Numerous lines of evidence have suggested that energy balance in animals and humans is tightly controlled. With the identification of leptin and its receptors by our laboratory, two of the molecular components of a system that maintains constant weight have been identified. Leptin is a hormone secreted by the adipose (fat) tissue in proportion to its mass that in turn modulates food intake relative to energy expenditure. Increased fat mass increases leptin levels, which in turn reduces body weight; decreased fat mass leads to a decrease in leptin levels and an increase in body weight. By this mechanism, weight is maintained within a relatively narrow range. Defects in the leptin gene are associated with severe obesity in animals and in humans. Leptin acts on sets of neurons in brain centers that control energy balance. Leptin also plays a general role in regulating many of the physiologic responses that are observed with changes in nutritional state, with clear effects on female reproduction, immune function, and the function of many other hormones, including insulin.
Our current research focuses on a series of questions pertinent to the regulation of body weight: How does the fat cell regulate how much leptin is made—i.e., how does the fat cell know how fat it is? How does a single molecule (leptin) change feeding, a complex behavior? How do brain pathways that are modulated by leptin in turn regulate peripheral metabolism and insulin action? Does variation in the genes that compose the physiologic circuit of which leptin is a component explain differences in body weight? Does leptin treatment change the biologic response to weight loss in humans?
The Neural Circuitry Controlling Feeding
The recent identification of the hypothalamic cells that express the leptin receptor is enabling us to delineate the precise neuronal effects of leptin and the mechanisms by which this single molecule can alter a complex behavior. Toward that end we have developed a novel system that employs an engineered virus to trace the neural circuit that is modulated by leptin. Results using these mice have suggested that a number of brain regions, including those known to regulate emotional behavior and higher brain functions, modulate leptin signaling in the hypothalamus. We have also shown that the neural circuits that respond to leptin are extremely dynamic and that leptin has rapid and dramatic effects on the number of synapses (connections between nerve cells) found on key neural pathways that regulate feeding. Another method known as laser photostimulation was used to map with great precision the inputs to leptin-responsive neurons. Our current studies are aimed at understanding how multiple relevant inputs are integrated to modulate the amount of food an animal eats. These studies may have general implications for our understanding of the regulation of a complex behavior.
Regulation of Leptin Production
We have initiated a new line of research aimed at elucidating the molecular mechanisms responsible for the regulation of leptin gene expression associated with changes in weight (These studies are funded by a grant from the National Institutes of Health). The amount of leptin that is expressed from fat is strongly regulated, with a 100-fold or more level of expression from ob/ob adipose tissue than from the adipose tissue of a lean or fasted animal. This suggests that the fat cell knows how much fat it has and adjusts leptin expression accordingly. The underlying mechanism responsible for this regulation is unknown. To address this question, we are using transgenic mice to identify DNA regulatory elements that change expression of a receptor gene controlled by the leptin gene in parallel with changes in adipose tissue mass. We have modified a series of leptin bacterial artificial chromosome (BAC) clones so that the leptin DNA regulatory elements direct the expression of luciferase. These modified BACs will be introduced into transgenic mice so that we can identify those BACs that replicate changes in the endogenous level of leptin expression that develop when weight is gained or lost. We are also analyzing the secretion of leptin in transgenic mice that express leptin-GFP (green fluorescent protein) fusion proteins. The relevant constructs have been made, and the generation of transgenic mice is under way.
The Molecular Basis of Leptin's Metabolic Effects
Leptin has potent metabolic effects to improve insulin action and reduce the lipid content of peripheral tissues. In previous studies we showed that leptin's effects on fat deposition resulted from the down-regulation of stearoyl-CoA desaturase (SCD-1), an enzyme that plays an important role in fat metabolism. Studies are now under way to establish the tissues where SCD-1 plays a role in regulating metabolism. In studies with Markus Stoffel's group (Rockefeller University), we showed that leptin's anti-diabetic effects are independent of SCD-1 and in part a result of activation of the FoxA2 transcription factor. Other studies are aimed at increasing understanding of the mechanism responsible for leptin's antidiabetic effects. (This work was supported by the National Institutes of Health.)
Leptin in Human Physiology
Diet-induced weight loss in humans results in a decrease in leptin concentration. This may explain the high failure rate of dieting, as low leptin is a potent stimulus to increase appetite and weight in animals and humans. Clinical studies at the Rockefeller Hospital are under way to explore the possibility that administration of leptin to patients who are dieting can alter the biologic response to weight loss and mitigate some of the unwanted side effects of a low-calorie diet.
Genetic Studies in Humans
Advances in genetics make it possible to use positional cloning to identify other genes that contribute to human disease. To implement this approach, we have been conducting genetic studies in collaboration with the Department of Health on the Pacific island of Kosrae in Micronesia and with the laboratories of Marcus Stoffel and Jan Breslow (Rockefeller University). The citizens of Kosrae have a high incidence of obesity, the basis of which is not understood. We have conducted a complete medical evaluation of the entire adult population of Kosrae over 20 years of age (some 2,500 individuals), including measurements of height, weight, blood pressure, glucose, cholesterol, triglycerides, serum insulin, and leptin. Recently we have used high-density DNA microarrays from Affymetrix to genotype the entire population for ~500,000 different genetic markers known as single nucleotide polymorphisms (SNPs). These results will allow us to assess which of these DNA variants correlate best with the different clinical parameters that were ascertained.