LncRNA MALAT1通过GADD45A抑制BM-MSCs放射后成骨分化机制研究

The structure and function impairment of bone morrow microenvironment is one of the main pathological basis of the hematopoietic failure of bone marrow-type acute radiation sickness. Previous researches displayed that GADD45A could promote the osteogenic differentiation of BM-MSCs by regulating the expression of osteogenic genes. Our studies revealed that irradiation induced the high expression of GADD45A. However, the osteogenic differentiation of BM-MSCs was reduced, which contradicts with GADD45A up-regulated after irradiation, and the molecular mechanism remains unknown. What’s more, the results of bioinformatic analysis and RNA Immunoprecipitation indicated that lncRNA MALAT1 could bind to GADD45A. After silenced long non-coding RNA MALAT1 by RNA interference, the osteogenic differentiation capability of BM-MSCs was partial recovery. Thus, we speculate that irradiation induced the high expression of lncRNA MALAT1 and GADD45A and lncRNA MALAT1 could directly bind to GADD45A, and then suppresses the osteogenesis differentiation of BM-MSCs via inhibiting the transcriptional expression of osteogenic genes. To uncover the molecular mechanism that how lncRNA MALAT1 regulates the osteoblastic differentiation via GADD45A, we propose to construct the truncated mutants, point mutation and RNA pull down, ChIP and BSP to determiner the key site of intercation of GADD45A with lncRNA MALAT1 and to investigate the lncRNA MALAT1 inhibit the transcriptional regulation of GADD45A on osteogenetic genes. We expect to reveal the molecular mechanism of inhibition osteogenetic differentiation of BM-MSCs induced by lncRNA MALAT1 after irradiation and to provide a new idea about recovery of hematopoietic function of bone marrow-type acute radiation sickness.

骨髓微环境结构与功能损伤是骨髓型急性放射病的主要病理机制之一。已知GADD45A可调控成骨分化相关基因转录表达促进BM-MSCs成骨分化。预研发现放射诱导BM-MSCs高表达GADD45A，与其成骨分化能力下降现象相左，机制未明；同时lncRNA MALAT1表达亦升高，生物信息学和RIP分析结果提示二者可直接结合；干扰lncRNA MALAT1后，可部分恢复BM-MSCs成骨分化能力。据此推测，放射后，lncRNA MALAT1、GADD45A表达升高并相互结合，抑制后者转录调控功能，降低BM-MSCs成骨分化能力。本项目拟通过截短突变、点突变、RNA pull down、ChIP、BSP等研究lncRNA MALAT1与GADD45A的互作关键位点及其抑制GADD45A对成骨分化相关基因转录调控作用，揭示BM-MSCs放射后成骨分化降低的分子机制，为放射损伤后造血功能恢复提供新思路。

骨髓微环境结构与功能损伤是骨髓型急性放射病的主要病理机制之一。研究发现放射诱导BM-MSCs高表达lncRNA MALAT1，与其成骨分化能力下降现象相左，机制未明；本项目从以下方面进行研究：干扰lncRNA MALAT1后，可部分恢复BM-MSCs成骨分化能力。分子和细胞实验分析干扰lncRNA MALAT1后，BM-MSCs中γH2AX的升高且DNA损伤相关的基因升高。同时，通过生物信息学分析与实验验证，证明了转录调控因子KLF9在BM-MSCs成脂分化过程中具有重要的作用。本项目拟通过一系列的实验研究lncRNA MALAT1对BM-MSCs放射后成骨分化相关基因调控作用，揭示BM-MSCs放射后成骨分化降低的分子机制，为放射损伤后造血功能恢复提供研究基础。