Modulation of vascular smooth muscle cell alignment by cyclic strain is dependent on reactive oxygen species and P38 mitogen-activated protein kinase.
OBJECTIVE
The aim of this study was to investigate the molecular targets of reactive oxygen species (ROS) and to determine whether cyclic strain induces smooth muscle cell (SMC) alignment via the ROS system. We assessed stretch-induced nicotinamide adenine dinucleotide phosphate (NAD(P)H) oxidase activation and the redox sensitivity of cyclic strain-stimulated activation of the mitogen-activated protein kinase (MAPK) family.


METHODS
SMCs were seeded on flexible collagen I-coated plates and exposed to cyclic strain. NAD(P)H oxidase activation was measured with lucigenin-enhanced chemiluminescent detection of superoxide. Activation of MAPK was detected by determining phosphorylation of extracellular signal-regulated protein kinase (ERK1/2), c-jun N-terminal kinase (JNK1/2), and p38 MAPK with immunoblotting. In other experiments, SMCs were exposed to diphenylene iodonium (DPI), an NAD(P)H inhibitor, 30 minutes before stretch. MAPK activation and cell orientation were then assessed.


RESULTS
Cyclic strain elicits a rapid increase in intracellular NADH/NADPH oxidase in SMCs. There was also a rapid and robust phosphorylation of ERK1/2, JNK1/2, and p38 MAPK. Cyclic strain-induced intracellular NAD(P)H generation was almost completely blocked with DPI. DPI also inhibited the strain-induced phosphorylation of ERK1/2, JNK1/2, and p38 MAPK. Both the p38 MAPK specific inhibitor, SB 202190, and DPI blocked cyclic strain-induced cell alignment, but PD98059, an ERK1/2-specific inhibitor, and SP600125, an anthrazolone inhibitor of JNK, did not.


CONCLUSIONS
Our results provide evidence that p38 MAPK is a critical component of the oxidant stress ROS-sensitive signaling pathway and plays a crucial role in vascular alignment induced by cyclic stain.
Chen
Quanhai
Q
Department of Surgery, Yale University School of Medicine and Connecticut VA Health Care System, 333 Cedar Street, New Haven, CT 06520-8062, USA.
Li
Wei
W
Quan
Zhiwei
Z
Sumpio
Bauer E
BE
D000818:Animals; D001011:Aorta; D002417:Cattle; D002478:Cells, Cultured; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D009097:Multienzyme Complexes / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009861:Onium Compounds / Q000494:pharmacology; D010084:Oxidation-Reduction; D010766:Phosphorylation; D011725:Pyridines / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013314:Stress, Mechanical; D048051:p38 Mitogen-Activated Protein Kinases
D004791:Enzyme Inhibitors; D007093:Imidazoles; D009097:Multienzyme Complexes; D009861:Onium Compounds; D011725:Pyridines; D017382:Reactive Oxygen Species; C007517:diphenyleneiodonium; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole