Inhibition of myosin phosphatase by upregulated rho-kinase plays a key role for coronary artery spasm in a porcine model with interleukin-1beta.
BACKGROUND
We recently demonstrated that the Rho-kinase-mediated pathway plays an important role for coronary artery spasm in our porcine model with interleukin-1beta (IL-1beta). In this study, we examined whether or not Rho-kinase is upregulated at the spastic site and if so, how it induces vascular smooth muscle hypercontraction.


RESULTS
Segments of the left porcine coronary artery were chronically treated from the adventitia with IL-1beta-bound microbeads. Two weeks after the operation, as reported previously, intracoronary serotonin repeatedly induced coronary hypercontractions at the IL-1beta-treated site both in vivo and in vitro, which were markedly inhibited by Y-27632, one of the specific inhibitors of Rho-kinase. Reverse transcription-polymerase chain reaction analysis demonstrated that the expression of Rho-kinase mRNA was significantly increased in the spastic compared with the control segment. Western blot analysis showed that during the serotonin-induced contractions, the extent of phosphorylation of the myosin-binding subunit of myosin phosphatase (MBS), one of the major substrates of Rho-kinase, was significantly greater in the spastic than in the control segment and that the increase in MBS phosphorylations was also markedly inhibited by Y-27632. There was a highly significant correlation between the extent of MBS phosphorylations and that of contractions.


CONCLUSIONS
These results indicate that Rho-kinase is upregulated at the spastic site and plays a key role in inducing vascular smooth muscle hypercontraction by inhibiting myosin phosphatase through the phosphorylation of MBS in our porcine model.
Kandabashi
T
T
Departments of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Shimokawa
H
H
Miyata
K
K
Kunihiro
I
I
Kawano
Y
Y
Fukata
Y
Y
Higo
T
T
Egashira
K
K
Takahashi
S
S
Kaibuchi
K
K
Takeshita
A
A
D000818:Animals; D003329:Coronary Vasospasm / Q000139:chemically induced* / Q000503:physiopathology*; D007375:Interleukin-1 / Q000008:administration & dosage; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000378:metabolism; D008863:Microspheres; D043263:Myosin-Light-Chain Phosphatase; D009218:Myosins / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine; D015854:Up-Regulation; D054460:rho-Associated Kinases
D007375:Interleukin-1; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase; D009218:Myosins