Perp调控少突胶质细胞发育在新生小鼠脑白质损伤中的作用

White matter injury (WMI) of preterm infants has been a major global health problem. Evidence indicates that failure of differentiation and myelination of oligodendrocyte precursor cells (OPCs) is the main course of WMI. Previous studies have reported that the upstream or related genes of Perp are closely related to oligodendrocytes (OLs) mediated myelination after cerebral ischemic injury. Our previous study has shown that PERP protein was expressed in OLs of neonatal mice and the expression increased in corpus callosum after WMI modeling. Through constructing WMI model of neonatal Olig2-Cre×Perpfl/fl mice, we found that the myelination disorder of Perp-/-mice alleviated, and the results of Morris water maze test were improved after WMI compared with littermate control at P30. Based on these results, we put forward the hypotheses that by regulating development of OLs, Perp may be closely related to WMI. Thus, in this study, we will construct WMI model in neonatal mice, and compare the changes in survival and differentiation of OLs, and in cognitive function among Perp conditional knockout mice or not. Furthermore, the primary Perp-/- OPCs were separated and cultured to analyze the underlying effects of Perp on the apoptosis, proliferation and differentiation of OPCs. The results will be conducive to clarify the intrinsic mechanism of Perp in regulating OLs development and function, and provide new ideas for medical intervention of WMI.

早产儿脑白质损伤(WMI)是世界性的健康问题。少突胶质前体细胞(OPCs)分化、成熟及髓鞘化障碍是造成WMI的主要原因。既往研究报道Perp上游及相关基因与脑缺血损伤后少突胶质细胞（OLs）介导的髓鞘化密切相关。我们前期研究显示，PERP蛋白在新生小鼠OLs表达，且在WMI造模后增加；采用Olig2-Cre×Perpfl/fl条件敲除小鼠造模发现，与同窝野生型小鼠相比，Perp-/-小鼠WMI后髓鞘化障碍减轻，Morris水迷宫行为改善。据此提出假设：Perp可能通过调控OLs发育而与WMI密切相关。因此，本项目将通过构建新生鼠WMI模型，比较条件敲除Perp对OLs存活与分化、白质髓鞘化及认知功能的影响。并分离原代Perp-/-OPCs，分析Perp对OPCs凋亡、增殖和分化的调控作用及相关分子机制。研究结果将有助于明晰Perp调控OLs发育的内在机制，为WMI医学干预提供新思路。

早产儿脑白质损伤(WMI)是世界性的健康问题。Perp（p53 effector related to PMP-22/gas 3）作为髓鞘磷脂蛋白/生长停滞特异性基因3跨膜蛋白家族成员，是p53基因独有的泛表达效应分子，在外胚层形成器官如表皮和神经组织等均有丰富的表达。既往研究报道Perp上游及相关基因与脑缺血损伤后少突胶质细胞（OLs）介导的髓鞘化密切相关。我们前期研究显示，Perp可能通过调控OLs发育而与脑白质损伤（WMI）密切相关。本项目中，我们采用Olig2-Cre×Perpfl/fl条件敲除小鼠进行WMI造模，免疫荧光、WB、电镜检测不同基因型小鼠髓鞘化情况，Morris水迷宫、平衡木行走实验及新物体识别实验检测小鼠行为学改变，结果显示，与同窝野生型小鼠相比，Perpfl/fl组小鼠髓鞘化增加，行为学改善，结果提示Perp基因在OLs条件性敲除可改善新生小鼠WMI白质损伤。Olig2-Cre×Perpfl/fl条件敲除小鼠进行WMI造模，免疫荧光检测WMI后OLs增殖、凋亡及不同分化阶段OLs分子标志物表达；分离培养Perp-/-少突胶质细胞前体细胞，糖氧剥夺处理模拟体内缺氧缺血，WB检测OLs分化、凋亡相关蛋白，体内外实验结果提示Perp基因在OLs条件性敲除可调控低氧后OLs存活，促进OLs存活分化。本项目相关研究结果课题负责人作为第一作者已发表在Frontiers in Pharmacology（JCR一区）及Immunology Letter（JCR二区）。项目后续实验研究结果已整理成文“Loss of Perp leads to amelioration of WMI by modulating the survival and differentiation of oligodendrocyte”，拟投稿至高质量国际期刊杂志。本项目从动物、细胞及分子水平深入研究Perp调控OLs存活及分化影响WMI发病的作用及机制，为WMI的基础研究和治疗提供新的理论依据和实验基础。