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Abstract Vascular restenosis involves contraction, proliferation, and remodeling of the arterial wall in response to overstretch injury. Mitogen-activated protein kinases (MAPKs) are implicated in both contraction and proliferation of vascular smooth muscle (VSM), and studies of porcine carotid arterial muscle strips have shown that mechanical stretch leads to the activation of the extracellular signal–regulated kinase (ERK) family of MAPKs in vivo. We, therefore, analyzed the acute effect of mechanical overstretch injury on ERK-MAPK (herein referred to simply as MAPK) activity in porcine coronary and carotid arteries in vivo. Balloon angioplasty catheters were inflated to 6 atm three times over 5 minutes at a balloon-artery ratio of 1.2:1 in either porcine coronary or carotid arteries. The arteries were snap-frozen after angioplasty, and MAPK activity was measured. Angioplasty of the left anterior descending (LAD, n=5), left circumflex (LCx, n=5), and carotid (n=5) arteries effected an increase in MAPK activity compared with the activity in uninstrumented right coronary arteries (RCAs) or carotid arteries from the same animals used for controls. Balloon angioplasty of carotid arteries led to an increase in MAPK activity that was 7.7-fold over the activity in control arteries and comparable to the activity in stretched carotid arterial muscle strips in vivo. The increase in coronary artery kinase activity on angioplasty was variable from animal to animal. The increase in MAPK activity over that in control arteries ranged from 4.5- to 31.7-fold (mean±SEM, 10.7±5.3) in the LAD and 1.8- to 31.3-fold (mean±SEM, 9.7±5.7) in the LCx. There were no apparent inherent differences in the levels of MAPK activity in the three different types of coronary arteries (RCA, LAD, and LCx) without instrumentation. MAPK activation occurs rapidly during angioplasty, suggesting that this kinase may play an early role in initiating the injury response in both porcine coronary and carotid arteries. MAPKs may be key enzymes targeted to treat or prevent restenosis.

Abstract The thin-filament protein h-caldesmon (the high molecular weight isoform of caldesmon) is phosphorylated in resting and contracted porcine carotid arteries. Phosphorylation of h-caldesmon in intact tissue occurs at sites that are covalently modified by mitogen-activated protein kinase (MAPK) in vitro. In this study, we have evaluated MAPK activation in arteries in response to mechanical load and pharmacological stimulation. MAPK was extracted from resting and stimulated porcine carotid arteries and then partially purified by anion-exchange fast-performance liquid chromatography. MAPK activity was separated into two peaks corresponding to the tyrosine-phosphorylated 42- and 44-kD isoforms of MAPK (p42 MAPK and p44 MAPK , respectively). Of the total MAPK activity, 42% was associated with p42 MAPK , and 58% was associated with p44 MAPK ; this percentage was not altered by stimulation of the muscles with either KCl (110 mmol/L) or phorbol 12,13-dibutyrate (PDBu, 1 μmol/L). Both p42 MAPK and p44 MAPK , purified from porcine carotid arteries, phosphorylated h-caldesmon at the same sites and to levels approaching or >1 mol phosphate per mole protein. In unloaded muscle strips, MAPK activity was 39 pmol · min −1 · mg protein −1 when assayed with the peptide substrate APRTPGGRR. MAPK activity increased in response to incremental mechanical loading to a maximum of 99 pmol · min −1 · mg protein −1 at 16×10 3 N/m 2 . MAPK activity could be further increased in loaded muscles by pharmacological stimulation. With KCl stimulation, MAPK activities rose to a peak of 205 pmol · min −1 · mg protein −1 at 10 minutes and then declined to basal values at 30 and 60 minutes. Stimulation with PDBu induced a gradual increase in MAPK activity that reached a value of 201 pmol · min −1 · mg protein −1 at 60 minutes. These results demonstrate that the level of MAPK activity in vascular smooth muscle is regulated in response to both mechanical load and pharmacological stimulation. Activation of MAPK and the subsequent phosphorylation of h-caldesmon may be important processes that modulate vascular smooth muscle contractility.