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Ryuji Okamoto,
Takaaki Kato,
Akira Mizoguchi,
Nobuaki Takahashi, Tetsuya Nakakuki,
Hideo Mizutani,
Naoki Isaka,
Kyoko Imanaka-Yoshida,
Kozo Kaibuchi,
Zhaojiang Lu,
Katsuhide Mabuchi,
Terenc Tao,
David J Hartshorne,
Takeshi Nakano,
Masaaki Ito
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ABSTRACT: Characterization of cardiac MYPT2 (an isoform of the smooth muscle phosphatase [MP] target subunit, MYPT1) is described. Several features of MYPT2 and MYPT1 were similar, including: a specific interaction with the catalytic subunit of type 1 phosphatase, delta isoform (PP1cdelta); interaction of MYPT2 with the small heart-specific MP subunit; interaction of the C-terminal region of MYPT2 with the active form of RhoA; phosphorylation by Rho-kinase at an inhibitory site, Thr646 and thiophosphorylation at Thr646 inhibited activity of the MYPT2-PP1cdelta complex. MYPT2 activated PP1cdelta activity, using light chains from smooth and cardiac muscle, by reducing K(m) and increasing k(cat). The extent of activation (k(cat)) was greater than for MYPT1 and could reflect distinct N-terminal sequences in the two MYPT isoforms. Adenovirus-mediated gene transfer of MYPT2 and PP1cdelta reduced the phosphorylation level of cardiac light chains following stimulation with A23187. Overexpression of MYPT2 and PP1cdelta blocked the angiotensin II-induced sarcomere organization in cultured cardiomyocytes. Electron microscopy indicated locations of MYPTs, at, or close to, the Z-line, the A band and mitochondria. Similarity of the two MYPT isoforms suggests common enzymatic mechanisms and regulation. Cardiac myosin is a substrate for the MYPT2 holoenzyme, but the Z-line location raises the possibility of other substrates.
Cellular Signalling 10/2006; 18(9):1408-16. · 4.06 Impact Factor
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Tetsuya Nakakuki,
Masaaki Ito,
Hitoshi Iwasaki,
Yasuko Kureishi,
Ryuji Okamoto,
Nobuyuki Moriki,
Mariko Kongo,
Shinya Kato,
Norikazu Yamada,
Naoki Isaka,
Takeshi Nakano
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ABSTRACT: Rho/Rho-kinase pathway plays pivotal roles in cardiovascular diseases including arteriosclerosis and hypertension. Recently it has become evident that C-reactive protein (CRP), a powerful marker for cardiovascular events, has direct proatherothrombotic effects on vascular cells. However, its molecular mechanism has not been fully investigated. We examined the involvement of Rho/Rho-kinase signaling in CRP-induced plasminogen activator inhibitor-1 (PAI-1) expression in bovine aortic endothelial cells (BAECs).
PAI-1 expression was determined by Western blotting. RhoA activation was determined by an affinity pull-down assay using Rho-binding fragment of rhotekin. NF-kappaB activity was determined using the luciferase reporter gene. Incubation of BAECs with human recombinant CRP (> or =25 microg/mL) induced a significant increase in PAI-1 expression. Stimulation of BAECs with CRP significantly increased RhoA activation. Pretreatment with TAT-C3 (a membrane-permeable RhoA inhibitor) and Y-27632 (Rho-kinase inhibitor) significantly inhibited CRP-induced PAI-1 expression. NF-kappaB activity was markedly enhanced by CRP and pretreatment with Y-27632 inhibited its activation. Parthenolide, SN50, and BAY 11-7082 (NF-kappaB inhibitors) significantly blocked CRP-mediated PAI-1 expression.
These data suggested that CRP activates Rho/Rho-kinase signaling, which in turn activates NF-kappaB activity, resulting in PAI-1 expression in BAEC. These observations provide evidence for the possible involvement of Rho/Rho-kinase signaling in CRP-induced atherothrombogenesis.
Arteriosclerosis Thrombosis and Vascular Biology 11/2005; 25(10):2088-93. · 6.37 Impact Factor
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ABSTRACT: Primary cardiac lymphoma is a rare disorder with a poor prognosis. We present here a case of 77-year-old woman who was diagnosed as having cardiac lymphoma antemortem according to a cytologic examination of the pericardial effusion. Determination of the levels of serum-soluble interleukin-2 receptor and serum deoxythymidine kinase was useful for the diagnosis. Echocardiography, computed tomography, magnetic resonance imaging, and gallium scan revealed neither lymphadenopathy nor tumor in the heart, so she was diagnosed as having malignant lymphoma that probably originated from the pericardium. Systemic chemotherapy with CHOP (cyclophosphamide, farmorubicin, oncovin, and prednisolone) resulted in a complete resolution of the pericardial effusion. She has been in remission 48 months after discontinuation of the chemotherapy.
Heart and Vessels 08/2004; 19(4):199-202. · 2.05 Impact Factor
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ABSTRACT: RhoA is commonly activated in the aorta in various hypertensive models, indicating that RhoA seems to be a molecular switch in hypertension. The molecular mechanisms for RhoA activation in stroke-prone spontaneously hypertensive rats (SHRSP) were here investigated using cultured aortic smooth muscle cells (VSMC). The level of the active form of RhoA was higher in VSMC from SHRSP than in those from Wistar-Kyoto rats (WKY). The phosphorylation level of myosin phosphatase target subunit 1 (MYPT1) at the inhibitory site was also significantly higher in SHRSP, and the phosphorylation levels in both VSMCs were strongly inhibited to a similar extent by treatment with Y-27632, a Rho-kinase inhibitor. The expression levels of RhoA/Rho-kinase related molecules, namely RhoA, Rho-kinase, MYPT1, CPI-17 (inhibitory phosphoprotein for myosin phosphatase) and myosin light chain kinase, were not different between SHRSP and WKY. Valsartan, an angiotensin II (Ang II)- type 1 receptor antagonist, selectively and significantly reduced the RhoA activation in VSMC from SHRSP. The expression levels of the Rho GDP-dissociation inhibitor (RhoGDI) and leukemia-associated Rho-specific guanine nucleotide exchange factor (RhoGEF) did not differ between SHRSP and WKY. In cyclic nucleotide signaling, cyclic GMP (cGMP)-dependent protein kinase Ialpha (cGKIalpha) was significantly downregulated in SHRSP cells, although there were no changes in the expression levels of guanylate cyclase beta and cyclic AMP (cAMP)-dependent protein kinase or the intracellular contents of cGMP and cAMP between the two rat models. These results suggest that the possible mechanisms underlying RhoA activation in VSMC from SHRSP are autocrine/paracrine regulation by Ang II and/or cGKIalpha downregulation.
Hypertension Research 05/2004; 27(4):263-70. · 2.58 Impact Factor
