骨髓间充质干细胞源性exosomes调控MAPK通路促进变性视网膜小胶质细胞M2型转化的研究

Stem cell retinal therapy has turned into a potentially useful strategy for treating retinal degeneration. However, a low rate of transplanted cell survival, presented by the diseased recipient retinal environment, is a major challenge in stem cell therapy. A prevalence of M1 (pro-inflammatory) over M2 (anti-inflammatory) microglia/macrophages has been reported in neurodegenerative pathologies, such as Alzheimer's disease and Parkinson's disease, and microglia polarization has been suggested to be a target for neuroprotective therapies. Microglia-mediated immune responses play a significant role in the survival of transplanted cells. For the first time, we used human retinal progenitor cells (hRPCs) and human bone marrow mesenchymal stem cells (hMSCs) for combined transplantation to treat a rodent model of human retinal disease-the Royal College of Surgeons (RCS) rat. We demonstrated that combined transplantation could better promote the survival of transplanted cells and photoreceptors in RCS rats, and hMSCs exhibited therapeutic effects on modulation of microglia activation. However, the mechanisms of microglia-mediated neurotoxicity or neuroprotection and the interaction between MSCs and microglia involved in retinal degeneration remain undefined. To explore whether MSCs-derived exosomes/miR-451 modulate microglia activation through the role of microglial MAPK pathway, MSCs-derived exosomes/miR-451were co-cultured with retinal microglia in vitro, and injected into the subretinal space of RCS rats in vivo, the expression of MAPK signaling and the function of microglia were measured. In this study, we investigated the mechanisms of microglia activation and MSCs-derived exosomes/miR-451 modulating microglial immune responses in retinal degeneration. Our primary aim was to improve retinal transplantation by manipulating the retinal recipient microenvironment, to determine MSCs exosomes’ providing a new therapeutic paradigm for cell-free MSCs-based therapies with decreased risk based on stem cell transplantation.

如何延长移植细胞的存活、减轻免疫排斥反应是干细胞移植治疗视网膜变性疾病的关键问题。视网膜中的小胶质细胞可发挥抗炎(M2型)和促炎(M1型)的双重作用。MAPK通路是神经元变性或炎症性疾病小胶质细胞M1型活化的促炎性通路。我们前期发现，视网膜前体细胞(RPCs)联合骨髓间充质干细胞(BMSCs)移植RCS变性大鼠视网膜，能有效抑制小胶质细胞的M1型作用，促进移植的RPCs更好存活，其作用机制不清楚。我们推测：“植入的BMSCs通过exosomes/miR-451调控MAPK通路，促进变性视网膜小胶质细胞由神经毒性的M1型向神经保护的M2型转化”。为此，本研究拟：体外和在体观察MAPK信号通路在变性视网膜小胶质细胞活化中的作用及BMSCs源性exosomes/miR-451对其调控的机制。本研究为干细胞移植改善视网膜微环境减轻免疫排斥反应、干细胞联合移植策略提供理论依据。

如何减少光感受器凋亡是治疗视网膜色素变性的核心问题。变性过程中视网膜中的小胶质细胞过度激活，免疫功能恶化，是导致光感受器继发性凋亡的关键因素。调控小胶质细胞，使其恢复稳态维持型免疫功能是延缓光感受器凋亡的可行方案。前期研究发现BMSCs源性外泌体移植，可在一定程度上改善视网膜变性微环境，抑制小胶质细胞的不良免疫反应。有报道称，用IFN-γ处理的BMSCs产生的外泌体包含更多的抗炎和神经保护内容物，对免疫性疾病产生抗炎等免疫调控作用。但在RP中尚未见报道。我们推测：“IFN-γ处理可以使BMSCs源性外泌体内容物发生变化，使其在调控小胶质细胞免疫功能，改善视网膜变性微环境，延缓RP进程上有更好的效果”。为此，本研究拟通过研究体内体外条件下，IFN-γ处理BMSCs产生的外泌体是否对小胶质细胞异常激活的免疫功能状态有更好的作用。本研究将为小胶质细胞异常激活的免疫性疾病尤其是视网膜色素变性的治疗提供重要理论依据。