lncRNA-RP11去甲基化调控Rab26介导肺血管重构的囊泡运输机制

Pulmonary vascular remodeling is the basic pathogenesis of chronic cor-pulmonale from COPD (chronic obstructive pulmonary disease); and its initiating key point is based on the disorder of vesicle trafficking and phenotypic transformation of PASMC (pulmonary arterial smooth muscle cells). We found that lncRNA-RP11, as a new type long non-coding RNA, was obviously increased in the tissue specimen from COPD pulmonary hypertension patients (cor-pulmonale) and animal models; there were remarkable positive correlation in the expression levels between lncRNA-RP11 and Rab26. Rab26 is also the target gene of miR135a, which binding loci is involved in the sequences of lncRNA-RP11. Here is our hypothesis: To manipulate hypoxia-mediated lncRNA-RP11 promoter demethylation can decrease the expression of RP11, maintain the stable levels of Rab26 via adsorption of miR135a, suppress the phenotype transformation of PASMC, and then control the progress of pulmonary remodeling. By using these techniques including gene kick-out mice, RNA-RIP (RNA binding protein immunoprecipitation) and FISH (fluorescence in situ hybridization), to investigate the regulation of hypoxia-mediated lncRNA-RP11 promoter methylation on the phenotypic modulation of PASMC; the rival effect of lncRNA RP11 and miR-135a as the competing endogenous RNA in PASMC proliferation; the modulation of co-localazation of lncRNA-RP11/miR-135a and Rab26 on vesicle transport from membrane to organelle in pulmonary vascular cells. The mechanism would be clarified about regulation of lncRNA-RP11 in Rab26-mediated the phenotype transformation and vesicle trafficking of PASMC. This study will provide the new targets and strategies for treatment of clinical diseases related to pulmonary vascular remodeling.

肺血管重构是COPD发展为肺心病的基本病理基础，而肺动脉平滑肌细胞（PASMC）囊泡运输紊乱及表型转化是其关键环节。我们在COPD肺高压患者及鼠模型肺组织标本中发现lncRNA-RP11表达显著增加，并与Rab26表达呈显著正相关；还发现Rab26是miR135a的靶基因，而RP11序列上含有miR135a的结合位点。故推测：调控低氧介导RP11启动子的去甲基化，影响其表达，经吸附miR135a再稳定Rab26介导的囊泡运输，抑制PASMC表型转化，从而控制肺血管重构。本项目应用基因敲除鼠、RNA-RIP和FISH等方法研究RP11启动子去甲基化对细胞表型转化的调节；RP11与miR135a的内源性竞争结合的效应；RP11/miR135a与Rab26共定位协调肺血管细胞胞膜转运进程。阐明RP11调节Rab26介导的PASMC表型转化和囊泡运输机制，为肺血管重构疾病的防治提供新靶点和新策略。

肺血管重构是COPD发展为肺心病的基本病理基础，而肺动脉平滑肌细胞（PASMC）囊泡运输紊乱及表型转化是其关键环节。我们在COPD肺高压患者及鼠模型肺组织标本中发现lncRNA-RP11表达显著增加，并与Rab26表达呈显著正相关；还发现Rab26是miR-135a的靶基因，而RP11序列上含有miR-135a的结合位点。故推测：调控低氧介导RP11启动子的去甲基化，影响其表达，经吸附miR-135a再稳定Rab26介导的囊泡运输，抑制PASMC表型转化，从而控制肺血管重构。研究的内容主要为：（1）低氧介导lncRNA-RP11去甲基化在PASMC表型转化中的作用及机制；（2）lncRNA-RP11对miR-135a的内源性竞争结合作用及效应；（3）lncRNA-RP11通过Rab26介导的囊泡运输进程调控PASMC表型转化；（4）lncRNA-RP11及Rab26在低氧肺动脉高压模型及基因敲除鼠中的重要作用；（5）lncRNA-RP11表达水平和COPD肺心病患者肺血管重构的相关性研究。（6）研究纳米材料负载Rab26的转染效率，并进一步研究了LPS刺激对Rab26表达和肺血管重构的影响。研究结果揭示：（1）阐释低氧介导lncRNA-RP11去甲基化；（2）在细胞水平上揭示miR-135靶向作用Rab26介导细胞增殖；（3）Rab26介导低氧诱导的PASMC表型转化；（4）Rab26在肺血管重构中起关键作用。在COPD肺心病患者组织等临床标本，阐述miR-135调节Rab26及其相关信号通路。本项目阐明RP11调节Rab26介导的PASMC表型转化和囊泡运输机制，为防止肺血管重构和肺动脉高压提供新的新靶点和新的防治策略。