活化小胶质细胞对骨髓间充质干细胞迁移、存活的作用及机制研究

Bone marrow mesenchymal stem cells（MSCs）have been shown to travel toward the injuried regions under the pathological conditions and repair the damaged neural tissues recently. However, only a few of transplanted MSCs can migrate to targeted regions and survive for a shorter time, which seriously influences the MSC secretion and neural differentiation and the effects of cell transplantation. So, it is important to elucidate the factors and the underlying mechanisms that affecting the migration and survival of MSCs in the microenvironment of diseased central nervous system (CNS), which would be helpful to take active measures to improve the migration and survival of the transplanted MSCs efficiently. Previous studies have proved that microglia, one of the major residential glial cells in the CNS, were significantly activated under pathological conditions and secreted a series of bioactive factors, which constituted the local microenvironment of CNS and played important roles in the nerve injuries and tissue repair. However, it remains unclear how the activated microglia regulate the migration and survival of transplanted MSCs at present. In the present study, we established both cell and animal models of microglia-mediated MSCs migration and survival. The MSCs were exposed to the microglia-conditioned medium or were pretreated with the exogenous factors (TNF-α,IL-6), the specific antibodies against the factors, the molecule inhibitors of MAPKs signaling pathway, or RNAi, respectively, and the effects and underlying mechanisms involved in the MSC migration and survival induced by microglia were investigated. The results would contribute to clarify the mechanisms of governing the migration and survival of MSCs in the damaged CNS, which would provide an experimental basis for the clinical application of MSCs to treat various CNS diseases, as well as provide new targets for developing new drugs.

研究发现，病理条件下骨髓间充质干细胞（MSCs）具有向损伤病灶定向迁移的趋势，但数量有限、存活时间短，直接影响其分泌、分化等功能，使移植效果大打折扣。阐明影响干细胞迁移、存活的微环境调控机制，有助于采取积极手段促进干细胞的迁移、存活，提高移植治疗效果。中枢神经系统损伤时，小胶质细胞作为构成中枢神经系统损伤局部微环境的重要细胞成分，在神经损伤及损伤修复中发挥重要作用。目前，小胶质细胞对移植MSCs定向迁移、存活的作用及其机制尚不清楚。本课题在建立小胶质细胞诱导的MSCs迁移、存活细胞模型和动物模型基础上，通过添加外源性活性因子或其中和抗体、信号分子抑制剂以及RNAi干预等，利用免疫荧光组织化学、免疫印迹等多种形态学、分子生物学和行为学方法，深入研究活化小胶质细胞对移植MSCs迁移、存活的影响及作用机制，为临床应用MSCs治疗中枢神经系统疾病提供实验依据及开发新的药物提供新的作用靶点。

虽然大量研究表明，移植骨髓间充质干细胞（MSC）治疗中枢神经系统损伤能促进受损神经功能的恢复，但由于移植后大多数细胞存活时间较短，且最终只有少部分细胞能够到达目的区域，其数量远远达不到实际需要的细胞量，从而影响移植干细胞的分泌及分化等功能，使移植效果大打折扣。因此，本课题主要研究病理微环境对移植的骨髓间充质干细胞（MSC）存活、迁移的影响，作用机制以及可能采取的预防措施。结果发现，1、炎症微环境中，活化小胶质细胞的条件培养基可促进MSCs迁移，其中小胶质细胞分泌的白细胞介素-6 (IL-6）等因子可通过调节黏着斑激酶（FAK）、MAPK、Stat3等信号通路等的表达来发挥作用，而褪黑素可明显促进这一作用。2、氧化应激和缺血缺氧等刺激，可引起MSCs细胞活性下降，凋亡增多。褪黑素预处理可促进细胞存活，减少凋亡，其作用机理可能为：褪黑素与其受体结合后，通过调控凋亡相关蛋白分子BAX、BCL-2，Caspase-3的表达，以及MAPK信号蛋白分子的磷酸化来实现。该课题的完成将有助于阐明中枢神经系统损伤微环境对移植MSCs存活、迁移的调控作用及其机理，为临床应用MSCs治疗相应疾病提供实验依据，并为探索新的治疗措施提供理论基础及作用靶点。