视黄酸经选择性剪接调控hBMSCs成脂分化的机制研究

Human bone marrow mesenchymal stem cells (hBMSCs) may differentiate into osteoblasts and adipocytes. The imbalance of these two cell differentiation may be an important factor in the development of osteoporosis and other diseases. A large number of studies have shown that the retinoic acid receptor signal regulates the adipogenesis of hBMSCs. Recent studies have revealed that retinoic acid significantly increases gene alternative splicing during adipocyte differentiation of hBMSCs, but its detailed mechanism remains to be elucidated. This research aims to illuminate the molecular mechanism of retinoic acid in regulating adipogenic differentiation of hBMSCs through alternative splicing mechanism, based on the previous study of retinoic acid.The alternative splicing of target genes regulated by retinoic acid are analyzed at the transcriptome level, splicing factors and alternative spliced genes are screened and identificated. The interaction mechanism between trans- acting splicing factors and cis-elements on target genes during the formation of the splicesome will be clarified. The relationship between lncRNAs-miRNAs signals regulated by retinoic acid with the alternative splicing will be determined. The results will be verified in vivo using gene knockout mice. This research will demonstrate the mechanisms regulating adipogenesis by retinoic acid via alternative splicing . It will provide a theoretical basis for profound understanding and effective treatment of degenerative bone diseases such as osteoporosis.

人类骨髓间充质干细胞(hBMSCs)成脂成骨分化失衡是骨质疏松症发生发展的重要因素。大量研究表明，视黄酸受体信号可调控hBMSCs成脂与成骨分化。最新研究揭示，视黄酸显著增加hBMSCs成脂分化过程中基因选择性剪接，但其详细机制有待阐明。本研究以hBMSCs成脂分化调控为视角，在前期视黄酸信号研究基础上，沿着视黄酸经选择性剪接机制调控成脂分化的思路开展工作。首先在转录组水平分析视黄酸对基因选择性剪接的影响，筛选并鉴定受视黄酸调控的剪接因子及选择性剪接目标基因，并明确视黄酸调控剪接复合体形成过程中反式剪接因子与目标基因顺式元件间相互作用的详细机制；然后筛选鉴定在成脂分化过程中受视黄酸调控的lncRNAs-miRNAs信号，分析其与选择性剪接的关系；上述结果采用基因敲除小鼠进行体内验证，最终系统阐述视黄酸经选择性剪接调控成脂分化的机制，为认识和治疗骨质疏松症这类骨退行性疾病提供理论依据。

人骨髓间充质干细胞（hBMSCs）成骨成脂分化失衡是骨质疏松症发生发展的重要因素，探讨hBMSCs成脂分化的调控机制对骨质疏松症发病机制的阐明具有重要意义。本项目从视黄酸经选择性剪接调控成脂分化的角度开展研究，明确了hBMSCs成脂分化与关键转录因子PPARG及SREBF1的选择性剪接密切相关，而视黄酸通过剪接因子hnRNPA1作用于这一过程并抑制成脂分化，结合转录组和蛋白质组学数据分析，系统阐明了一种新的成脂分化调控机制，是对传统转录调控的重要补充。另外，在对视黄酸选择性剪接目标基因的发掘及功能研究过程中，还发现ROCK，CTGF 等众多基因与视黄酸的成脂分化调控密切相关，大大拓展了对视黄酸功能的认识。部分下游基因在结合临床检验数据分析后，被确定为与成脂分化相关疾病有关的检测靶点，为后续的临床应用开辟了新的线路。重要研究结果如下：.1. 视黄酸经剪接因子hnRNPA1调控成脂转录因子PPARG及SREBF1选择性剪接，从而抑制成脂分化；.2. 转录组联合蛋白质组筛选确定了参与视黄酸成脂分化调控的剪接因子及剪接目标基因；.3. 视黄酸通过ROCK和CTGF等参与对成脂分化抑制的调控；.4. 基于组学及临床检测数据分析确定IL-38等与成脂分化失衡类疾病密切相关。. 上述结果初步明确了视黄酸经选择性剪接调控成脂分化的机制，不但有利于骨质疏松症发病机制的阐明，对其它成脂分化失衡相关疾病的研究也有重要参考价值。