Dai Guo-Hua, Ma Pei-Ze, Song Xian-Bo, Liu Ning, Zhang Tong, Wu Bo
Affiliated Hospital of Shandong University of Traditional Chinese Medicine, Jinan, China.
Shandong University of Traditional Chinese Medicine, Jinan, China.
PLoS One. 2014 Oct 14;9(10):e108468. doi: 10.1371/journal.pone.0108468. eCollection 2014.
MicroRNAs (miRNAs) are a recently discovered class of posttranscriptional regulators of gene expression with critical functions in the angiogenesis and cardiovascular diseases; however, the details of miRNAs regulating mechanism of angiogenesis of ischemic cardiac microvascular endothelial cells (CMECs) are not yet reported.
This study analyzes the changes of the dynamic expression of miRNAs during the process of angiogenesis of ischemic CMECs by applying miRNA chip and real-time PCR for the first time. Compared with normal CMECs, ischemic CMECs have a specific miRNAs expression profile, in which mir-223-3p has the most significant up-regulation, especially during the process of migration and proliferation, while the up-regulation is the most significant during migration, reaching 11.02 times. Rps6kb1 is identified as a potential direct and functional target of mir-223-3p by applying bioinformatic prediction, real-time PCR and Western blot. Pathway analysis report indicates Rps6kb1 regulates the angiogenesis by participating into hif-1a signal pathway. Further analysis reveals that both the gene and protein expression of the downstream molecules VEGF, MAPK, PI3K and Akt of Rps6kb1/hif-1a signal pathway decrease significantly during the process of migration and proliferation in the ischemic CMECs. Therefore, it is confirmed that mir-223-3p inhibits the angiogenesis of CMECs, at least partly, via intervening RPS6KB1/hif-1a signal pathway and affecting the process of migration and proliferation.
This study elucidates the miRNA regulating law in the angiogenesis of CMECs; mir-223-3p inhibits the process of migration and proliferation of ischemic CMECs probably via affecting RPS6KB1/hif-1a signal pathway, which in turn suppresses the angiogenesis. It is highly possible that mir-223-3p becomes a novel intervention core target in the treatment of angiogenesis of ischemic heart diseases.
微小RNA（miRNA）是最近发现的一类基因表达的转录后调节因子，在血管生成和心血管疾病中具有关键作用；然而，miRNA调节缺血性心脏微血管内皮细胞（CMECs）血管生成机制的细节尚未见报道。
本研究首次应用miRNA芯片和实时定量PCR分析缺血性CMECs血管生成过程中miRNA动态表达的变化。与正常CMECs相比，缺血性CMECs具有特定的miRNA表达谱，其中mir-223-3p上调最为显著，尤其是在迁移和增殖过程中，而在迁移过程中上调最为明显，达到11.02倍。通过生物信息学预测、实时定量PCR和蛋白质印迹法，确定Rps6kb1是mir-223-3p潜在的直接功能靶点。通路分析报告表明，Rps6kb1通过参与hif-1a信号通路调节血管生成。进一步分析发现，在缺血性CMECs迁移和增殖过程中，Rps6kb1/hif-1a信号通路下游分子VEGF、MAPK、PI3K和Akt的基因和蛋白表达均显著降低。因此，证实mir-223-3p至少部分通过干预RPS6KB1/hif-1a信号通路并影响迁移和增殖过程来抑制CMECs的血管生成。
本研究阐明了CMECs血管生成中miRNA的调节规律；mir-223-3p可能通过影响RPS6KB1/hif-1a信号通路抑制缺血性CMECs的迁移和增殖过程，进而抑制血管生成。mir-223-3p极有可能成为治疗缺血性心脏病血管生成的新型干预核心靶点。
