Send Orders for Reprints to reprints@benthamscience.net Current Medicinal Chemistry, 2013, 20, ????-???? 1 0929-8673/13 $58.00+.00 © 2013 Bentham Science Publishers Molecular Effects of Diphenyl Diselenide on Cholesterol and Glucose Cell Metabolism Juliana Trevisan da Rocha 1,# , Laura Trapani 2,# , Marco Segatto 2 , Piergiorgio La Rosa 2 , Cristina Wayne Nogueira 1 , Gilson Zeni 1 and Valentina Pallottini 2,* 1 Laboratório de Síntese, Reatividade e Avaliação Farmacológica e Toxicológica de Organocalcogênios, Centro de Ciências Naturais e Exatas, Universidade Federal de Santa Maria, Santa Maria, CEP 97105-900, RS, Brazil; 2 Department of Biology, University Roma Tre, Viale Marconi, 446 00146 Rome, Italy Abstract: This study was designed to investigate the molecular effects of diphenyl diselenide ((PhSe) 2 ) on cholesterol metabolism in HepG2 cell line in a dose-dependent manner. The protein levels of both total and phosphorylated 3- hydroxy-3-methylglutaryl coenzyme A reductase (HMGR and P-HMGR), low-density lipoprotein receptors (LDLr) and the proteins involved in their regulatory network were analyzed by Western blotting, and the effect of (PhSe) 2 on HMGR activity was measured. Additionally, we also evaluated the effects of this compound on glucose transporter type 4 (GLUT4) translocation using fluorescence microscopy in L6 skeletal muscle cell line. Results demonstrated that (PhSe) 2 increased P-HMGR, HMGR, and LDLr protein levels as well as simvastatin treatment, which was used as positive control, without directly affecting HMGR activity. We observed that both long- and short-term HMGR regulation mechanisms are involved in the effects of (PhSe) 2 , as this compound was able to augment Sterol regu- latory element binding proteins (SREBP)-1 and Insulin induced gene (Insig)1 protein levels, and to increase AMP acti- vated kinase (AMPK) activation state. We also found that, in L6 skeletal myotubes, 10 M (PhSe) 2 increases GLUT4 translocation through AMPK activation Taken together, these findings suggest that (PhSe) 2 can modulate the expression of some proteins involved in cholesterol and glucose cell metabolism. Keywords: AMPK, cholesterol, diphenyl diselenide, glucose, HepG2 cells, L6 myoblasts, HMGR, LDLr. 1. INTRODUCTION Cholesterol is both synthesized by cells and introduced by food intake. The liver represents the principal site for cholesterol homeostasis maintenance [1], carried out by many mechanisms, such as the uptake through low-density lipoprotein (LDL) receptors (LDLr), lipoprotein release in the blood, storage by esterification, conversion into bile ac- ids and the modulation of the key and rate-limiting enzyme of cholesterol biosynthetic pathway, the 3-hydroxy-3- methylglutaryl coenzyme A reductase (HMGR, E.C. 1.1.1.34) [2]. When cells accumulate excess sterols, the activity of HMGR declines by more than 90% and the amount of LDLr decreases. By contrast, upon depletion of intracellular cho- lesterol, cells increase HMGR transcription and activity and enhance the expression and the membrane exposition of LDLr [2]. HMGR is regulated at multiple levels: the long-term regulation is operated by the control of both the biosynthesis and the degradation of the enzyme, while the short-term *Address correspondence to this author at the Department of Science, University Roma Tre, Viale Marconi, 446, 00146- Rome, Italy; Tel: +39 06 57336320; Fax: +39 06 57336321; Email: valentina.pallottini@uniroma3.it #These authors contributed equally to this work. regulation is attained by the modulation of phosphorylation or dephosphorylation processes [3, 4]. Sterol regulatory ele- ment-binding proteins (SREBPs) enhance cholesterol syn- thesis and uptake by modulating genes encoding cholesterol biosynthetic enzymes, including HMGR and the LDLr [5]. SREBPs are produced as membrane-bound precursors that require cleavage by a two-step proteolytic process into Golgi apparatus, in order to release into the nucleus their amino- terminal domain (nSREBP) which are responsible for the expression of their target genes involved in cholesterol and fatty acid synthesis [6]. Two proteins, Insig1 and Insig2 (In- sulin Induced Gene), are shown to cooperate with sterols to inhibit the migration of SCAP (SREBP cleavage-activating protein)/SREBP complex from the endoplasmic reticulum [7, 8]. When cholesterol builds up in the endoplasmic reticulum (ER) membranes, the SREBP/SCAP complex fails to exit the ER, the proteolitic processing of SREBPs is abolished and the transcription of target genes declines. ER retention of SCAP/SREBP is mediated by sterol dependent binding of SCAP/SREBP to Insigs [9]. Intracellular accumulation of sterols also induces HMGR to bind Insigs which promote the ubiquitination and proteasomal degradation of the enzyme [10]. HMGR short-term regulation depends on cellular meta- bolic state through the AMP-activated kinase (AMPK) that optimizes ATP expenditure during metabolic stress into cells