lncRNA HK2P1, PGK1P1/P2作为竞争性RNA抑制蜕膜细胞瓦氏效应诱发先兆子痫的机制

Glycolysis, which converts glucose into pyruvate, it is an oxygen independent metabolic pathway. Then, when oxygen is used in the disposal of pyruvate, the process is usually referred to as aerobic which pyruvate is oxidized to carbon dioxide and water, whereas if the disposal uses no oxygen the process is said to be anaerobic when the oxygen is unavailable, and the pyruvate is deoxidized into lactic acid. While in rapidly growing tumor cells，the large amount of pyruvate is converted into lactic acid even under normal oxygen environment, that is the Warburg effect. There exists Warburg effect in the decidual cells of pregnant women. Preeclampsia (PE) is a disorder of pregnancy characterized by high blood pressure and a large amount of protein in the urine which occurs after 20 weeks of pregnancy. PE increases the risk of poor outcomes for both the mother and the baby. Our previous research has demonstrated that the deficiency of decidualization would cause PE, yet the mechanism is unknown. So we collected the decidual tissues of the parturient women and conducted long noncoding RNA (lncRNA) sequencing. The co- expression network analysis found that the two key enzymes of glycolysis, which are HK2 (hexokinase 2) and its pseudogene HK2P1，PGK1( phosphoglycerate kinase 1) and its two pseudogenes appear in pairs，and their transcription levels were all decreased in the tissues with PE complications, at the same time, the main glucose transporting protein, SLC2A1（also called GLUT) and pyruvate dehydrogenase kinase which inhibits aerobic oxidation were reduced significantly, too. Furthermore, we found 648 miRNAs by bioinformatics analysis that interact with them. Therefore, this application wants to carry out a comprehensive study of these RNAs and lncRNAs by a serials of methods and technics, so to explicit the roles of Warburg effect and mechanisms underlying the pathogenesis of PE.

葡萄糖经过糖酵解形成丙酮酸，之后缺氧时丙酮酸生成乳酸；有氧时生成CO2和水。而肿瘤细胞在有氧环境下，丙酮酸也生成乳酸，即瓦氏效应。妊娠子宫蜕膜中也存在瓦氏效应。先兆子痫（PE）是指孕妇出现的高血压、蛋白尿等症状，它严重危害母婴健康。我们前期研究发现蜕膜化缺陷会导致PE发生，但机制仍不清楚，为此我们收取了蜕膜进行了长非编码RNA（lncRNA）测序，共表达网络分析发现糖酵解通路的HK (己糖激酶) 2及其假基因HK2P1；PGK (磷酸甘油酸激酶) 1 及其2个假基因PGK1P1、PGK1P2在核心基因中成对出现，它们在PE里都显著降低，同时该通路的葡萄糖转运蛋白SLC2A1（GLUT)及抑制有氧氧化的丙酮酸脱氢酶激酶也显著降低。此外生物信息学分析发现了648个和它们相互作用的miRNA。因此本申请拟通过一系列的手段对这些基因/假基因进行研究，以明确瓦氏效应障碍在PE发病中的作用及机制。

葡萄糖是生物的主要供能物质和活细胞的能量来源。葡萄糖经过分解形成丙酮酸后，在无氧时生成乳酸；有氧时生成CO2和水。而肿瘤细胞在有氧环境下，丙酮酸也被代谢成乳酸，即有氧糖酵解也叫作瓦氏效应。我们先前的研究发现妊娠子宫蜕膜中也存在瓦氏效应；并且经RNAseq分析后发现瓦氏效应通路关键的酶己糖激酶(HK) 2及其假基因HK2P1；磷酸甘油酸激酶(PGK) 1 及其2个假基因PGK1P1、PGK1P2在PE患者蜕膜组织的核心基因中成对出现，并且表达量显著降低，同时该通路的葡萄糖转运蛋白SLC2A1（GLUT1)及抑制有氧氧化的丙酮酸脱氢酶激酶也显著降低。因此本研究通过一系列的实验方法对这些基因/假基因进行了研究，以明确糖酵解障碍在PE发病中的作用及机制。我们发现HK2P1和HK2对于hESCs的蜕膜化至关重要。假基因HK2P1通过竞争性的和 miR-6887-3p结合调控HK2的表达。而miR-6887-3p在PE患者蜕膜组织中显著升高，hESCs中miR-6887-3p 抑制蜕膜化和糖酵解。并且HK2P1和 HK2 都是miR-6887-3p 的直接靶标。PE患者蜕膜中HK2P1的下调减少了对miR-6887-3p的结合，使得更多的miR-6887-3p能够抑制HK2的表达，而HK2 的下调抑制了hESCs的糖酵解导致蜕膜化缺陷，并最终促成PE的发生。而PGK1和PGK1P2是一对通过miR-330-5p连接的竞争性内源RNA，二者都是miR-330-5p的靶基因，它们通过竞争性地结合miR-330-5p而达到调节彼此表达水平的目的，二者通过调节糖酵解通路及血管生成在人类蜕膜化中发挥至关重要的作用。PGK1和PGK1P2在蜕膜组织中的缺乏将引起子宫蜕膜化不足，这和PE的发生有一定的关系。总之，糖酵解受阻导致蜕膜化缺陷最终促成PE的发生。另外我们还研究了核心基因中的另外2个：BNIP3和DDIT4,发现BNIP3通过mTOR/ p70s6k/ BCL-2信号通路影响蜕膜细胞的凋亡从而参与蜕膜化进程；DDIT4对于正常蜕膜化及蜕膜细胞的凋亡来说也至关重要，它也通过影响细胞的凋亡影响蜕膜化从而参与PE的发生。即本研究揭示了一个新的调控网络，为PE的防治提供了新的靶点，也为今后进一步研究PE的发病机制提供了重要的依据和研究方向。