Extracellular redox environments regulate adipocyte differentiation Barry R. Imhoff, Jason M. Hansen n Division of Pulmonary, Allergy/Immunology, Cystic Fibrosis and Sleep, Emory Department of Pediatrics, Emory University, 2015 Uppergate Drive, Room 350, Atlanta, GA 30322, USA article info Article history: Received 1 December 2009 Received in revised form 4 March 2010 Accepted 28 April 2010 Keywords: Redox Extracellular Cysteine Adipogenesis Oxidative stress abstract Oxidized extracellular redox states have been associated with many diseases related to obesity, including heart disease and diabetes, but relatively little is known about the relationship between extracellular redox states and obesity. In 3T3-L1 preadipocytes, oxidizing extracellular redox potentials (E h ) increased intracellular and mitochondrial reactive oxygen species (ROS) production. 3T3-L1 adipocytes showed a greater response to extracellular E h , producing more intracellular ROS, than preadipocytes. 3T3-L1 adipocytes also produced more extracellular ROS and re-regulated the extracellular E h to a more oxidizing state than preadipocytes. During 3T3-L1 differentiation, cellular glutathione and mitochondrial thioredoxin-2 become oxidized, suggesting that adipogenesis may be enhanced under conditions promoting intracellular and mitochondrial compartment oxidation. Under various extracellular E h , 3T3-L1 adipogenesis, as determined by lipid accumulation and the expression of early genetic markers of adipogenesis, was sensitive to the extracellular redox environment, where it was enhanced under oxidizing conditions and lower under reducing conditions. Using a diet-induced obesity mouse model, plasma was collected before and after the 8 week diet regimens. Plasma GSH E h was unchanged as a consequence of weight gain but plasma cystiene (Cys) E h was significantly oxidized in overweight animals. Data presented here show that adipocytes/excessive adipose preferentially alter extracellular E h to a more oxidized state in vivo and in vitro and may promote further adipogenesis. & 2010 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved. Introduction Compartmentation of redox process has been proposed as a means by which redox signaling can be fine tuned to yield a specific response to specific stimuli (Hansen et al., 2006a). Recently, there have been multiple reports concerning the redox status of the extracellular compartment as a regulator of intracellular processes and cell function. An oxidized extracellular environment has been associated with many different diseases, including atherosclerosis and diabetes (Ashfaq et al., 2006; Samiec et al., 1998). In addition, changes in the extracellular redox states are correlated with alcohol consumption and cancer chemotherapies (Jonas et al., 2000; Moriarty et al., 2003). These studies support the hypothesis that the extracellular redox status is an important regulator of pathophysiologic processes and exposure to environmental insults. While the exact mechanisms of extracellular redox control are just beginning to be understood, there is some evidence that extracellular redox status affects intracellular signaling pathways. In in vitro models of cardiovascular disease, monocyte adhesion, an early event of atherosclerosis, was dramatically increased in bovine aortic endothelial cells that were maintained in oxidizing culture conditions (Go and Jones, 2005). Furthermore, the redox- sensitive transcription factor, NF-kB, was shown to be more active under oxidizing extracellular environments and resulted in an increase in the expression of NF-kB-dependent cell adhesion molecules (Go and Jones, 2005). These data suggest that the extracellular redox state may play an important regulatory role in plaque formation and heart disease. The intestinal cell line, Caco-2, was used to show that cellular proliferation was decreased in cells maintained in an oxidizing environment compared to a reducing one (Jonas et al., 2002). In human retinal pigment epithelial cells, cells in an oxidizing medium were more prone to undergo oxidant-induced, mito- chondria-mediated apoptosis than cells in a reducing medium (Jiang et al., 2005). These changes may be due to alterations in intracellular ROS generation, which occur when cells are incu- bated under oxidizing conditions (Go and Jones, 2005). In the plasma, cysteine (Cys)/cystine (CySS) and glutathione (GSH)/glutathione disulfide (GSSG) are present. However, unlike intracellular concentrations where GSH dominates, plasma GSH and GSSG concentrations are dramatically reduced. Intracellular GSH concentrations are estimated to be in the range 2–10 mM, while extracellular concentrations are estimated between ARTICLE IN PRESS Contents lists available at ScienceDirect journal homepage: www.elsevier.com/locate/diff Differentiation 0301-4681/$ - see front matter & 2010 International Society of Differentiation. Published by Elsevier Ltd. All rights reserved. Join the International Society for Differentiation (www.isdifferentiation.org) doi:10.1016/j.diff.2010.04.005 n Corresponding author. Tel.: + 404 727 3145; fax: + 404 712 9712. E-mail address: jhansen@emory.edu (J.M. Hansen). Differentiation 80 (2010) 31–39