Differential expression of alpha-actin mRNA and immunoreactive protein in brain microvascular pericytes and smooth muscle cells.
Hypertension has been linked to opening of the blood-brain barrier and may be related to the expression of the smooth muscle alpha-actin gene in contractile cells at the brain microvasculature. However, the cellular origin (i.e., endothelial cells, pericytes, smooth muscle cells) of the alpha-actin mRNA in the brain microvasculature is not clearly identified. Therefore, we investigated the abundance of actin mRNA by Northern blot analysis in isolated brain microvessels and in brain microvascular endothelial or pericytes in tissue culture. All samples showed the characteristic 2.1 kb transcript corresponding to cytoplasmic beta and gamma isoform mRNA. The 1.7 kb transcript corresponding to smooth muscle alpha-actin was detected in freshly isolated bovine brain microvessels, in primary cultures of brain microvascular pericytes, or endothelial cells; the latter cultures contain both endothelial cells and pericytes. The alpha-actin mRNA was absent in a cloned bovine brain endothelial cell line. The relative abundance of the alpha/(beta + gamma) actin transcript ratio was: cultured pericytes > freshly isolated microvessels > endothelial primary. The cellular distribution of the smooth muscle alpha-actin immunoreactive protein was studied by immunocytochemistry in cytospun/methanol-fixed isolated bovine brain microvessels with a monoclonal antibody directed to the amino-terminal decapeptide of the smooth muscle alpha-actin isoform. This antibody reacted strongly with precapillary arterioles of isolated microvessels, whereas no immunostaining was observed in either capillary endothelial cells or in pericytes. In conclusion, the alpha-actin mRNA is expressed in brain microvascular pericytes in tissue culture, but the immunoreactive alpha-actin protein is not expressed in brain microvascular pericytes in vivo. These data suggest that either 1) alpha-actin gene expression is induced in capillary pericytes in tissue culture or 2) alpha-actin mRNA in brain capillary pericytes in vivo is subject to translational repression resulting in no detectable alpha-actin protein under normal conditions.
Boado
R J
RJ
Department of Medicine, UCLA School of Medicine, 90024.
Pardridge
W M
WM
D000199:Actins / Q000096:biosynthesis*; D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D002560:Cerebrovascular Circulation; D002831:Choroid Plexus / Q000098:blood supply / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D015870:Gene Expression; D007150:Immunohistochemistry; D008833:Microcirculation / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000302:isolation & purification; D014158:Transcription, Genetic
D000199:Actins; D012333:RNA, Messenger