Endothelin-1 activation of JAK2 in vascular smooth muscle cells involves NAD(P)H oxidase-derived reactive oxygen species.
Endothelin-1 (ET-1) and JAK2 are both implicated in diabetic complications. Therefore, we investigated whether ET-1 differentially activates JAK2 under conditions of normal (5 mM) and high (25 mM) glucose. We tested the hypothesis that reactive oxygen species mediate the activation of JAK2 in response to ET-1. In rat aortic vascular smooth muscle cells (VSMC), ET-1 (10 (- 7) M, 5 min) stimulated the activation of JAK2, which was further enhanced under high glucose conditions. Allopurinol (xanthine oxidase inhibitor, 1 microM) and l-NAME (nitric oxide synthase inhibitor, 1 mM) had no effect on ET-1-induced JAK2 activation, while apocynin (NAD(P)H oxidase inhibitor 100 microM) resulted in a significant inhibition of ET-1-induced JAK2 and MAPK activation. Overexpression of SOD did not inhibit ET-1-induced activation of JAK2, but catalase (50 units/mL) treatment resulted in complete inhibition. In vivo administration of apocynin (1.5 mM) resulted in a significant decrease ( 50%), while the ETA receptor antagonist ABT-627 completely inhibited phosphorylation of JAK2 in aortae from STZ-induced diabetic rats. Additionally, DHE staining of aortic sections was significantly reduced in diabetic rats treated with ABT-627. These data suggest that in VSMC, ET-1 via the ETA receptor, utilizes NAD(P)H oxidase to activate JAK2.
Banes-Berceli
Amy K L
AK
Vascular Biology Center, Medical College of Georgia, Augusta, GA 30912, USA. abanes@mcg.edu
Ogobi
Safia
S
Tawfik
Amany
A
Patel
Bela
B
Shirley
Amanda
A
Pollock
David M
DM
Fulton
David
D
Marrero
Mario B
MB
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001013:Aorta, Thoracic / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D002469:Cell Separation; D003921:Diabetes Mellitus, Experimental / Q000378:metabolism / Q000473:pathology; D019332:Endothelin-1 / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D004996:Ethidium / Q000031:analogs & derivatives; D005456:Fluorescent Dyes; D005947:Glucose / Q000494:pharmacology; D015151:Immunoblotting; D066298:In Vitro Techniques; D053614:Janus Kinase 2; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014162:Transfection
D019332:Endothelin-1; D005456:Fluorescent Dyes; D011518:Proto-Oncogene Proteins; D017382:Reactive Oxygen Species; C067883:dihydroethidium; D019255:NADPH Oxidases; D011505:Protein-Tyrosine Kinases; C507927:Jak2 protein, rat; D053614:Janus Kinase 2; D020928:Mitogen-Activated Protein Kinases; D004996:Ethidium; D005947:Glucose