脑血管内皮细胞敲除Prmt5导致脑血管损伤及星形胶质细胞活化  

Deletion of Prmt5 in Cerebral Endothelial Cells Leads to Cerebrovascular Disease and Astrocyte Activation

作　　者：宁慧敏 张一哲 韩钰莹 张翀 宋晓朋 梁爽 杨晓[1,2] 王俊 NING Hui-min;ZHANG Yi-zhe;HAN Yu-ying;ZHANG Chong;SONG Xiao-peng;LIANG Shuang;YANG Xiao;WANG Jun(School of Basic Medicine,Qingdao University,Qingdao 266071,China;State Key Laboratory of Proteomics,Beijing National Center for Protein Sciences(Beijing),Beijing Institute of Lifeomics,Beijing 102206,China)

机构地区：[1]青岛大学基础医学院,青岛266071 [2]军事科学院军事医学研究院生命组学研究所,蛋白质组学国家重点实验室,北京102206

出　　处：《中国生物工程杂志》2022年第4期1-8,共8页China Biotechnology

基　　金：国家自然科学基金重点项目(82030011)资助项目。

摘　　要：目的:研究蛋白质精氨酸甲基转移酶5(protein arginine methyltransferase 5,Prmt5)在小鼠脑血管发育、稳态维持中的功能,并考察脑血管内皮细胞特异性敲除Prmt5后对中枢神经系统的影响。方法:利用脑血管内皮细胞特异性表达SP-A-Cre转基因小鼠和Prmt5条件基因打靶小鼠交配,构建脑血管内皮细胞特异性Prmt5敲除小鼠。利用H-E染色、免疫荧光染色、激光散斑成像、Sulfo-NHS-Biotin染料灌注等方法评价脑血管内皮细胞特异性Prmt5敲除小鼠脑血管结构、脑血流量、血脑屏障渗透性等;利用实时定量PCR进一步检测补体C1q(complement C1q,C1q)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和白细胞介素-1β(interleukin 1β,IL-1β)等细胞因子的表达水平。通过免疫荧光、Western blot等检测胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)、S100钙结合蛋白β(S100 calcium-binding proteinβprotein,S100β)和补体C3(complement C3,C3)的表达,检测小鼠皮层、丘脑和小脑中星形胶质细胞活化水平。结果:脑血管内皮细胞特异性敲除Prmt5导致血管损伤,C1q、TNF-α和IL-1β等炎症因子表达水平上调,活化星形胶质细胞比例明显增加。结论:脑血管内皮细胞中Prmt5在小鼠脑血管稳态维持中发挥了重要功能。Objective:To study the role of protein arginine methyltransferase 5(Prmt5)in cerebral vascular development and homeostasis maintenance in mice,and to investigate the effect of Prmt5 specific knockout on the central nervous system.Methods:We crossed Prmt5;mice with SP-A Cre transgenic mice that express Cre recombinase in cerebrovascular endothelial to generate cerebrovascular endothelial cell-specific Prmt5 knockout mice.H-E staining and immunostaining were performed to observe the vascular structures of control and Prmt5;mutant mice.Laser speckle contrast imaging was used to detect cerebral blood flow in control and mutant mice.Sulfo-NHS-Biotin was intraperitoneally injected into control and mutant mice to examine the blood brain barrier(BBB)integrity.The expression levels of astrocyte glial fibrillary acidic protein(GFAP),S100 calcium-binding proteinβ(S100β),complement C3(C3),C1 q,tumor necrosis factor alpha(TNF-α)and Interleukin-1 beta(IL-1β)were detected by immunofluorescence and Western blot to evaluate the activation level of astrocytes in cortex,thalamus and cerebellum of knockout mice and control mice.Furthermore,activators of astrocytes,such as C1 q,TNF-α,IL-1βand other cytokines,were also detected by real-time PCR.Results:We found that cerebrovascular endothelial cell-specific Prmt5 knockout mice exhibited aberrant cerebrovascular structure,and increased the number of reactive astrocytes.The expression levels of TNF-αand IL-1βin the whole brain,as well as the C1 q,TNF-αand IL-1β,were all increased in Prmt5;mutant mice.Conclusion:Prmt5 plays an essential role in the maintenance of cerebrovascular homeostasis,suggesting that it might act as a potential therapeutic target for cerebrovascular diseases.

关 键 词：脑血管内皮细胞 Prmt5 星形胶质细胞 

分 类 号：Q819[生物学—生物工程]