Prostaglandin A₂ Enhances Cellular Insulin Sensitivity via a Mechanism that Involves the Orphan Nuclear Receptor NR4A3

Abstract

We have previously reported that members of the NR4A family of orphan nuclear receptors can augment insulin’s ability to stimulate glucose transport in adipocytes. In the current study, we endeavored to test for an insulin-sensitizing effect in muscle cells and to identify a potential transactivator. Lentiviral constructs were used to engineer both hyperexpression and shRNA silencing of NR4A3 in C2C12 myocytes. The NR4A3 hyper-expression construct led to a significant increase in glucose transport rates in the presence of maximal insulin while the NR4A3 knock-down exhibited a significant reduction in insulin-stimulated glucose transport rates. Consistently, insulin-mediated AKT phosphorylation was increased by NR4A3 hyperexpression and decreased following shRNA NR4A3 suppression. Then, we examined effects of prostaglandin A₂ (PGA₂) on insulin action and NR4A3 transactivation. PGA₂ augmented insulin-stimulated glucose uptake in C2C12 myocytes and AKT phosphorylation after 12-h treatment, without significant effects on basal transport or basal AKT phosphorylation. More importantly, we demonstrated that PGA₂ led to a greater improvement in insulin-stimulated glucose rates in NR4A3 overexpressing C2C12 myocytes, when compared with Lac-Z controls stimulated with insulin and PGA₂. Moreover, the sensitizing effect of PGA₂ was significantly diminished in NR4A3 knockdown myocytes compared to scramble controls. These results show for the first time that: (i) PGA₂ augments insulin action in myocytes as manifested by enhanced stimulation of glucose transport and AKT phosphorylation; and (ii) the insulin sensitizing effect is dependent upon the orphan nuclear receptor NR4A3.

Key words

NR4A3 - MINOR - insulin sensitivity - prostaglandin A₂ - glucose transport - skeletal muscle - C2C12 cells

• References

• 1 Wilson TE, Fahrner TJ, Johnston M, Milbrand J. Identification of the DNA binding site for NGFI-B by genetic selection in yeast. Science 1991; 252: 1296-1300
• 2 Davis IJ, Lau LF. Endocrine and neurogenic regulation of the orphan nuclear receptors Nur77 and Nurr-1 in the adrenal glands. Mol Cell Biol 1994; 14: 3469-3483
• 3 Borghaei RC, Rawlings Jr PL, Mochan E. Interleukin-4 suppression of interleukin-1-induced transcription of collagenase (MMP-1) and stromelysin 1 (MMP-3) in human synovial fibroblasts. Arthritis Rheum 1998; 41: 1398-1406
• 4 Kagaya S, Ohkura N, Tsukada T, Miyagawa M, Sugita Y, Tsujimoto G, Matsumoto K, Saito H, Hashida R. Prostaglandin A2 acts as a transactivator for NOR1 (NR4A3) within the nuclear receptor superfamily. Biol Pharm Bull 2005; 28: 1603-1607
• 5 Hazel TG, Nathans D, Lau LF. A gene inducible by serum growth factors encodes a member of the steroid and thyroid hormone receptor superfamily. Proc Natl Acad Sci USA 1988; 85: 8444-8448
• 6 Le WD, Xu P, Jankovic J, Jiang H, Appel SH, Smith RG, Vassilatis DK. Mutations in NR4A2 associated with familial Parkinson disease. Nat Genet 2003; 33: 85-89
• 7 Pei L, Castrillo A, Tontonoz P. Regulation of macrophage inflammatory gene expression by the orphan nuclear receptor Nur77. Mol Endocrinol 2006; 20: 786-794
• 8 Rius J, Martinez-Gonzalez J, Crespo J, Badimon L. NOR-1 is involved in VEGF-induced endothelial cell growth. Atherosclerosis 2006; 184: 276-282
• 9 Ohkubo T, Ohkura N, Maruyama K, Sasaki K, Nagasaki K, Hanzawa H, Tsukada T, Yamaguchi K. Early induction of the orphan nuclear receptor NOR-1 during cell death of the human breast cancer cell line MCF-7. Mol Cell Endocrinol 2000; 162: 151-156
• 10 Oita RC, Mazzatti DJ, Lim FL, Powell JR, Merry BJ. Whole-genome microarray analysis identifies up-regulation of Nr4a nuclear receptors in muscle and liver from diet-restricted rats. Mech Ageing Dev 2009; 130: 240-247
• 11 Fontana L, Klein S, Holloszy JO. Effects of long-term calorie restriction and endurance exercise on glucose tolerance, insulin action, and adipokine production. Age (Dordr) 2010; 32: 97-108
• 12 Chao LC, Wroblewski K, Zhang Z, Pei L, Vergnes L, Ilkayeva OR, Ding SY, Reue K, Watt MJ, Newgard CB, Pilch PF, Hevener AL, Tontonoz P. Insulin resistance and altered systemic glucose metabolism in mice lacking Nur77. Diabetes 2009; 58: 2788-2796
• 13 Fu Y, Luo L, Luo N, Zhu X, Garvey T. NR4A orphan nuclear receptors modulate insulin action and the glucose transport system: potential role in insulin resistance. J Biol Chem 2007; 282: 31525-31533
• 14 Straus DS, Glass CK. Cyclopentenone prostaglandins: new insights on biological activities and cellular targets. Med Res Rev 2001; 21: 185-210
• 15 Mayor P, Maianu L, Garvey WT. Glucose and insulin chronically regulate insulin action via different mechanisms in BC3H1 myocytes. Effects on glucose transporter gene expression. Diabetes 1992; 41: 274-285
• 16 Nedachi T, Kanzaki M. Regulation of glucose transporters by insulin and extracellular glucose in C2C12 myotubes. Am J Physiol Endocrinol Metab 2006; 291: E817-E828
• 17 Pei L, Waki H, Vaitheesvaran B, Wilpitz DC, Kurland IJ, Tontonoz P. NR4A orphan nuclear receptors are transcriptional regulators of hepatic glucose metabolism. Nat Med 2006; 12: 1048-1055
• 18 Chao LC, Zhang Z, Pei L, Saito T, Tontonoz P, Pilch PF. Nur77 coordinately regulates expression of genes linked to glucose metabolism in skeletal muscle. Mol Endocrinol 2007; 21: 2152-2163
• 19 Kim EH, Surh YJ. 15-deoxy-Delta12,14-prostaglandin J2 as a potential endogenous regulator of redox-sensitive transcription factors. Biochem Pharmacol 2006; 72: 1516-1528
• 20 Kim HJ, Kim JY, Meng Z, Wang LH, Liu F, Conrads TP, Burke TR, Veenstra TD, Farrar WL. 15-deoxy-Delta12,14-prostaglandin J2 inhibits transcriptional activity of estrogen receptor-alpha via covalent modification of DNA-binding domain. Cancer Res 2007; 67: 2595-2602
• 21 Milne GL, Musiek ES, Morrow JD. The cyclopentenone (A2/J2) isoprostanes – unique, highly reactive products of arachidonate peroxidation. Antioxid Redox Signal 2005; 7: 210-220
• 22 Oh JY, Giles N, Landar A, Darley-Usmar V. Accumulation of 15-deoxy-delta(12,14)-prostaglandin J2 adduct formation with Keap1 over time: effects on potency for intracellular antioxidant defence induction. Biochem J 2008; 411: 297-306
• 23 Levonen AL, Dickinson DA, Moellering DR, Mulcahy RT, Forman HJ, Darley-Usmar VM. Biphasic effects of 15-deoxy-delta-(12,14)-prostaglandin J(2) on glutathione induction and apoptosis in human endothelial cells. Arterioscler Thromb Vasc Biol 2001; 21: 1846-1851
• 24 Landar A, Zmijewski JW, Dickinson DA, Le Goffe C, Johnson MS, Milne GL, Zanoni G, Vidari G, Morrow JD, Darley-Usmar VM. Interaction of electrophilic lipid oxidation products with mitochondria in endothelial cells and formation of reactive oxygen species. Am J Physiol Heart Circ Physiol 2006; 290: H1777-H1787
• 25 Fukushima M, Kato T, Ota K, Arai Y, Narumiya S, Hayaishi O. 9-deoxy-delta 9-prostaglandin D2, a prostaglandin D2 derivative with potent antineoplastic and weak smooth muscle-contracting activities. Biochem Biophys Res Commun 1982; 109: 626-633
• 26 Fitzpatrick FA, Wynalda MA. Albumin-catalyzed metabolism of prostaglandin D2. Identification of products formed in vitro. J Biol Chem 1983; 258: 11713-11718
• 27 Forman BM, Tontonoz P, Chen J, Brun RP, Spiegelman BM, Evans RM. 15-Deoxy-delta 12, 14-prostaglandin J2 is a ligand for the adipocyte determination factor PPAR gamma. Cell 1995; 83: 803-812
• 28 Kliewer SA, Lenhard JM, Willson TM, Patel I, Morris DC, Lehmann JM. A prostaglandin J2 metabolite binds peroxisome proliferator-activated receptor gamma and promotes adipocyte differentiation. Cell 1995; 83: 813-819
• 29 Tortorella LL, Pilch PF. C2C12 myocytes lack an insulin-responsive vesicular compartment despite dexamethasone-induced GLUT4 expression. Am J Physiol Endocrinol Metab 2002; 283: E514-E524