Abstract
Although diacylglycerol kinase (DGK) is known to be expressed in vascular smooth muscle cell, its functional significance remains to be clarified. We hypothesized that DGK is involved in the pathway of cytokine-induced nitric oxide (NO) production in vascular smooth muscle cells. The purpose of this study was to investigate the effects of R59949, a diacylglycerol kinase inhibitor, on inducible nitric oxide production in vascular smooth muscle cell. Cultured rat aortic smooth muscle cells (RASMCs) were used to elucidate the effects of R59949 on basal and interleukin-1β (IL-1β)-induced NO production. The effects of R59949 on protein and mRNA expression of induced nitric oxide synthase (iNOS) and on transplasmalemmal l-arginine uptake were also evaluated using RASMCs. Treatment of RASMCs with R59949 (10 μM) inhibited IL-1β (10 ng/ml)-induced NO production but not basal NO production. Neither protein nor mRNA expression level of iNOS after stimulation with IL-1β was significantly affected by R59949. Estimated enzymatic activities of iNOS in RASMCs were comparable in the absence and presence of R59949. Stimulation of RASMCs with IL-1β caused a marked increase in transplasmalemmal l-arginine uptake into RASMCs. l-Arginine uptake in the presence of IL-1β was markedly inhibited by R59949, while basal l-arginine uptake was not significantly affected by R59949. Both IL-1β-induced NO production and l-arginine uptake were abolished in the presence of cycloheximide (1 μM). The results indicate that R59949 inhibits inducible NO production through decreasing transplasmalemmal l-arginine uptake. DGK is suggested to be involved in cytokine-stimulated l-arginine transport and regulate its intracellular concentration in vascular smooth muscle cell.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Albert AP (2011) Gating mechanisms of canonical transient receptor potential channel proteins: role of phosphoinositols and diacylglycerol. Adv Exp Med Biol 704:391–411
Albert AP, Piper AS, Large WA (2005) Role of phospholipase D and diacylglycerol in activating constitutive TRPC-like cation channels in rabbit ear artery myocytes. J Physiol 566:769–780
Baydoun AR, Wileman SM, Wheeler-Jones CPD, Marber MS, Mann GE, Pearson JD, Closs EI (1999) Transmembrane signalling mechanisms regulating expression of cationic amino acid transporters and inducible nitric oxide synthase in rat vascular smooth muscle cells. Biochem J 344:265–272
Choi H, Allahdadi KJ, Tostes RCA, Webb RC (2009) Diacylglycerol kinase inhibition and vascular function. Curr Enzym Inhib 5:148–152
Clarke CJ, Ohanian V, Ohanian J (2007) Norepinephrine and endothelin activate diacylglycerol kinases in caveolae/rafts of rat mesenteric arteries: agonist-specific role of PI3-kinase. Am J Physiol Heart Circ Physiol 292:H2248–H2256
Flores I, Jones DR, Ciprés A, Díaz-Flores E, Sanjuan MA, Mérida I (1999) Diacylglycerol kinase inhibition prevents IL-2-induced G1 to S transition through a phosphatidylinositol-3 kinase-independent mechanism. J Immunol 163:708–714
Goto K, Hozumi Y, Nakano T, Saino-Saito S, Kondo H (2007) Cell biology and pathophysiology of the diacylglycerol kinase family: morphological aspects in tissues and organs. Int Rev Cytol 264:25–63
Goto K, Hozumi Y, Nakano T, Saino-Saito S, Martelli AM (2008) Lipid messenger, diacylglycerol, and its regulator, diacylglycerol kinase, in cells, organs, and animals: History and perspective. Tohoku J Exp Med 214:199–212
Hattori Y, Kasai K, Gross SS (1999) Cationic amino acid transporter gene expression in cultured vascular smooth muscle cells and in rats. Am J Physiol 276:H2020–H2028
Jiang Y, Sakane F, Kanoh H, Walsh JP (2000) Selectivity of the diacylglycerol kinase inhibitor 3-[2-(4-[bis-(4-fluorophenyl)methylene]-1-piperidinyl)ethyl]-2, 3-dihydro-2-thioxo-4(1H)quinazolinone (R59949) among diacylglycerol kinase subtypes. Biochem Pharmacol 59:763–772
Kohlhaas CF, Morrow VA, Jhakra N, Patil V, Connell JMC, Petrie JR, Salt IP (2011) Insulin rapidly stimulates L-arginine transport in human aortic endothelial cells via Akt. Biochem Biophys Res Commun 412:747–751
Li S, Huang FL, Feng Q, Liu J, Fan SX, McKenna TM (1998) Overexpression of protein kinase Cα enhances lipopolysaccharide-induced nitric oxide formation in vascular smooth muscle cells. J Cell Physiol 176:402–411
Mann GE, Yudilevich DL, Sobrevia L (2003) Regulation of amino acid and glucose transporters in endothelial and smooth muscle cells. Physiol Rev 83:183–252
Marumo M, Nakano T, Takeda Y, Goto K, Wakabayashi I (2012) Inhibition of thrombin-induced Ca2+ influx in platelets by R59949, an inhibitor of diacylglycerol kinase. J Pharm Pharmacol 64:855–861
Mérida I, Ávila-Flores A, Merino E (2008) Diacylglycerol kinases: at the hub of cell signalling. Biochem J 409:1–18
Nakano T, Hozumi Y, Goto K, Wakabayashi I (2009) Localization of diacylglycerol kinase ε on stress fibers in vascular smooth muscle cells. Cell Tissue Res 337:167–175
Nakano T, Hozumi Y, Goto K, Wakabayashi I (2012) Involvement of diacylglycerol kinase γ in modulation of iNOS synthesis in Golgi apparatus of vascular endothelial cells. Naunyn Schmiedeberg's Arch Pharmacol 385:787–795
Nobe K, Sakai Y, Maruyama Y, Momose K (2002) Hyper-reactivity of diacylglycerol kinase is involved in the dysfunction of aortic smooth muscle contractility in streptozotocin-induced diabetic rats. Br J Pharmacol 136:441–451
Nobe K, Sakai Y, Nobe H, Momose K (2003) Dysfunction of aorta involves different patterns of intracellular signaling pathways in diabetic rats. Eur J Pharmacol 471:195–204
Ohanian J, Heagerty AM (1994) Membrane-associated diacylglycerol kinase activity is increased by noradrenaline, but not by angiotensin II, in arterial smooth muscle. Biochem J 300:51–56
Ohanian J, Ohanian V (2001) Lipid second messenger regulation: the role of diacylglycerol kinases and their relevance to hypertension. J Hum Hypertens 15:93–98
Richman JG, Regan JW (1998) α2-Adrenergic receptors increase cell migration and decrease F-actin labeling in rat aortic smooth muscle cells. Am J Physiol 274:C654–C662
Rivera-Correa M, Altieri PI, Escobales N (1996) Parallel regulation of arginine transport and nitric oxide synthesis by angiotensin II in vascular smooth muscle cells role of protein kinase C. Amino Acids 11:153–170
Sakane F, Imai S, Kai M, Yasuda S, Kanoh H (2007) Diacylglycerol kinases: Why so many of them? Biochim Biophys Acta 1771:793–806
Sanjuán MA, Jones DR, Izquierdo M, Mérida I (2001) Role of diacylglycerol kinase α in the attenuation of receptor signaling. J Cell Biol 153:207–219
Sato M, Liu K, Sasaki S, Kunii N, Sakai H, Mizuno H, Saga H, Sakane F (2013) Evaluations of the selectivities of the diacylglycerol kinase inhibitors R59022 and R59949 among diacylglycerol kinase isozymes using a new non-radioactive assay method. Pharmacology 92:99–107
Schwartz IF, Ayalon R, Chernichovski T, Reshef R, Chernin G, Weinstein T, Litvak A, Levo Y, Schwartz D (2006) Arginine uptake is attenuated through modulation of cationic amino-acid transporter-1, in uremic rats. Kidney Int 69:298–303
Schwartz IF, Ingbir M, Chernichovski T, Reshef R, Chernin G, Litvak A, Weinstein T, Levo Y, Schwartz D (2007) Arginine uptake is attenuated, through post-translational regulation of cationic amino acid transporter-1, in hyperlipidemic rats. Atherosclerosis 194:357–363
Schwartz IF, Chernichovski T, Schwartz D (2010) Aortic arginine transport is attenuated, through post-translational modulation of CAT-1 by PKCα, in old male rats. Vasc Med 15:55–59
Schwartz IF, Grupper A, Chernichovski T, Grupper A, Hillel O, Engel A, Schwartz D (2011) Hyperuricemia attenuates aortic nitric oxide generation, through inhibition of arginine transport, in rats. J Vasc Res 48:252–260
Tsuchiya R, Tanaka T, Hozumi Y, Nakano T, Okada M, Topham MK, Iino M, Goto K (2015) Downregulation of diacylglycerol kinase ζ enhances activation of cytokine-induced NF-κB signaling pathway. Biochim Biophys Acta 1853:361–369
Wakabayashi I, Mayer B, Groschner K (2000) Inhibitory effects of aclarubicin on nitric oxide production in aortic smooth muscle cells and macrophages. Biochem Pharmacol 59:719–726
Wileman SM, Mann GE, Baydoun AR (1995) Induction of L-arginine transport and nitric oxide synthase in vascular smooth muscle cells: synergistic actions of pro-inflammatory cytokines and bacterial lipopolysaccharide. Br J Pharmacol 116:3243–3250
Zhang Y, Du G (2009) Phosphatidic acid signaling regulation of Ras superfamily of small guanosine triphosphatases. Biochim Biophys Acta 1791:850–855
Zhao Y, Vanhoutte PM, Leung SWS (2015) Vascular nitric oxide: Beyond eNOS. J Pharmacol Sci 129:83–94
Acknowledgements
This work was supported by a Grant-in-Aid for graduated students (2013) and that for researchers (2014) at Hyogo College of Medicine.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
The study protocols for the treatment of animals were in accordance with the “Guidelines for Experiments Using Laboratory Animals in Hyogo College of Medicine”.
Conflict of interest
The authors declare that they have no conflict of interest.
Rights and permissions
About this article
Cite this article
Shimomura, T., Nakano, T., Goto, K. et al. R59949, a diacylglycerol kinase inhibitor, inhibits inducible nitric oxide production through decreasing transplasmalemmal l-arginine uptake in vascular smooth muscle cells. Naunyn-Schmiedeberg's Arch Pharmacol 390, 207–214 (2017). https://doi.org/10.1007/s00210-016-1316-5
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00210-016-1316-5