miR-378对缺血缺氧条件下骨髓间充质干细胞增殖及凋亡的影响

miR-378 effects on the proliferation and apoptosis of bone marrow mesenchymal stem cells under hypoxic-ischemic condition

背景:研究miR-378对缺血缺氧条件下骨髓间充质干细胞生长增殖、细胞凋亡等生物学行为的影响,为进一步改善骨髓间充质干细胞在梗死心肌中的生存提供新方法。目的:观察miR-378转染后的骨髓间充质干细胞耐受缺血缺氧及促进血管形成的能力。方法:体外培养的骨髓间充质干细胞分为未转染组和转染组,未转染组为未转染miR-378的骨髓间充质干细胞,转染组则采用化学合成的miR-378模拟物转染SD大鼠骨髓间充质干细胞;将两组骨髓间充质干细胞置于缺血缺氧(无血清,体积分数为1%O2、5%CO2,94%N2)环境中培养24 h后,应用锥虫蓝染色计数法、MTS法检测两组细胞在不同时间点的生长及增殖情况,TUNEL法检测细胞凋亡情况;将两组缺血缺氧后的培养基上清液分别刺激人脐静脉内皮细胞,观察血管形成情况。结果与结论:缺血缺氧干预后48 h和72 h,转染组的骨髓间充质干细胞活细胞数明显高于未转染组(均为P<0.01);转染组的骨髓间充质干细胞表现出较高的增殖能力,缺血缺氧干预后24 h及48 h细胞增殖升高较未转染组明显,差异有显著性意义(P<0.01,P<0.05);缺血缺氧后转染组骨髓间充质干细胞的凋亡比例明显下降(P<0.05)。两组细胞均可促进血管腔样结构形成,但转染组血管管腔样结构较未转染组显著增多(P<0.01)。研究结果提示miR-378可促进缺血缺氧后骨髓间充质干细胞的生长增殖并抑制其在缺血缺氧条件下的细胞凋亡,同时可提高其促血管生成的能力。

BACKGROUND：To investigate the influence of miR-378 on bone marrow mesenchymal stem cels proliferation and apoptosis under hypoxic-ischemic condition wil provide a new method for further improving the survival of bone marrow mesenchymal stem cels in the infarcted myocardium. OBJECTIVE：To observe the viability of bone marrow mesenchymal stem cels and their ability of angiogenesis under hypoxic-ischemic condition after miR-378 transfection. METHODS：The bone marrow mesenchymal stem cels of Sprague-Dawley rat were culturedin vitro and divided into the non-transfection group and the transfection group. Mesenchymal stem cels were transfected with miR-378 mimic in the transfection group while the non-transfection group was taken as the control group. After transfection, the two groups were incubated under hypoxic-ischemic condition （serum-free medium,1% O2, 5% CO2, 94% N2） at 37℃ for 24 hours. Viability and proliferation of bone marrow mesenchymal stem cels were evaluated by the trypan-blue exclusion assay and MTS assay respectively. The cellapoptosis was detected by TUNEL assay. The culture supernatant of the two groups was colected separately and used as the conditioned medium after their exposure to hypoxic-ischemic environment. Endothelial cels were then co-cultured with the conditioned medium for the vasculature formation assay. RESULTS AND CONCLUSION： After the experience of hypoxia-ischemia for 24 hours and 72 hours, there was a larger number of the living cels in the transfection group in contrast to the non-transfection group （bothP 〈 0.01）. Compared with the non-transfection group, miR-378-MSCs group presented a stronger ability of proliferation, and their proliferation rates were obviously higher at 24 and 48 hours （P 〈 0.01 andP 〈 0.05, respectively）. The percentage of apoptotic cels in the transfection group was significantly declined under the hypoxic-ischemic condition （P 〈 0.05）. The vasculature formation assay indicated that the lumen-like structures were found in both of the two groups, however, the number of lumen-like structures was remarkably increased in the transfection group （P 〈 0.01）. miR-378 could promote the proliferation of bone marrow mesenchymal stem cels and suppress their apoptosis under hypoxic-ischemic condition. It could also enhance their ability of vasculogenesis.