Hyperleptinemia Is Required for the Development of Leptin Resistance Zachary A. Knight, K. Schot Hannan, Matthew L. Greenberg, Jeffrey M. Friedman* Howard Hughes Medical Institute, Laboratory of Molecular Genetics, The Rockefeller University, New York, New York, United States of America Abstract Leptin regulates body weight by signaling to the brain the availability of energy stored as fat. This negative feedback loop becomes disrupted in most obese individuals, resulting in a state known as leptin resistance. The physiological causes of leptin resistance remain poorly understood. Here we test the hypothesis that hyperleptinemia is required for the development of leptin resistance in diet-induced obese mice. We show that mice whose plasma leptin has been clamped to lean levels develop obesity in response to a high-fat diet, and the magnitude of this obesity is indistinguishable from wild- type controls. Yet these obese animals with constant low levels of plasma leptin remain highly sensitive to exogenous leptin even after long-term exposure to a high fat diet. This shows that dietary fats alone are insufficient to block the response to leptin. The data also suggest that hyperleptinemia itself can contribute to leptin resistance by downregulating cellular response to leptin as has been shown for other hormones. Citation: Knight ZA, Hannan KS, Greenberg ML, Friedman JM (2010) Hyperleptinemia Is Required for the Development of Leptin Resistance. PLoS ONE 5(6): e11376. doi:10.1371/journal.pone.0011376 Editor: Krisztian Stadler, Louisiana State University, United States of America Received March 30, 2010; Accepted June 7, 2010; Published June 29, 2010 Copyright: 2010 Knight et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by NIH grants DK083531 (Z.A.K.) and DK041096 (J.M.F.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: friedj@rockefeller.edu Introduction Body weight in mammals is controlled by a physiological system that balances energy intake and expenditure over the long term[1]. The core component of this system that signals the availability of body fat is the hormone leptin[2]. Leptin is secreted by adipocytes in proportion to their size and number, such that the concentration of leptin in the blood is proportional to the total amount of adipose tissue[3,4]. Binding of leptin to its target neurons, which are expressed in the hypothalamus, brain stem and other brain regions, inhibits feeding and stimulates energy expenditure. Leptin thus functions as the afferent signal in a negative feedback loop that maintains a stable level of body fat reserves. Leptin deficient mice (ob/ob) and humans are obese and hyperphagic[5], and, in these individuals, leptin replacement therapy induces dramatic weight loss[6,7,8,9]. However most obesity is associated with elevated plasma leptin levels[3,4], implying resistance to leptin’s weight reducing effects[10]. The contribution of leptin resistance to obesity has also been established by the demonstration that hyperleptinemic animals and humans have a blunted response to exogenous leptin. Despite the importance of delineating the causes of leptin resistance, the cellular and molecular mechanisms responsible remain poorly understood. The diet-induced obese mouse is a well characterized system for studying the development leptin resistance and pathogenesis of obesity. In this model, C57Bl/6J mice fed a high-fat diet (45% to 60% calories from fat) become progressively obese and hyperlep- tinemic over a span of 4 to 6 months. As these animals become obese, they lose the ability to reduce their food intake and body weight in response to leptin treatment. In the early stages of obesity, mice develop resistance to leptin delivered peripherally, but not centrally; this has been attributed to downregulation or saturation of the transport system that transports leptin across the blood-brain barrier[11,12]. After long-term exposure to high-fat diet (.20 weeks), mice become resistant to leptin even when it is directly infused into the brain via the cerebral ventri- cle[10,12,13,14]. In these animals, the first-order, leptin-respon- sive neurons have apparently lost the ability to activate the signaling pathways downstream of the leptin receptor. How does exposure to a high-fat diet impair the leptin sensitivity of these neurons? Two models have been proposed. The first is that leptin resistance is caused by elevated plasma leptin levels, which result in chronic overstimulation of the leptin receptor and activation of negative feedback pathways that block further leptin signaling. This model is supported by the fact that leptin stimulates the expression of SOCS-3, a protein that directly inhibits leptin signaling[15,16,17], and that ablation of SOCS-3 in neurons enhances leptin sensitivity and protects against diet-induced obesity[18,19,20]. Moreover, targeted expression of a constitu- tively active form of STAT3, which is a key mediator of leptin signaling, is sufficient to induce leptin resistance in the hypothal- amus[21]. This mechanism is analogous to the decrease in insulin receptor signaling that is associated with chronic insulin treatment and believed to result from the activation of negative feedback pathways such as serine phosphorylation of IRS-1[22]. An alternative explanation for the development of leptin resistance is that dietary fats themselves, rather than hyperlepti- nemia, are responsible. Fats could either directly block leptin signaling or activate cellular processes, such as endoplasmic reticulum (ER) stress and inflammation, that impair leptin PLoS ONE | www.plosone.org 1 June 2010 | Volume 5 | Issue 6 | e11376