RESUMO
Salusinß and adiponectin receptor 1 (adipoR1) serve important roles in the development of certain cardiovascular diseases and lipid metabolism. However, to the best of our knowledge, the relationship between salusinß and adipoR1, and their underlying mechanisms of action, currently remain unclear. In the present study, lentiviral vectors designed to overexpress salusinß or knock down salusinß expression were used in 293T and HepG2 cells. Semiquantitative PCR was performed to investigate the relationship between salusinß and adipoR1 mRNA expression in 293T cells. Western blotting was used to assess the protein expression levels of adipoR1, adenosine monophosphateactivated protein kinase (AMPK), acetylCoA carboxylase (ACC) and carnitine palmitoyl transferase 1A (CPT1A) in transfected HepG2 cells. Simultaneously, HepG2 cells were treated with an adipoR1 inhibitor (thapsigargin) or agonist (AdipoRon) and the resultant changes in the expression levels of the aforementioned proteins were observed. Oil Red O staining and measurements of cellular triglyceride levels were performed to assess the extent of lipid accumulation in HepG2 cells. The results demonstrated that salusinß overexpression downregulated adipoR1 expression and inhibited the phosphorylation of AMPK and ACC, which led to decreased CPT1A protein expression. By contrast, salusinß knockdown increased adipoR1 expression and promoted the phosphorylation of AMPK and ACC, which conversely enhanced CPT1A protein expression. Treatment with adipoR1 agonist, AdipoRon, reversed the effects of salusinß overexpression. In addition, salusinß overexpression enhanced intracellular lipid accumulation in HepG2 cells induced by free fatty acid treatment. These findings highlighted the potential regulatory role of salusinß in adipoR1mediated signaling pathways. To conclude, the present study provided insights into the regulation of fatty acid metabolism by the liver. In particular, salusinß may serve as a potential target for the therapeutic intervention of metabolic disorders of lipids.
Assuntos
Proteínas Quinases Ativadas por AMP , Metabolismo dos Lipídeos , Receptores de Adiponectina , Humanos , Proteínas Quinases Ativadas por AMP/metabolismo , Ácidos Graxos não Esterificados/metabolismo , Células Hep G2 , Fígado/metabolismo , Receptores de Adiponectina/genética , Receptores de Adiponectina/metabolismoRESUMO
Salusinα and adiponectin, are vasoactive peptides with numerous similar biological effects related to lipid metabolism. Adiponectin has been shown to reduce fatty acid oxidation and to inhibit lipid synthesis of liver cells through its receptor, adiponectin receptor 2 (AdipoR2), but whether salusinα is able to interact with AdipoR2, was not previously reported. To investigate this, in vitro experiments were carried out. The overexpression and interference recombinant plasmids were constructed with salusinα. The lentiviral expression systems of salusinα overexpression and interference were respectively synthesized in 293T cells, and 293T cells were infected with the lentivirus. Finally, the association between salusinα and AdipoR2 was analyzed by semiquantitative PCR. Subsequently, HepG2 cells were also infected with these viruses. The expression levels of AdipoR2, peroxisome proliferatoractivated receptorα (PPARα), apolipoprotein A5 (ApoA5) and sterol regulatory elementbinding transcription factor 1 (SREBP1c) were detected by western blotting, and AdipoR2 inhibitor (thapsigargin) and agonist [4phenyl butyric acid (PBA)] were used to observe the resultant changes in the aforementioned molecules. The results obtained revealed that the overexpression of salusinα increased the level of AdipoR2 in 293T and HepG2 cells, led to an upregulation of the levels of PPARα and ApoA5, and inhibited the expression of SREBP1c, whereas the salusinα interference lentivirus exerted the opposite effects. Notably, thapsigargin inhibited the expression of AdipoR2, PPARα and ApoA5 in HepG2 cells of pHAGESalusinα group, and caused an increase in the level of SREBP1c, whereas the opposite effects were observed in pLKO.1shSalusinα#1 group upon treatment with PBA. Taken together, these data demonstrated that overexpression of salusinα upregulated AdipoR2, which in turn activated the PPARα/ApoA5/SREBP1c signaling pathway to inhibit lipid synthesis in HepG2 cells, thereby providing theoretical data on which to base the clinical application of salusinα as a novel peptide for molecular intervention in fatty liver disease.