Hypoxic Preconditioning Inhibits Hypoxia-induced Apoptosis of Cardiac Progenitor Cells via the PI3K/Akt-DNMT1-p53 Pathway.
Hypoxic Preconditioning Inhibits Hypoxia-induced Apoptosis of Cardiac Progenitor Cells via the PI3K/Akt-DNMT1-p53 Pathway.
复制标题
缺氧预处理通过 PI3K/Akt-DNMT1-p53 途径抑制缺氧诱导的心脏祖细胞凋亡
DOI：
10.1038/srep30922
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发表时间：
2016-08-04
影响因子：
4.6
通讯作者：
Ma G
中科院分区：
文献类型：
--
作者：
Xu R;Sun Y;Chen Z;Yao Y;Ma G
研究表明，低氧预处理（HP）可以增强心脏祖细胞（CPC）的存活和增殖；然而，其根本机制尚未完全了解。在此，我们报告 c-kit (+) CPC 的 HP 通过 PI3K/Akt-DNMT1 途径抑制 p53。首先，从 C57BL/6 小鼠的心脏中分离出 CPC，并通过磁激活细胞分选进一步纯化。接下来，这些细胞在常氧 (H0) 或 HP 下培养 6 小时 (H6)，然后缺氧-血清剥夺 24 小时 (24 小时)。流式细胞术分析和 MTT 测定表明，缺氧预处理的 CPC 表现出更高的存活率。 Western blot和实时定量PCR检测显示，HP明显抑制p53，而DNMT1和DNMT3β均显着上调。亚硫酸氢盐测序分析表明DNMT1和DNMT3β不会引起p53启动子高甲基化。报告基因测定和染色质免疫沉淀分析进一步证明，DNMT1 在缺氧预处理的 CPC 中与 p53 的启动子位点结合。总之，这些观察结果表明，CPC 的 HP 可能通过上调 DNMT1 和 DNMT3β 导致 p53 抑制，这不会导致 p53 启动子高甲基化，并且 DNMT1 可能通过与 p53 启动子位点结合直接抑制 p53，至少部分抑制。
Research has demonstrated that hypoxic preconditioning (HP) can enhance the survival and proliferation of cardiac progenitor cells (CPCs); however, the underlying mechanisms are not fully understood. Here, we report that HP of c-kit (+) CPCs inhibits p53 via the PI3K/Akt-DNMT1 pathway. First, CPCs were isolated from the hearts of C57BL/6 mice and further purified by magnetic-activated cell sorting. Next, these cells were cultured under either normoxia (H0) or HP for 6 hours (H6) followed by oxygen–serum deprivation for 24 hours (24h). Flow cytometric analysis and MTT assays revealed that hypoxia-preconditioned CPCs exhibited an increased survival rate. Western blot and quantitative real-time PCR assays showed that p53 was obviously inhibited, while DNMT1 and DNMT3β were both significantly up-regulated by HP. Bisulphite sequencing analysis indicated that DNMT1 and DNMT3β did not cause p53 promoter hypermethylation. A reporter gene assay and chromatin immunoprecipitation analysis further demonstrated that DNMT1 bound to the promoter locus of p53 in hypoxia-preconditioned CPCs. Together, these observations suggest that HP of CPCs could lead to p53 inhibition by up-regulating DNMT1 and DNMT3β, which does not result in p53 promoter hypermethylation, and that DNMT1 might directly repress p53, at least in part, by binding to the p53 promoter locus.