HO-1过表达可提高急性肾损伤微环境中骨髓间充质干细胞存活能力的机制探讨

Mechanism Investigation of Increasing the Viability of Bone Marrow-derived Mesenchymal Stem Cells by the HO-1 Overexpression Under the Acute Kidney Injury Mocroenvironment

目的:以急性肾损伤(AKI)大鼠的肾脏匀浆上清孵育骨髓间充质干细胞(BMSCs),体外模拟AKI微环境,观察经血红素加氧酶-1(HO-1)基因修饰的BMSCs的存活能力,并探讨其机制。方法:基因转染技术获得HO-1-BMSCs和e GFP-BMSCs(空载体对照),检测HO-1水平。制作缺血再灌注诱导的AKI大鼠肾脏匀浆上清(AKI-KHS)和正常大鼠肾脏匀浆上清(N-KHS)用于孵育BMSCs,检查各组BMSCs的存活差异,并对影响BMSCs存活的凋亡蛋白及炎症因子水平进行检测。结果:成功转染的HO-1-BMSCs可高效表达HO-1。AKI-KHS孵育的BMSCs有丝分裂停滞,伴随着PCNA表达减弱。HO-1过表达可显著降低G0/G1期BMSCs细胞比例,增加PCNA+BMSCs的比例,细胞内cleaved Caspase-3水平显著降低,Bcl-2表达升高。AKI-KHS孵育的BMSCs上清中可检测到大量的炎症因子,HO-1过表达可降低炎症因子水平。结论:HO-1过表达可恢复AKI微环境中培养BMSCs正常的有丝分裂,存活能力增强,抗氧化、抗凋亡和抗炎为其发挥作用的可能机制。

Objective:Bone marrow-derived mesenchymal stem cells(BMSCs)were cultured by the kidney homogenate supernatant of the acute kidney injury rats.Effect of HO-1 overexpression on the survival of BMSCs were observed.The possible mechanisms were also investigated.Methods:HO-1-BMSCs and e GFP-BMSCs were obtained by the gene transfection technique.The HO-1 level was detected.I/R-AKI kidney homogenate supernatant(AKI-KHS)and the control N-KHS were harvested to culture BMSCs.The survival differences of BMSCs in each group were examined.Levels of the apoptotic proteins and the inflammatory factors which affected the survival of BMSCs were also detected.Results:Expression of HO-1 was enhanced in HO-1-BMSCs.Mitotic stagnation of BMSCs incubated by AKI-KHS was observed.The PCNA expression was also weakened.HO-1 overexpression significantly decreased the proportion of BMSCs in G0/G1 phase.It also increased the proportion of PCNA+BMSCs.The decreased expression of cleaved Caspase-3 and the increased Bcl-2 expression were also observed.Many inflammatory factors were detected in the BMSCs/AKI-KHS group.HO-1 overexpression significantly decreased the levels of these inflammatory factors.Conclusion:HO-1 overexpression could restore the normal mitosis of BMSCs and enhance their survival ability cultured in the AKI microenvironment.The possible mechanisms include the anti-oxidation effect,the anti-apoptotic effect and the anti-inflammatory effect.