Abstract
Background/Aims: Valproic acid (VPA) is the drug of choice for treating epilepsy, but has the unwanted effects of inducing weight gain and increasing the risk of developing insulin resistance. The mechanism through which these side effects occur is unknown. VPA inhibits histone deacetylase (HDAC), but also decreases the transcriptional activity of CCAAT enhancer binding protein α (CEBPα). Given the possible association between VPA, CEBPα and adipokine gene regulation, we hypothesized that they would alter the expression of resistin (rstn), fasting-induced adipose factor (fiaf) and suppressor of cytokine signaling-3 (socs-3), genes implicated in the development of leptin and insulin resistance. Methods: We investigated the effects of VPA (1 mM; 24 or 48 h) on gene expression using real-time RT-PCR in 3 distinct models: N-1 hypothalamic neurons, 3T3-L1 adipocytes and male CD-1 mice. Subsequently, cells were treated with 5 nM of the more specific HDAC inhibitor trichostatin A (TSA). CEBPα expression was also modified in N-1 neurons using either RNA interference (RNAi) or an overexpression vector to evaluate its effects on target gene expression. Results: In N-1 neurons, VPA induced significant increases in CEBP α and socs-3, but inhibited rstn and fiaf gene expression. In contrast, TSA induced rstn and socs-3, but inhibited fiaf. VPA also induced the expression of CEBP αin 3T3-L1 adipocytes, but had no effect on other target genes, and TSA suppressed fiaf and socs-3.Subsequently, CEBPα was overexpressed (24 h) or silenced using RNAi (24 and 48 h) in N-1 neurons. The silencing of CEBPα led to significant decreases in rstn mRNA, but increased fiaf and socs-3 expression, whereas its overexpression had the opposite effect. When male CD-1 mice were treated with either a single (100 mg/kg; 24 h), or multiple (200 mg/kg; 72 h) daily injections of VPA, no changes in body weight or gene expression were detected in either hypothalamic or adipose tissues. Conclusions: In summary, these experiments reveal a potentially important role for CEBPα in the regulation of hypothalamic gene expression in N-1 neurons and suggest that it might modulate central energy metabolism. Although VPA also modified hypothalamic gene expression in vitro, it remains to be determined whether it has similar effects in vivo.
References
1.
El-Khatib F, Rauchenzauner M, Lechleitner M, Hoppichler F, Naser A, Waldmann M, Trinka E, Unterberger I, Bauer G, Luef GJ: Valproate, weight gain and carbohydrate craving: a gender study. Seizure 2007;16:226–232.
2.
Pylvanen V, Knip M, Pakarinen A, Kotila M, Turkka J, Isojarvi JI: Serum insulin and leptin levels in valproate-associated obesity. Epilepsia 2002;43:514–517.
3.
Greco R, Latini G, Chiarelli F, Iannetti P, Verrotti A: Leptin, ghrelin, and adiponectin in epileptic patients treated with valproic acid. Neurology 2005;65:1808–1809.
4.
Isojarvi JI: Reproductive dysfunction in women with epilepsy. Neurology 2003;61:S27–S34.
5.
Hamed SA: Leptin and insulin homeostasis in epilepsy: relation to weight adverse conditions. Epilepsy Res 2007;75:1–9.
6.
Lagace DC, Nachtigal MW: Inhibition of histone deacetylase activity by valproic acid blocks adipogenesis. J Biol Chem 2004;279:18851–18860.
7.
Gottlicher M, Minucci S, Zhu P, Kramer OH, Schimpf A, Giavara S, Sleeman JP, Lo Coco F, Nervi C, Pelicci PG, Heinzel T: Valproic acid defines a novel class of HDAC inhibitors inducing differentiation of transformed cells. EMBO J 2001;20:6969–6978.
8.
Qiao L, Schaack J, Shao J: Suppression of adiponectin gene expression by histone deacetylase inhibitor valproic acid. Endocrinology 2006;147:865–874.
9.
Lagace DC, McLeod RS, Nachtigal MW: Valproic acid inhibits leptin secretion and reduces leptin messenger ribonucleic acid levels in adipocytes. Endocrinology 2004;145:5493–5503.
10.
Trayhurn P, Wood IS: Adipokines: Inflammation and the pleiotropic role of white adipose tissue. Br J Nutr 2004;92:347–355.
11.
Ahima RS, Qi Y, Singhal NS: Adipokines that link obesity and diabetes to the hypothalamus. Prog Brain Res 2006;153:155–174.
12.
Hartman HB, Hu X, Tyler KX, Dalal CK, Lazar MA: Mechanisms regulating adipocyte expression of resistin. J Biol Chem 2002;277:19754–19761.
13.
Steppan CM, Bailey ST, Bhat S, Brown EJ, Banerjee RR, Wright CM, Patel HR, Ahima RS, Lazar MA: The hormone resistin links obesity to diabetes. Nature 2001;409:307–312.
14.
Kim I, Moon SO, Koh KN, Kim H, Uhm CS, Kwak HJ, Kim NG, Koh GY: Molecular cloning, expression, and characterization of angiopoietin-related protein: angiopoietin-related protein induces endothelial cell sprouting. J Biol Chem 1999;274:26523–26528.
15.
Kersten S, Mandard S, Tan NS, Escher P, Metzger D, Chambon P, Gonzalez FJ, Desvergne B, Wahli W: Characterization of the fasting-induced adipose factor FIAF, a novel peroxisome proliferator-activated receptor target gene. J Biol Chem 2000;275:28488–28493.
16.
Kersten S: Regulation of lipid metabolism via angiopoietin-like proteins. Biochem Soc Trans 2005;33:1059–1062.
17.
Banerjee RR, Rangwala SM, Shapiro JS, Rich AS, Rhoades B, Qi Y, Wang J, Rajala MW, Pocai A, Scherer PE, Steppan CM, Ahima RS, Obici S, Rossetti L, Lazar MA: Regulation of fasted blood glucose by resistin. Science 2004;303:1195–1198.
18.
Rangwala SM, Rich AS, Rhoades B, Shapiro JS, Obici S, Rossetti L, Lazar MA: Abnormal glucose homeostasis due to chronic hyperresistinemia. Diabetes 2004;53:1937–1941.
19.
McTernan PG, Kusminski CM, Kumar S: Resistin. Curr Opin Lipidol 2006;17:170–175.
20.
Muse ED, Lam TK, Scherer PE, Rossetti L: Hypothalamic resistin induces hepatic insulin resistance. J Clin Invest 2007;117:1670–1678.
21.
Tovar S, Nogueiras R, Tung LY, Castaneda TR, Vazquez MJ, Morris A, Williams LM, Dickson SL, Dieguez C: Central administration of resistin promotes short-term satiety in rats. Eur J Endocrinol 2005;153:R1–R5.
22.
Wilkinson M, Brown R, Imran SA, Ur E: Adipokine gene expression in brain and pituitary gland. Neuroendocrinology 2007;86:191–209.
23.
Brown R, Imran SA, Belsham DD, Ur E, Wilkinson M: Adipokine gene expression in a novel hypothalamic neuronal cell line: resistin-dependent regulation of fasting-induced adipose factor and SOCS-3. Neuroendocrinology 2007;85:232–241.
24.
Belsham DD, Cai F, Cui H, Smukler SR, Salapatek AM, Shkreta L: Generation of a phenotypic array of hypothalamic neuronal cell models to study complex neuroendocrine disorders. Endocrinology 2004;145:393–400.
25.
Kievit P, Howard JK, Badman MK, Balthasar N, Coppari R, Mori H, Lee CE, Elmquist JK, Yoshimura A, Flier JS: Enhanced leptin sensitivity and improved glucose homeostasis in mice lacking suppressor of cytokine signaling-3 in POMC-expressing cells. Cell Metab 2006;4:123–132.
26.
Kahn BB, Alquier T, Carling D, Hardie DG: AMP-activated protein kinase: ancient energy gauge provides clues to modern understanding of metabolism. Cell Metab 2005;1:15–25.
27.
Williams SC, Cantwell CA, Johnson PF: A family of C/EBP-related proteins capable of forming covalently linked leucine zipper dimers in vitro. Genes Dev 1991;5:1553–1567.
28.
Blalock EM, Geddes JW, Chen KC, Porter NM, Markesbery WR, Landfield PW: Incipient Alzheimer’s disease: microarray correlation analyses reveal major transcriptional and tumor suppressor responses. Proc Natl Acad Sci USA 2004;101:2173–2178.
29.
Song H, Shojima N, Sakoda H, Ogihara T, Fujishiro M, Katagiri H, Anai M, Onishi Y, Ono H, Inukai K, Fukushima Y, Kikuchi M, Shimano H, Yamada N, Oka Y, Asano T: Resistin is regulated by C/EBPS, PPARS, and signal-transducing molecules. Biochem Biophys Res Commun 2002;299:291–298.
30.
Jackson AL, Linsley PS: Noise amidst the silence: off-target effects of siRNAs? Trends Genet 2004;20:521–524.
31.
Schaufele F, Wang X, Liu X, Day RN: Conformation of CCAAT/enhancer-binding protein α dimers varies with intranuclear location in living cells. J Biol Chem 2003;278:10578–10587.
32.
Brown R, Wiesner G, Ur E, Wilkinson M: Pituitary resistin gene expression is upregulated in vitro and in vivo by dexamethasone but is unaffected by rosiglitazone. Neuroendocrinology 2005;81:41–48.
33.
Steppan CM, Wang J, Whiteman EL, Birnbaum MJ, Lazar MA: Activation of SOCS-3 by resistin. Mol Cell Biol 2005;25:1569–1575.
34.
Wiesner G, Brown RE, Robertson GS, Imran SA, Ur E, Wilkinson M: Increased expression of the adipokine genes resistin and fasting-induced adipose factor in hypoxic/ischaemic mouse brain. Neuroreport 2006;17:1195–1198.
35.
Stewart MJ, Smoak K, Blum MA, Sherry B: Basal and reovirus-induced β interferon (IFN-β) and IFN-β-stimulated gene expression are cell type specific in the cardiac protective response. J Virol 2005;79:2979–2987.
36.
Jochheim A, Hillemann T, Kania G, Scharf J, Attaran M, Manns MP, Wobus AM, Ott M: Quantitative gene expression profiling reveals a fetal hepatic phenotype of murine ES-derived hepatocytes. Int J Dev Biol 2004;48:23–29.
37.
Brown R, Thompson HJ, Imran SA, Ur E, Wilkinson M: Traumatic brain injury induces adipokine gene expression in rat brain. Neurosci Lett 2007, E-pub ahead of print.
38.
Nau H: Teratogenic valproic acid concentrations: infusion by implanted minipumps vs conventional injection regimen in the mouse. Toxicol Appl Pharmacol 1985;80:243–250.
39.
Schank JR, Liles LC, Weinshenker D: Reduced anticonvulsant efficacy of valproic acid in dopamine beta-hydroxylase knockout mice. Epilepsy Res 2005;65:23–31.
40.
Walker RM, Smith GS, Barsoum NJ, Macallum GE: Preclinical toxicology of the anticonvulsant calcium valproate. Toxicology 1990;63:137–155.
41.
Munzberg H, Flier JS, Bjorbaek C: Region-specific leptin resistance within the hypothalamus of diet-induced obese mice. Endocrinology 2004;145:4880–4889.
42.
Enriori PJ, Evans AE, Sinnayah P, Jobst EE, Tonelli-Lemos L, Billes SK, Glavas MM, Grayson BE, Perello M, Nillni EA, Grove KL, Cowley MA: Diet-induced obesity causes severe but reversible leptin resistance in arcuate melanocortin neurons. Cell Metab 2007;5:181–194.
43.
Howard JK, Flier JS: Attenuation of leptin and insulin signaling by SOCS proteins. Trends Endocrinol Metab 2006;17:365–371.
44.
Munzberg H, Myers MG Jr: Molecular and anatomical determinants of central leptin resistance. Nat Neurosci 2005;8:566–570.
45.
Cai F, Gyulkhandanyan AV, Wheeler MB, Belsham DD: Glucose regulates amp-activated protein kinase activity and gene expression in clonal, hypothalamic neurons expressing proopiomelanocortin: additive effects of leptin or insulin. J Endocrinol 2007;192:605–614.
46.
Cui H, Cai F, Belsham DD: Anorexigenic hormones leptin, insulin, and α-melanocyte-stimulating hormone directly induce neurotensin (NT) gene expression in novel NT-expressing cell models. J Neurosci 2005;25:9497–9506.
47.
Cheng H, Isoda F, Belsham DD, Mobbs CV: Inhibition of AgRP expression by glucose in a clonal hypothalamic neuronal cell line is mediated by glycolysis, not oxidative phosphorylation. Endocrinology 2007, E-pub ahead of print.
48.
Abbud RA, Kelleher R, Melmed S: Cell-specific pituitary gene expression profiles after treatment with leukemia inhibitory factor reveal novel modulators for proopiomelanocortin expression. Endocrinology 2004;145:867–880.
49.
Snove O Jr, Holen T: Many commonly used siRNAs risk off-target activity. Biochem Biophys Res Commun 2004;319:256–263.
50.
Snove O Jr, Rossi JJ: Chemical modifications rescue off-target effects of RNAi. ACS Chem Biol 2006;1:274–276.
51.
Li AW, Morash B, Hollenberg AN, Ur E, Wilkinson M, Murphy PR: Transcriptional regulation of the leptin gene promoter in rat GH3 pituitary and C6 glioma cells. Mol Cell Endocrinol 2001;176:57–65.
52.
Zhou L, Xia T, Li Y, Chen X, Peng Y, Yang Z: Transcriptional regulation of the resistin gene. Domest Anim Endocrinol 2006;30:98–107.
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