Sirtuin在低氧诱导人脐带间充质干细胞增殖中的作用

发布时间：2018-06-20 17:42

本文选题：人脐带间充质干细胞 + 低氧　；参考：《山东大学学报(医学版)》2017年07期

【摘要】：目的探讨低氧对人脐带间充质干细胞(h UC-MSCs)中Sirtuin(SIRT)蛋白表达影响及与细胞增殖的关系。方法组织块贴壁法分离和培养h UC-MSCs;流式细胞仪鉴定细胞表面标志物;CCK-8法测定低氧与常氧下不同时间点h UC-MSCs的增殖能力;流式细胞仪检测低氧与常氧下细胞周期的变化;免疫荧光与Western blotting测定低氧与常氧下细胞SIRT蛋白定位与表达。结果成功分离h UC-MSCs;细胞CD90、CD105、CD29、CD44高表达,CD14、CD19、CD34、CD45、HLA-DR无表达;CCK-8测定低氧培养24、48、72 h细胞的增殖能力均高于常氧培养(P均0.05);在低氧下培养24、48 h后S期细胞比率高于常氧培养(P均0.05);免疫荧光检测SIRT1、6位于细胞核内,SIRT2定位于胞浆,SIRT3、4、5定位于线粒体。Western blotting测定SIRT1在低氧培养24、48 h蛋白表达量均高于常氧条件(P均0.05),常氧培养72 h的SIRT1表达量高于24 h及48 h(P均0.05);SIRT2表达量在低氧培养24 h高于常氧培养(P0.05),48 h后低于常氧培养,72 h后高于常氧培养(P0.05);低氧培养24、48、72 h后SIRT5的表达量均高于常氧处理(P均0.05),常氧与低氧培养细胞48 h和72 h后均高于24 h时SIRT5的表达(P均0.05)。结论低氧处理24、48、72 h均可以促进h UC-MSCs的增殖。低氧诱导SIRT1、2与5表达改变提示SIRT1、2、5可能参与了低氧对h UC-MSCs增殖的调控。
[Abstract]:Objective to investigate the effect of hypoxia on the expression of Sirtuintin-SIRT protein in human umbilical cord mesenchymal stem cells (hUC-MSCs) and its relationship with cell proliferation. Methods hUC-MSCs were isolated and cultured by tissue mass adherence method, cell surface marker (CCK-8) was identified by flow cytometry, the proliferative ability of hUC-MSCs at different time points under hypoxia and normoxia was determined by flow cytometry, the changes of cell cycle under hypoxia and normoxia were detected by flow cytometry. The localization and expression of SIRT protein in hypoxic and normoxic cells were detected by immunofluorescence and Western blotting. Results Human UC-MSCs were successfully isolated, the high expression of CD90, CD105, CD29, CD44, CD14, CD19, CD34, CD45, CD45, HLA-DR, and CCK-8 were all higher than those in normoxic culture (P = 0.05, P = 0.05), and the percentage of S phase cells was higher than that in normoxic culture (P < 0.05). Localization of SIRT6 in cytoplasm SIRT2 and mitochondria. Western blotting Detection of SIRT1 protein expression at 24 h in hypoxia culture was higher than that in normoxic condition (P 0.05). The expression of SIRT1 in normal oxygen culture at 72 h was higher than that in 24 h and 48 h P The expression of SIRT2 in 24 h hypoxia culture was higher than that in normal oxygen culture after 48 h, and was higher than that in normal oxygen culture (72 h after 72 h), the expression of SIRT5 in hypoxia culture was higher than that in normal oxygen treatment (P 0.05), the expression of SIRT5 in hypoxia culture was higher than that in normal oxygen culture (P 0.05), and the expression of SIRT5 in hypoxia culture was higher than that in normal oxygen culture (48 h) and hypoxia culture (48 h). The expression of SIRT5 at 72 h was higher than that at 24 h. Conclusion hypoxia treatment can promote the proliferation of hUC-MSCs for 72 h. Hypoxia induced changes in the expression of SIRT1O-2 and 5 suggest that SIRT1m2O5 may be involved in the regulation of hypoxia on the proliferation of hUC-MSCs.
【作者单位】：南方医科大学第三临床医学院附属奉贤区中心医院神经外科;上海交通大学生命科学技术学院;南方医科大学第三临床医学院附属奉贤区中心医院中心实验室;
【基金】：国家自然科学基金(10972141,11172176,31570949)
【分类号】：R329.2

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