摘要
目的应用原代分离培养BALB/c小鼠脑微血管内皮细胞(brain microvascular endothelial cells,BMVEC)建立体外血脑屏障模型,探讨跨血脑屏障(blood brainb arrier,BBB)电阻与屏障渗透功能的动态关系以及最佳构建条件。方法用酶消化、机械分离结合密度离心的方法得到原代BALB/c小鼠脑血管内皮细胞,通过培养在具有特殊质材和孔径的Transwell小室上建立BBB体外实验模型,采用倒置显微镜、电镜观察细胞形态结构和生长规律,紧密连接ZO-1蛋白免疫组化检测,比较血脑屏障形成前后膜两侧电阻动态变化与3H葡萄糖通透性的关系等方法,探讨血脑屏障模型的建立及生长特性。结果BMVEC培养至汇合后具有典型的"铺路石"样外观;扫描电镜显示细胞形成致密单层,透射电镜、ZO-1蛋白免疫组化证实细胞间形成光滑、连续、高密度的紧密连接;3H葡萄糖的通透量与实时电阻呈负相关,内皮细胞电阻随着通透性的增加而降低,通透率最低时跨细胞电阻为(346±10)Ω/cm2。结论建立的BBB体外模型在形态学、电阻和通透性方面具备了BBB的基本特性。
Objective To study the dynamic changes of transendothelial electrical resistance （TEER） on our in vitro model of brain blood barrier （BBB） made from primary culture of BALB/c mouse brain microvascular endothelial cells （BMVEC） , and to explore the relation between TEER and BBB permeability, and search for the best culture condition. Methods BMVEC were isolated from BALB/c mouse and cultured on a transwell insert with special micro-pore. The cells were identified with immunohistochemical methods and electron microscope. TEER over BMVEC was measured after BBB model establishment for determining the 3H-Glucose permeability of BBB in vitro. Results BMVEC cultured in the transwell insert exhibited typical ＂flagstone＂ appearance and in a tight monolayer structure under electron microscope. Immunohistochemical detection of ZO- 1 protein, a marker antigen of tight junction, showed smooth, continuous and tight junctions between confluent BMVEC. TEER over BMVEC monolayer increased to （346±10）Ω/cm2 when the permeability for 3H-C, lucose was decreased to the minimum. Conclusion BBB model in vitro made from primary culture of BMVEC in transwell has the basic characteristics of BBB in morphology, electrical resistance and permeability.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2008年第22期2093-2095,共3页
Journal of Third Military Medical University
基金
国家自然科学基金(30670545)~~
关键词
血脑屏障
体外模型
通透率
电阻
blood brain barrier
in vitro model
permeability
electric resistance
