Colgan Olga C, Collins Nora T, Ferguson Gail, Murphy Ronan P, Birney Yvonne A, Cahill Paul A, Cummins Philip M
Faculty of Science and Health, School of Biotechnology, Dublin City University, Glasnevin, Dublin 9, Ireland.
Brain Res. 2008 Feb 8;1193:84-92. doi: 10.1016/j.brainres.2007.11.072. Epub 2007 Dec 14.
Basolateral condition of the brain microvascular endothelium is believed to influence blood-brain barrier (BBB) phenotype, although the precise transcriptional and post-translational mechanisms involved are poorly defined. In vivo, the basolateral surface of the blood-brain endothelium is bathed in serum-free interstitial fluid and encompassed by astrocytic end-feet. We hypothesized that these conditions impact on BBB function by directly modulating expression and biochemical properties of tight junctions. To investigate this, an in vitro transwell culture model was employed to selectively modify the basolateral environment of bovine brain microvascular endothelial cells (BBMvECs). In the absence of basolateral (but not apical) serum, we observed higher levels of expression, association and plasma membrane localization for the tight junction proteins, occludin and zonula occludens-1 (ZO-1), in parallel with elevated transendothelial electrical resistance (TEER) and reduced (14)[C]-sucrose permeability of BBMvEC monolayers. We further examined the effects of non-contact co-culture with basolateral astrocytes (C6 glioma) on indices of BBMvEC barrier function in both the presence and absence of serum. Astrocyte co-culture with serum led to enhanced occludin protein expression, occludin/ZO-1 association, and ZO-1 membrane localization, in parallel with increased TEER of BBMvEC monolayers. Astrocyte co-culture in the absence of serum (i.e. basolateral conditions most consistent with in vivo BBB physiology) however, gave the highest increases in BBMvEC barrier indices. Thus, we can conclude that factors influencing condition of the basolateral environment of the brain microvasculature can directly, and independently, modify BBB properties by regulating the expression and biochemical properties of the tight junction proteins, occludin and ZO-1.
尽管尚不清楚其中确切的转录和翻译后机制，但人们认为脑微血管内皮细胞的基底外侧状态会影响血脑屏障（BBB）的表型。在体内，血脑内皮细胞的基底外侧表面浸泡在无血清的间质液中，并被星形胶质细胞的终足所包围。我们推测，这些条件通过直接调节紧密连接的表达和生化特性来影响血脑屏障功能。为了对此进行研究，我们采用体外transwell培养模型来选择性地改变牛脑微血管内皮细胞（BBMvECs）的基底外侧环境。在没有基底外侧（而非顶端）血清的情况下，我们观察到紧密连接蛋白闭合蛋白和闭合小环蛋白-1（ZO-1）的表达水平、结合情况及质膜定位均有所升高，同时BBMvEC单层的跨内皮电阻（TEER）升高，[¹⁴C]-蔗糖通透性降低。我们进一步研究了在有血清和无血清情况下，与基底外侧星形胶质细胞（C6胶质瘤）进行非接触共培养对BBMvEC屏障功能指标的影响。血清存在时与星形胶质细胞共培养导致闭合蛋白表达增强、闭合蛋白/ZO-1结合增加以及ZO-1膜定位增加，同时BBMvEC单层的TEER升高。然而，在无血清情况下（即与体内血脑屏障生理学最一致的基底外侧条件）与星形胶质细胞共培养，BBMvEC屏障指标的升高最为显著。因此，我们可以得出结论，影响脑微血管基底外侧环境条件的因素可通过调节紧密连接蛋白闭合蛋白和ZO-1的表达及生化特性，直接且独立地改变血脑屏障特性。
