E Lusha, Cheng Ying, Zhao Xingsheng
Department of Cardiology, the Inner Mongolia Autonomous Region People's Hospital, Hohhot 010017, Inner Mongolia, China (E LS, Zhao XS); Department of Clinical Laboratory, the Second Hospital of Tianjin Medical University, Tianjin 300201, China (Cheng Y). Corresponding author: Zhao Xingsheng, Email:
Zhonghua Wei Zhong Bing Ji Jiu Yi Xue. 2018 Aug;30(8):795-799. doi: 10.3760/cma.j.issn.2095-4352.2018.08.016.
To investigate the protective effect of high-density lipoprotein (HDL) on the mice cardiac myocytes induced by oxygen and glucose deprivation (OGD).
Cardiac cells of primary scavenger receptor-B1 knockout mice (SR-B1) and normal C57 mice (SR-B1) were obtained by protease digestion and differential adhesion method. (1) The two kinds of cells were divided into normal control group (Con group), OGD group, OGD+HDL group. Propidium iodide (PI) staining were used to determine the necrosis of cardiac myocytes. (2) SR-B1 cardiac cells were divided into Con group, OGD group, OGD+HDL group, and phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) inhibitor LY294002 group. PI staining were used to determine the necrosis of cardiac myocytes. TUNEL staining was used to determine the cell apoptosis. The kit was used to determine the contents of MB isoenzyme of creatine kinase (CK-MB) and lactate dehydrogenase (LDH) in the culture medium supernatant. The expressions of SR-B1 and Akt protein were determined by Western Blot.
(1) In SR-B1 cardiomyocytes, HDL could inhibit cell necrosis induced by OGD. There was no protective effect of HDL on OGD in the SR-B1 cardiomyocytes. (2) The study of SR-B1 cells was showed that compared with Con group, necrotic cells were significantly increased and cell activity were significantly decreased, the cell viability were significantly decreased, the contents of LDH and CK-MB in supernatant were significantly increased, the expressions of phosphorylated Akt (p-Akt) and SR-B1 were significantly decreased in OGD group. Compared with OGD group, the number of necrotic cells in the OGD+HDL group was significantly decreased [PI positive cells rate: (26.71±5.94)% vs. (64.24±18.34)%], the cell activity was significantly increased [(63.84±6.95)% vs. (26.71±5.13)%], the contents of LDH and CK-MB in supernatant were significantly decreased [LDH (U/L): 896.3±161.5 vs. 1 568.3±243.5, CK-MB (U/L): 304.3±72.9 vs. 583.6±81.6], the expressions of p-Akt and SR-B1 were significantly increased (p-Akt/t-Akt: 0.84±0.13 vs. 0.18±0.06, SR-B1/β-actin: 1.23±0.19 vs. 0.09±0.02), with statistically significant differences (all P < 0.05). Compared with OGD+HDL group, necrotic cells in LY294002 group were increased, cell activity was decreased, LDH and CK-MB contents in supernatant were increased, p-Akt and SR-B1 expressions were decreased; there was no statistical difference between LY294002 group and OGD group. There was no significant difference in cell apoptosis among the 4 groups.
HDL has protective effect on the mice myocardial cells. The mechanism may be related with the up regulation of the expression of SR-B1 protein by the activation of PI3K/Akt pathway.
探讨高密度脂蛋白（HDL）对氧糖剥夺（OGD）诱导的小鼠心肌细胞的保护作用。
采用蛋白酶消化法和差速贴壁法获取原代清道夫受体-B1基因敲除小鼠（SR-B1）和正常C57小鼠（SR-B1）的心肌细胞。（1）将两种细胞分为正常对照组（Con组）、OGD组、OGD+HDL组。采用碘化丙啶（PI）染色检测心肌细胞坏死情况。（2）将SR-B1心肌细胞分为Con组、OGD组、OGD+HDL组、磷脂酰肌醇3激酶/蛋白激酶B（PI3K/Akt）抑制剂LY294002组。采用PI染色检测心肌细胞坏死情况，采用TUNEL染色检测细胞凋亡情况。采用试剂盒检测培养基上清液中肌酸激酶同工酶MB（CK-MB）和乳酸脱氢酶（LDH）含量。采用蛋白质免疫印迹法检测SR-B1和Akt蛋白表达。
（1）在SR-B1心肌细胞中，HDL可抑制OGD诱导的细胞坏死。在SR-B1心肌细胞中HDL对OGD无保护作用。（2）SR-B1细胞研究结果显示，与Con组比较，OGD组坏死细胞明显增多，细胞活性明显降低，细胞存活率明显降低，上清液中LDH和CK-MB含量明显升高，磷酸化Akt（p-Akt）和SR-B1表达明显降低。与OGD组比较，OGD+HDL组坏死细胞数量明显减少[PI阳性细胞率：（26.71±5.94）%比（64.24±18.34）%]，细胞活性明显升高[（63.84±6.95）%比（26.71±5.13）%]，上清液中LDH和CK-MB含量明显降低[LDH（U/L）：896.3±161.5比1 568.3±243.5，CK-MB（U/L）：304.3±72.9比583.6±81.6]，p-Akt和SR-B1表达明显升高（p-Akt/t-Akt：0.84±0.13比0.18±0.06，SR-B1/β-肌动蛋白：1.23±0.19比0.09±0.02），差异均有统计学意义（均P<0.05）。与OGD+HDL组比较，LY294002组坏死细胞增多，细胞活性降低，上清液中LDH和CK-MB含量升高，p-Akt和SR-B1表达降低；LY294002组与OGD组比较差异无统计学意义。4组细胞凋亡情况差异无统计学意义。
HDL对小鼠心肌细胞具有保护作用。其机制可能与通过激活PI3K/Akt通路上调SR-B1蛋白表达有关。
