Smooth muscle--specific expression of CYP4A1 induces endothelial sprouting in renal arterial microvessels.
Cytochrome P450 (CYP) 4A1 has been characterized as the most efficient arachidonic acid omega-hydroxylase catalyzing the formation of 20-hydroxyeicosatetraenoic acid (20-HETE), a potent constrictor of the renal and cerebral microcirculation and a mitogen for smooth muscle cells. We constructed adenoviruses expressing the CYP4A1 cDNA or LacZ under the control of the smooth muscle cell-specific promoter SM22alpha (Ad-SM22-4A1 and Ad-SM22-nLacZ, respectively). Beta-galactosidase expression was detected in Ad-SM22-nLacZ-transduced vascular smooth muscle A7r5 and PAC1 cells, but not in Ad-SM22-nLacZ-transduced 3T3 fibroblasts or vascular endothelial cells. Likewise, CYP4A1 mRNA and protein were detected in Ad-SM22-4A1-transduced A7r5 and PAC1 cells. Ad-SM22-4A1-transduced A7r5 cells metabolized lauric acid to 12-hydroxy-lauric acid at a rate 5 times greater than that of cells transduced with Ad-SM22-nLacZ (4.79+/-1.77 versus 0.97+/-0.57 nmol 12-hydroxy lauric acid/10(6) cells per h). Smooth muscle-specific LacZ expression was also detected in microdissected renal interlobar arteries transduced with Ad-SM22-nLacZ. Arteries transduced with Ad-SM22-4A1 produced higher levels of 20-HETE (4.04+/-0.29 and 13.43+/-2.84 ng/mg protein in Ad-SM22-nLacZ-transduced and Ad-SM22-4A1-transduced arteries, respectively) and demonstrated a marked angiogenic activity measured as the total length of sprouting neovessels (12.63+/-3.66 mm in Ad-SM22-4A1-transduced vessels versus 1.79+/-0.89 mm in Ad-SM22-nLacZ-transduced vessels). This angiogenic activity represented endothelial cell sprouting and was fully blocked by treatment with HET0016, a selective inhibitor of CYP4A-catalyzed reactions. The inhibitory effect of HET0016 was reversed by addition of a 20-HETE agonist. We conclude that Ad-SM22-4A1 drives a smooth muscle-specific functional expression of CYP4A1 and demonstrates increased angiogenesis, presumably via increased production of 20-HETE.
Jiang
Miao
M
Department of Pharmacology, New York Medical College, Valhalla, NY 10595, USA.
Mezentsev
Alexandre
A
Kemp
Rowena
R
Byun
Kihwan
K
Falck
John R
JR
Miano
Joseph M
JM
Nasjletti
Alberto
A
Abraham
Nader G
NG
Laniado-Schwartzman
Michal
M
D016475:3T3 Cells; D000256:Adenoviridae / Q000235:genetics; D000578:Amidines / Q000494:pharmacology; D000818:Animals; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000502:physiology*; D000072470:Cytochrome P450 Family 4; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000502:physiology; D007850:Lauric Acids / Q000378:metabolism; D008297:Male; D051379:Mice; D008840:Microfilament Proteins / Q000235:genetics; D009124:Muscle Proteins / Q000235:genetics; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D012077:Renal Artery / Q000166:cytology / Q000378:metabolism; D014161:Transduction, Genetic; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
D000578:Amidines; C465387:Cyp4a1protein, rat; D004791:Enzyme Inhibitors; C000708209:HET0016; D006893:Hydroxyeicosatetraenoic Acids; D007850:Lauric Acids; D008840:Microfilament Proteins; D009124:Muscle Proteins; C058992:Tagln protein, mouse; D042461:Vascular Endothelial Growth Factor A; C051615:transgelin; C030358:lauric acid; C055987:20-hydroxy-5,8,11,14-eicosatetraenoic acid; D003577:Cytochrome P-450 Enzyme System; D000072470:Cytochrome P450 Family 4