Abstract:

Objective To investigate whether mesenchymal stem cells lead to induction of autophagy and change at early apoptotic rate under hypoxia condition on coronary artery endothelial cells. Methods We constructed the co?culture model of HCAECs（human coronary artery endothelial cells） and hBMMSCs（human bone marrow?derived mesenchymal stem cells）. In experiment group，HCAEC co?cultured with MSC，HCAEC cultured alone in blank group. And in control group ，HCAEC cultured alone using same medium with experiment group. Each group separately cultured 24 h under nomoxia （5% CO2 ，95% air）and hypoxia（1% O2 ，5% CO2 ，94% N2）condition. The HCAEC′s expression of LC3 and Beclin?1 at mRNA and protein level was detected by Real?Time PCR and Western blotting ， respectively. Flow cytometry was used to detect early apoptotic rate. Results The protein expression of LC3 and Beclin?1 in co?culture groups and hypoxia groups significantly higher than separate culture groups and nomoxia groups respectively（P<0.05），while mRNA expression of LC3 and Beclin?1 in each group shows no difference （P>0.05）. The early apoptotic rate of co?culture group under hypoxia was 8.86%±1.48%，which was significantly lower than those in separate culture group under hypoxia and control group under hypoxia（15.23%±2.56% and 14.49%±3.31% ，P<0.05）. Conclusion MSC induces autophagy in HCAEC，along with decrease of early apoptosis rate of HCAEC under hypoxia condition.

Key words: apoptosis, autophagy, bone marrow?derived mesenchymal stem cells , hypoxia, human coronary artery endothelial cells, co?culture

摘要：

目的 探索缺氧状况下MSC对内皮细胞自噬的诱导以及对早期凋亡率的影响。方法 构建HCAEC与MSC间接共培养模型，实验分组：共培养组：MSC与HCAEC共培养；单独培养组：HCAEC单独培养；对照组：HCAEC使用与共培养组相同的培养基进行单独培养。上述各组分别予常氧（5% CO2，95%空气）和缺氧（1% O2，5% CO2，94% N2）培养24 h，Westernblotting检测各组HCAEC细胞LC3蛋白和Beclin?1蛋白的表达量，RT?qPCR检测各组HCAEC细胞LC3基因和Beclin?1基因的表达水平，流式细胞术检测各组HCAEC细胞早期凋亡率。结果 共培养组与单独培养组相比、以及缺氧组与常氧组相比，LC3蛋白和Beclin?1蛋白表达量均有升高（P<0.05）。各组间LC3基因和Beclin?1基因的表达水平没有检测到明显差异（P>0.05）。缺氧组中共培养组的早期凋亡率为（8.86%±1.48%），明显低于单独培养组（15.23%±2.56%）和对照组（14.49%±3.31%）（P<0.05）。结论 MSC可以诱导HCAEC产生自噬，并在缺氧状况下改善HCAEC的生存状况，减少早期凋亡率。

关键词: 冠状动脉内皮细胞, 自噬, 缺氧, 骨髓间充质干细胞, 共培养, 凋亡