Bestrophin protein is involved in ion transport across the basolateral membrane of the retinal pigment epithelium. The mammalian genome encodes 4 members of the bestrophin family. Bestrophins have been proposed to comprise a new family of Ca2+-activated Cl- channels1. We have recently demonstrated (abstract by Matchkov et. al) that siRNA mediated downregulation of bestrophin-4 is associated with the disappearance of a recently demonstrated2 cGMP-dependent Ca2+-activated Cl- current in vascular smooth muscle cells (SMCs). Here we study the distribution of bestrophin-4-and cGMP dependent Cl- channel activity in the vasculature to further test the possibility that bestrophin-4 is important for SMC Cl--channel activity. We combined electrophysiological detection (patch clamp) of Ca2+-activated Cl- currents in SMCs from vascular beds of different origin with the expression pattern of bestrophin-4 using PCR, Western blot and immunohistochemistry. Using RT-PCR we identified bestrophin-4 mRNA expression in rat mesenteric small arteries, aorta and renal arteries, while no expression was seen in pulmonary arteries. Using a bestrophin-4 specific antibody (verified by Western blotting of the heterologously expressed epitope) Western blot detected a ~65 kDa band in cell lysates from rat mesenteric small arteries and aorta, which was not seen in pulmonary arteries and when preincubated with the immunizing peptide. The distribution of bestrophin-4 mRNA and protein has a pattern similar to the cGMP-dependent Cl- current in SMCs of different origins. Immunohistochemistry identified bestrophin-4 both in endothelial and SMCs of the vascular tree in the brain, heart, kidney and mesentery, but not in the lungs. We suggest that bestrophin-4 is important for the cGMP dependent, Ca2+ activated Cl- conductance in many vascular beds and may play a role for both SMC and endothelial function.
Journal of Vascular Research, 2008
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9TH INTERNATIONAL SYMPOSIUM ON RESISTANCE ARTERIES (ISRA), 2008