ANGPTL3 facilitates lipolysis through insulin-resistance development in 3T3-L1 cells.

Abstract Detection of loss-of-function (LOF) mutations in ANGPTL3 gene in humans revealed the existence of a new lipid phenotype, called familial combined hypolipidemia (FHBL2) and characterized by markedly reduced levels of all lipoproteins. ANGPLT3 protein is known to be involved in the inhibitory regulation of extracellular lipases (LPL and HL), so that its absence increases the lipolytic degradation of TG-rich lipoproteins (in particular VLDL) and HDL. Accordingly, the FHBL2 status is also characterized by reduced levels of circulating free fatty acids (FFA), revealing that ANGPTL3 also plays a role in regulating the metabolism of FFA. Our previous data indicate ANGPTL3 facilitates β-adrenergic-dependent lipolysis in adipocytes and ANGPTL3 treated 3T3-L1 cells showed high levels of ERK1/2 as well as pPKA/PKA and pHSL/HSL, downstream the cAMP-dependent signaling pathway. In order to understand mechanisms through which ANGPTL3 regulates lipolysis in adipose tissue, our purpose is to demonstrate ANGPTL3 can be involved in insulin-resistance development. Methods: Therefore, the main aim of present study is to investigate changes in cAMP-dependent pathways induced by the addition of ANGPTL3 to 3T3-L1 cell line. Changes in release of FFA and in expression of genes (AMPK, P38, AKT and HSL) involved in the canonical and non-canonical activation of lipolytic pathways will be investigated by a combination of biochemical measures, Western blot and RT-qPCR analyses. Results: Compared to β-agonist (isoproterenol) stimulated cells, adipocytes treated with isoproterenol (Iso), in presence of ANGPTL3, showed highest levels of intracellular cAMP (Iso 4,25 nM; ANGPTL3 pre-treatment 7,89 nM; p<0,001) and increased activation of kinases. Conclusion: Recently, it was shown that AMPK-P38-AKT pathway activation increases insulin-resistance in adipocytes, which leads to a greater availability of cAMP within the cell. According to literature, our findings indicate ANGPTL3 facilitates FFA release in adipose tissue counteracting the insulin inhibitory action against β-adrenergic-dependent lipolysis.