ALPL在BMMSCs调控血管内皮细胞管腔形成中的作用研究

An experimental study of the role of ALPL in regulating tube formation of endothelial cells promoted by bone marrow mesecnchymal stem cells

目的:探究ALPL基因在骨髓间充质干细胞对内皮细胞(ECs)管腔形成及骨内H型血管的影响。方法:以正常C57BL/6小鼠骨髓间充质干细胞(mBMMSCs)及人骨髓间充质干细胞(hBMMSCs)为对照组,取ALPL^(+/-)基因敲除小鼠mBMMSCs、HPP患者hBMMSCs及通过慢病毒浸染法敲减ALPL基因的hBMMSCs为实验组,通过Western-Blot及ALP染色法检测细胞中ALPL蛋白的表达水平。采用管腔形成实验和免疫荧光染色分别检测各组成血管能力及H型血管的情况。结果:Western-Blot及ALP染色结果显示,经过慢病毒转染成功调控胞内ALPL基因表达,敲除ALPL基因导致小鼠干骺端附近H型血管数量及管腔形成能力明显降低(P<0.05)。结论:ALPL基因参与骨髓间充质干细胞调控骨内血管形成。

Objective:To explore the effects of ALPL in the regulation of mouse bone marrow mesenchymal cells(mBMMSCs)on the tube formation of endothelial cells(ECs).Methods:mBMMSCs from C57BL/6 mice and hBMMSCs from healthy people were cultured as the controls,BMMSCs from ALPL knockdown(ALPL^(+/-))mice and HPP patient as well as hBMMSCs with ALPL down-expression was designed as the experimental groups.The ALPL expression in BMMSCs were examined by Western blot and ALP staining.Immunofluorescence staining was performed to detect H type vessel in femur of ALPL^(+/-)mice,and BMMSCs co-cultured with ECs were performed to detect the effect of ALPL expression on tube formation.Results:Western blot and ALP staining verified a lower ALPL expression in BMMSCs from ALPL^(+/-)mice,HPP patient as well as hBMMSCs transfected with ALPL shRNA lentivirus compaired with that of control groups.In contrast to WT mice,ALPL^(+/-)mice displayed less H type vessels in distal femoral metaphysis.ALPL deficiency impaired the ability of BMMSCs to induce tube formation of ECs(P<0.05).Conclusion:ALPL play a significant role in BMMSCs regulating angiogenesis in bone.