长链非编码RNA NR024122作为ceRNA在低氧性肺动脉高压发病中的作用和机制研究

Our results and other studies showed that CAT (cationic amino acid transporter) and ACE2 (angiotensin converting enzyme 2) were down-regulated under hypoxia, which could inhibit the generation of NO and Ang-(1-7)/Mas. The lower NO and Ang-(1-7)/Mas contribute to the pathogenesis of hypoxic pulmonary hypertension. The miRNA array results showed that miR-210 and miR-let-7b were up-regulated under hypoxia, which could inhibit CAT and ACE2 expression respectively. The lncRNA array results showed that NR024122 was down-regulated under hypoxia. By further analysis, we found NR024122 sequence contained miRNA response elements both for miR-210 and miR-let-7b. According to these results, we would investigate whether lncRNA NR024122 could regulate miR-210 and miR-let-7b by competing endogenous binding or not, and the role of NR024122 in the regulation of miR-210/CAT/NO and miR-let-7b/ACE2/Ang-(1-7)/Mas pathways and the related mechanism. We would also investigate the protective role of NR024122 in the pathogenesis of hypoxic pulmonary hypertension.

既往的研究显示，低氧通过下调阳离子氨基酸转运体(CAT)和血管紧张素转换酶2(ACE2)的表达，从而分别抑制NO和Ang-(1-7)/Mas的产生，促进肺动脉高压的发生。本课题组的miRNA芯片结果显示，低氧可诱导miR-210和miR-let-7b的表达，且miR-210和miR-let-7b分别可抑制CAT和ACE2的表达。进一步的lncRNA芯片结果显示，低氧可抑制NR024122序列的表达，且该序列含有能和miR-210和miR-let-7b结合的miRNAs反应原件。在此基础上，我们将探讨lncRNA NR024122对miR-210和miR-let-7b是否具有竞争性内源性RNA(ceRNA)作用，其对miR-210/CAT/NO和miR-let-7b/ACE2/Ang-(1-7)/Mas通路的调控作用和相关机制；以及NR024122在低氧性肺动脉高压发病中是否具有保护作用。

在课题执行中，我们发现lncRNA NR024122对miR-let-7b和miR-210并没有竞争性内源性结合作用，这与之前的设想不符合。基于这个试验结果，对于原先计划的进一步验证lncRNA NR024122对miR-let-7b和miR-210的结合作用，以及具体机制，就显得没有必要。因此，对原定的研究计划作出调整，停止原定计划中lncRNA NR024122对miR-let-7b和miR-210调控机制的相关研究。同时进行文献检索，我们发现miR-let-7b和miR-210，尤其是miR-let-7b可能在肺动脉高压的发生中具有重要的作用。为此我们对原计划进行了相应的调整，调整后的研究计划主要是针对miR-let-7b进行研究，包括（1）在体外和体内水平，miR-let-7b在HPH发生中所起的作用；（2）低氧下，miR-let-7b对下游相关通路的调控作用。随后的研究发现，低氧通过HIF-1α上调miR-let-7b的表达，而miR-let-7b可抑制ACE2的表达，ACE2是miR-let-7b的1个新的靶基因。进一步研究发现，miR-let-7b能刺激PASMCs的增殖和迁移；miR-let-7b敲除后，低氧处理小鼠的右心室肥厚、肺小动脉重塑等指标明显好转。本课题阐明了低氧通过HIF-1α-miR-let-7b-ACE2轴发挥对ACE2的调控作用和相关机制，并在动物水平证实了敲除miR-let-7b具有治疗HPH的潜在作用，为HPH的发病机制研究及其治疗提供新的思路。