Proceedings of The Physiological Society
Kings College London (2005) J Physiol 565P, PC154
Caveolae membrane system and astrocyte interactions within porcine brain microvascular capillary endothelial cell cultures
Smith, Mathew Wayne; Connell, David ; Omidi, Yadollah ; Gumbleton, Mark ;
1. Welsh School of Pharmacy, Cardiff University, Cardiff, United Kingdom.
Caveolae are classically defined as omega shaped invaginations of the plasma membrane enriched in cholesterol, sphingolipids, and their unique and functional protein, caveolin-1. Caveolae are abundant in many cell types including peripheral capillary endothelial cells, smooth muscle cells and type I pneumocytes, and are involved in the transport of solutes including macromolecules (Gumbleton et al 2000). Previous morphometric studies (Stewart 2000) determining the numerical density of caveolae-like vesicles within the Blood Brain Barrier (BBB) report a limited presence. Unlike clathrin coated pits, caveolae can exist in morphologically undetectable forms and a paucity of vesicles may not necessarily reflect functionality. Our aim is to investigate the function and modulation of the caveolae membrane system within the blood brain barrier. In our studies we utilise a primary culture of porcine brain microvascular endothelial capillary (PBMVEC) cells, isolated and cultured as previously described by our laboratory (Omidi et al 2003). We report that while whole brain tissue (containing by volume 0.1% capillary endothelial cells) shows negligible levels of caveolin-1 expression, freshly isolated PBMVEC cells display high caveolin-1 expression comparable to model cell lines widely used in caveolae research i.e. MA104 & A431. Expression is not diminished in PBMVEC primary or 1st passage culture. In freshly isolated cells the level of phosphorylated-caveolin-1 is negligible while in primary and 1st passage cells the phosphorylation state is abundant. In non-brain endothelial cell types this change in phosporylation state is implicated in the endocytic potential of caveolae (Aoki et al 1999) and warrants further investigation. Exposing PBMVEC cells to astrocyte (C6 glioma) conditioned media or astrocyte co-culture systems did not alter either the expression of caveolin-1 or its phosphorylation state in primary or 1st passage cells. Microscopy did not reveal any significant effect of C6 treatments upon vesicle density within the PBMVEC cells. Whilst astrocyte derived factors do not appear to alter expression of caveolin-1 or its phosphorylation state, this does not mean that astrocyte derived factors do not modulate the caveolae biology of the BBB. The effect of astrocyte derived factors on caveolae function and solute transport is currently under investigation.
Where applicable, experiments conform with Society ethical requirements