差异过筛法体外分离培养原代大鼠脑微血管内皮细胞及其鉴定  

Utilizing differential sieving method for culturing primary rat brain mi⁃crovascular endothelial cells

作　　者：刘海琴 张帆 李柏霖 唐元瑜[3] LIU Haiqin;ZHANG Fan;Li BoLin;TANG Yuanyu(Department of Respiratory and Critical Care Medicine,Shanghai Changzheng Hospital,Second Affiliated Hospital of Na-val Military Medical University,Shanghai 200003,China;College of Acupuncture and Moxibustion,Fujian University of Traditional Chinese Medicine,Fuzhou 350122,China;College of Traditional Chinese Medicine,Fujian University of Tra-ditional Chinese Medicine,Fuzhou 350122,China)

机构地区：[1]海军军医大学第二附属医院上海长征医院呼吸与危重症医学科,上海200003 [2]福建中医药大学针灸学院,福建福州350122 [3]福建中医药大学中医学院,福建福州350122

出　　处：《中国病理生理杂志》2023年第9期1724-1728,共5页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81072714);福建省自然科学基金资助项目(No.2017J01545)。

摘　　要：目的:采用差异过筛法培养纯度高、活性好的原代大鼠脑微血管内皮细胞(BMECs)。方法:取4周龄SD大鼠脑皮质,经剪碎、细胞筛网过滤、收集网下滤过物、Ⅱ型胶原酶消化后,置于CO_(2)培养箱中进行原代培养。通过细胞形态学观察、血管性血友病因子(vWF)免疫细胞化学染色鉴定所培养的目的细胞。结果:培养24 h后血管段周围爬出短梭形的细胞;48 h后岛屿状的细胞集落形成;72 h后细胞铺满瓶底,呈典型的单层、铺路石样、镶嵌式贴壁生长;免疫细胞化学染色显示,所培养细胞的细胞质呈现棕红色,vWF表达为阳性。结论:差异过筛法能够成功分离培养出原代大鼠BMECs。AIM:To establish a method for isolating and culturing primary rat brain microvascular endothelial cells(BMECs)with high purity using the differential sieving technique.METHODS:Brain tissue from 4-week-old SD rats was minced,passed through a cell strainer,and digested with type Ⅱ collagenase to obtain brain microvascular seg-ments,which were then placed in a CO_(2) incubator for primary culture.The cells were identified by morphological observa-tion and immunocytochemical staining for von Willebrand factor(vWF).RESULTS:After 24 h of culture,short spindle-shaped endothelial cells migrated out from the microvascular segments.Forty-eight hours later,the cells formed island-like colonies.After 72 h,the cells covered the bottom of the container and exhibited a typical cobblestone-like pattern.Immunocytochemical staining for vWF showed brownish-red cytoplasm,indicating positive expression.CONCLUSION:The differential sieving method can successfully isolate and culture primary rat BMECs with exceptional purity.

关 键 词：脑微血管内皮细胞 原代培养 差异过筛法 血管性血友病因子 大鼠 

分 类 号：R329.21[医药卫生—人体解剖和组织胚胎学] R363.2[医药卫生—基础医学]