷移能力增強。用抑制劑LY294002及雷帕霉素處理后,與SI/R組相比,SI/R+HBSP組AKT、m TOR及p70S6K的磷酸化水平均顯著提高(P0.05);與SI/R+HBSP組相比,SI/R+HBSP+LY294002組的AKT、m TOR及p70S6K的磷酸化水平均顯著下降(P0.05),SI/R+HBSP+雷帕霉素組m TOR磷酸化水平及p70S6K磷酸化水平顯著下降(P0.05)。結(jié)論:HBSP能夠抑制缺血/再灌注誘導的CMECs的損傷,其保護作用可能與PI3K-AKT/mT OR信號通路激活有關(guān)。
[Abstract]:Aim: to investigate the inhibitory effect of erythropoietin derived peptide (HBSP) on myocardial microvascular endothelial cells (CMECs) ischemia / reperfusion injury in rats and its mechanism. Methods: CMECs were isolated and cultured in rats. The model of ischemia / reperfusion injury was established in rats. The rats were randomly divided into four groups: the control group, the simulated ischemia / reperfusion group and the SI-R / R recombinant human erythropoietin group. The rats in the SIP-R / R HBSP group were divided into four concentrations of 2.5 ng / ml 25 ng / ml 50 ng / ml 100 ng/ml, respectively. The optimal concentration of CMECs was determined by MTT colorimetric assay. Then the cell migration ability was detected by cell scratch assay and the apoptosis rate was detected by terminal deoxynucleotidyl transferase-mediated d UTP Nick end labeling method to further verify its protective effect. The experimental groups to study the protective mechanism were as follows: the optimal concentration of SIR / R HBSP(HBSP in the control group was 25 ng / ml), and the SIR HBSP phosphatidylinositol 3 kinase PI3K inhibitor, LY294002, was used in the SIP-R HBSP rapamycin group. The expression and phosphorylation level of protein kinase (BK), mammalian rapamycin target protein (mTORA) and ribosomal S6 protein kinase (p70S6K) were detected by Western blot assay. Results: compared with SI/R group, the proliferative ability of CMECs in SIR rh EPO group and SI/R HBSP group was significantly increased (P 0.05), the apoptosis rate was decreased significantly (P 0.01), and the migration ability was enhanced. After treated with inhibitor LY294002 and rapamycin, Compared with SI/R group, the phosphorylation levels of p70S6K and p70S6K in Sirs / Sirs HBSP group were significantly higher than those in SI/R HBSP group, and the phosphorylation levels of TOR and p70S6K in Sirs / R HBSP LY294002 group were significantly lower than those in SI/R HBSP group, and the phosphorylation levels of m TOR and p70S6K in Sirs / Sirs HBSP rapamycin group were significantly lower than those in SI/R HBSP group. Conclusion CMECs injury induced by ischemia / reperfusion can be inhibited by 1: HBSP, and its protective effect may be related to the activation of PI3K-AKT/mT OR signaling pathway.
【作者單位】： 中國人民解放軍第四軍醫(yī)大學第一附屬醫(yī)院心內(nèi)科;
【基金】：國家自然科學基金(81200100)
【分類號】：R965

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本文編號：1910394