Mechanisms Underlying the Anti-inflammatory Effects of the Ca2+/Calmodulin Antagonist CV-159 in Cultured Vascular Smooth Muscle Cells

Usui, Tatsuya; Yamawaki, Hideyuki; Kamibayashi, Masato; Okada, Muneyoshi; Hara, Yukio

doi:10.1254/jphs.10100fp

Cited by 9 publications

(7 citation statements)

References 39 publications

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“…In the previous study, we showed that an antioxidant drug, N-acetyl l-cysteine inhibited TNF-induced expression of VCAM-1 and e-selectin, as well as activation of Akt, JNK, and p38, in cultured SMCs or endothelial cells. 14,15 In addition, we confirmed that the inhibitor of Akt, JNK, or p38 prevented TNF-induced expression of VCAM-1 or e-selectin. Collectively, our results indicate that DAPK3 mediates TNFinduced inflammation via induction of proinflammatory molecules (VCAM-1, e-selectin, and COX-2) through activation of ROS-dependent signals in cultured SMCs and endothelial cells.…”

Section: October 2012supporting

confidence: 64%

Death-Associated Protein Kinase 3 Mediates Vascular Inflammation and Development of Hypertension in Spontaneously Hypertensive Rats

et al. 2012

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C almodulin (CaM) is recognized as a key regulatory molecule for diverse biological functions, such as muscular contraction.1 In addition, a recent study demonstrated that CaM-dependent protein kinases, such as CaM-dependent protein kinase II, may regulate hypertension through the mechanisms including promotion of vascular smooth muscle hypertrophy. 2 In the previous study, we compared expression levels of several CaM-related proteins with almost unknown functions in vasculature. As a result, we demonstrated that protein expression of eukaryotic elongation factor 2 kinase (also known as CaM-dependent protein kinase III), histone deacetylase 4, and death associated protein kinase (DAPK) 3 increased in the mesenteric artery from spontaneously hypertensive rats (SHRs) compared with Wistar Kyoto rats (WKYs). 3 In the recent study, we also showed that histone deacetylase 4 promotes reactive oxygen species (ROS)-dependent vascular inflammation and mediates the development of hypertension in SHRs. 4 However, it remains to be clarified how DAPK3, might affect vascular patholophysiology and control the development of hypertension. The DAPK protein has been considered as a Ca 2+ /CaMregulated serine/threonine kinase that mainly mediates cell death. 5 The DAPK protein consists of multiple domains, including an N-terminal kinase domain (known as a Ca 2+ /CaM regulatory region), ankyrin repeats, a cytoskeleton binding region, and a C-terminal death domain.6 Various stimulation-induced intracellular calcium spikes promote the CaM binding to the CaM regulatory region of DAPK, which in turn activates DAPK.7 Five protein kinases, including DAPK1, DAPK2, DAPK3 (also known as Zipper-interacting protein kinase), DAPK-related apoptosis-inducing protein kinase 1, and DAPK-related apoptosis-inducing protein kinase 2, are known as DAPK family proteins.8 DAPK1, DAPK2, and DAPK3 are all ubiquitously expressed in various tissues from mice and rats.6 Recently, the functions of DAPK3 were gradually clarified. The mutation of DAPK3 Abstract-Death-associated protein kinase (DAPK) is a Ca 2+ /calmodulin-regulated serine/threonine kinase that mediates cell death. Our recent study demonstrated that DAPK3 protein increases in the mesenteric artery from spontaneously hypertensive rats compared with Wistar Kyoto rats. Pathogenesis of hypertension is modulated at least in part by vascular inflammation. We examined whether DAPK3 mediates vascular inflammatory responses and development of hypertension. In rat mesenteric arterial smooth muscle cells, small interfering RNA against DAPK3 inhibited vascular cell adhesion molecule 1 expression and monocyte adhesion induced by tumor necrosis factor-α. DAPK3 small interfering RNA inhibited phosphorylation of c-Jun amino-terminal kinase, p38, and Akt, as well as reactive oxygen species (ROS) production induced by tumor necrosis factor-α. In human umbilical vein endothelial cells, expressions of vascular cell adhesion molecule 1, endothelial selectin, and cyclooxygenase 2, as well as ROS production induced ...

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Section: October 2012supporting

confidence: 64%

Death-Associated Protein Kinase 3 Mediates Vascular Inflammation and Development of Hypertension in Spontaneously Hypertensive Rats

et al. 2012

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“…7). We have previously shown that antioxidant, N-acetyl L-cysteine prevented TNF-␣-induced ROS, VCAM-1, and phosphorylation of JNK and NF-B, suggesting that ROS mediates inflammatory responses in vascular SMCs and ECs (39,40). In addition, we showed that a CaM inhibitor, W-7, inhibited TNF-␣-induced activation of eEF2K, JNK, and NF-B as well as TNF-␣-induced expression of VCAM-1 and E-selectin in ECs (Fig.…”

Section: Discussionmentioning

confidence: 98%

“…The superior mesenteric artery was isolated. SMCs isolated from mesenteric artery by an explant method were cultured in DMEM supplemented with 10% FBS (Invitrogen, Carlsbad, CA) (40). Passages 4 to 20 SMCs at 80 to 90% confluence were growth arrested by incubating in DMEM containing 0.5% FBS for 24 h before stimulation.…”

Section: Methodsmentioning

confidence: 99%

Eukaryotic elongation factor 2 kinase regulates the development of hypertension through oxidative stress-dependent vascular inflammation

Usui

Okada

Hara

et al. 2013

American Journal of Physiology-Heart and Circulatory Physiology

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Eukaryotic elongation factor 2 kinase (eEF2K) is a Ca2+/calmodulin-dependent protein kinase. We recently demonstrated that eEF2K protein increases in mesenteric artery from spontaneously hypertensive rats (SHR). Pathogenesis of hypertension is regulated in part by vascular inflammation. We tested the hypothesis whether eEF2K mediates vascular inflammatory responses and development of hypertension. In vascular endothelial cells, small interfering RNA (siRNA) against eEF2K inhibited induction of VCAM-1 and endothelial-selectin as well as monocyte adhesion by TNF-α (10 ng/ml). eEF2K siRNA inhibited phosphorylation of JNK and NF-κB p65 as well as reactive oxygen species (ROS) production by TNF-α. In vascular smooth muscle cells, eEF2K siRNA also inhibited VCAM-1 induction and phosphorylation of JNK and NF-κB by TNF-α. In vivo, increased blood pressure in SHR and ROS production, induction of inflammatory molecules, and hypertrophy in SHR superior mesenteric artery were reduced by an eEF2K inhibitor NH125 (500 μg·kg(-1)·day(-1)). In SHR superior mesenteric artery, impairment of ACh-induced relaxation was normalized by NH125. The present results for the first time demonstrate that eEF2K mediates TNF-α-induced vascular inflammation via ROS-dependent mechanism, which is at least partly responsible for the development of hypertension in SHR.

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“…Western blotting : Western blotting was performed as described previously [19, 29, 30, 32]. Protein lysates were obtained by homogenizing mesenteric artery with Triton-based lysis buffer (20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM EDTA, 1 mM ethylene glycol tetraacetic acid, 1% Triton, 2.5 mM sodium pyrophosphate, 1 mM β-glycerophosphate, 1 mM Na 3 VO 4 and 1 µ g/m l leupeptin).…”

Section: Methodsmentioning

confidence: 99%

Mechanisms Underlying a Decrease in KCl-Induced Contraction after Long-Term Serum-Free Organ Culture of Rat Isolated Mesenteric Artery

Morita

Okada

Yamawaki

2014

The Journal of Veterinary Medical Science

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Organ culture of blood vessel is a better technique to investigate the long-term effects of drugs. However, some functional changes may occur from freshly isolated vessel (Fresh). Mammalian/mechanistic target of rapamycin (mTOR) regulates smooth muscle differentiation and Ca2+ mobilization. We thus investigated mechanisms of alteration in smooth muscle contractility after serum-free organ culture focusing on mTOR. Rat isolated mesenteric arteries were cultured for 5 days without (0% serum) or with rapamycin. In 0% serum, absolute contraction by KCl significantly decreased from Fresh, which was significantly rescued by rapamycin. In 0% serum, mTOR expression significantly increased from Fresh, which was significantly rescued by rapamycin. In 0% serum, expression of myocardin, a key regulator of smooth muscle differentiation markers, significantly decreased from Fresh, which was significantly rescued by rapamycin. However, the decrease in expression of contractile proteins, including SM22α and calponin, was not changed by rapamycin. Basal phosphorylation of calmodulin-dependent protein kinase II significantly increased in 0% serum, which was significantly rescued by rapamycin. In 0% serum, absolute contraction by caffeine significantly decreased from Fresh, which was significantly rescued by rapamycin. In conclusion, expression of mTOR increased during serum-free organ culture of rat isolated mesenteric artery for 5 days, which may be at least partly responsible for the decreased smooth muscle contractility perhaps due to the decrease in the stored Ca2+ in smooth muscle.

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Mechanisms Underlying the Anti-inflammatory Effects of the Ca2+/Calmodulin Antagonist CV-159 in Cultured Vascular Smooth Muscle Cells

Cited by 9 publications

References 39 publications

Death-Associated Protein Kinase 3 Mediates Vascular Inflammation and Development of Hypertension in Spontaneously Hypertensive Rats

Death-Associated Protein Kinase 3 Mediates Vascular Inflammation and Development of Hypertension in Spontaneously Hypertensive Rats

Eukaryotic elongation factor 2 kinase regulates the development of hypertension through oxidative stress-dependent vascular inflammation

Mechanisms Underlying a Decrease in KCl-Induced Contraction after Long-Term Serum-Free Organ Culture of Rat Isolated Mesenteric Artery

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