Molecular and functional mechanisms involved in the central effects of FGF19 on hypothalamic dysregulation

Abstract

Obesity is defined as an excessive and abnormal accumulation of fat and is often associated with hypothalamic dysregulation. The consumption of high-fat diets has been considered the main factor responsible for inducing hypothalamic inflammation, commonly found in obese individuals. This inflammation has been associated with chronic hypothalamic resistance to the signs of anorexigenic and pro-thermogenic hormones leptin and insulin, which are unable to promote efficient reduction in food intake and increase in energy expenditure. Therefore, several strategies have been used in the treatment of Obesity and the inflammatory process associated with it, among which we can highlight the Fibroblast Growth Factor (FGF) 19. FGF19 and its orthologue in rodents, the FGF15, are hormones produced by the intestine, and secreted into the circulation in response to the release of bile acids in postprandial situations. In addition to its well-established role in inhibiting hepatic biosynthesis of bile acids, FGF15/19 also acts on the hypothalamus, reducing food intake and increasing energy expenditure in obese mice. However, the molecular mechanisms involved in these effects are not fully understood yet, as well as the central effects of FGF19 in metabolically active peripheral tissues. In this work, using C57BL/6 mice that will receive a high-fat diet for 8 weeks, for Obesity induction, and treated centrally with 3 µg of FGF19 for 5 days, our aim is to evaluate the hypothalamic effects of FGF19 on the glucose and energy metabolism of obese mice, as well as the molecular mechanisms involved in their responses. In the hypothalamus, gene expression of neuropeptides and inflammatory markers will be evaluated using RT-PCR. By means of western blot, we will analyze the hypothalamic signaling of FGF19, through the protein content of ERK1/2 phosphorylation. The protein content of the phosphorylation of AKT and IRS-1, as well as of JAK2 and STAT3, will also be quantified, to assess hypothalamic sensitivity to insulin and leptin, respectively. In these animals, we will also assess glucose metabolism, through glucose and insulin tolerance tests, insulin secretion stimulated by glucose in isolated islets and calculation of insulin clearance. Energy metabolism will be assessed by indirect calorimetry, where energy expenditure, respiratory quotient and locomotor activity will be analyzed, in addition to food intake. In the periphery, the gene expression of thermogenic markers, in Brown Adipose Tissue (BAT) and in subcutaneous White Adipose Tissue (WAT), and of lipolysis and lipogenesis markers in perigonadal WAT will be quantified. In addition, the temperature of the BAT will be determined through an infrared detection camera, and the amount of triglycerides will be measured in the liver. Finally, we will investigate the metabolic repercussions of the hypothalamic-specific knockdown of FGFR1 and FGFR3 receptors, through lentiviral transfection. The results of this project will be important to understand the molecular mechanisms involved in the central FGF19 responses, which may lead to the development of new strategies for the treatment of diseases where hypothalamic and metabolic dysregulation is present, such as Diabetes and Obesity. (AU)