The Multimerization And Secretion Of Adiponectin Are Regulated By TNF-α

Adiponectin is the most abundant adipokine in the circulation of humans and other mammals. In addition to regulation of energy homeostasis, adiponectin has a protective role against insulin resistance and is antiatherogenic, anti-inflammatory. Treatment of mice with adiponectin causes glucose depression and ameliorates insulin resistance.Conversely, depletion of adiponectin causes insulin resistance and glucose intolerance. In obese individuals, serum adiponectin levels are significantly reduced.Adiponectin exists in the circulation as three complexes: trimer(LMW), hexamer(MMW), and the high-molecular-weight(HMW) complexes of 18–36 monomers.Recently groups have proposed that the ratio of HMW/total adiponectin rather than absolute amount of adiponectin determines insulin sensitivity in humans and rodents. The multimerization of adiponectin depends on formation of intermolecular disulfide bonds.Development of disulfide bonds occurs in the endoplasmic reticulum(ER), and early reports revealed that ER-resident chaperone proteins, ERO1-Lα, Dsb A-L and ERp44 are involved in the assembly and secretion of adiponectin oligomers, especially for secretion of HMW adiponectin.As an endocrine organ, adipose tissue secrets a wide range of cytokines, many of which are increased in obese subjects, including TNF-α, leptin, IL-6, and visfatin.Moreover, in obese individuals, macrophage infiltration of adipose tissue promotes a proinflammatory microenvironment by secreting inflammatory cytokines. Whether these cytokines regulate adipocyte secretion of adiponectin in an autocrine or paracrine manner remains unknown. TNF-α, a cytokine secreted by mature adipocytes and macrophages,plays an important role in regulating glucose and lipid metabolism. It is also one of the major cytokine responsible for inflammation-induced insulin resistance.Clinical studies revealed an inverse correlation between circulating levels of TNF-α and adiponectin in obese and diabetic patients, but whether TNF-α regulates the circulating adiponectin by directly targeting adipocytes needs further study. Currently, westudied the effect of TNF-α on adiponectin multimerization and secretion both in vivo and vitro, and further explored the mechanisms involved in this regulation.1. We established mouse model of insulin resistance with the method of high-fat diet or intraperitoneal injection of TNF-α respectively, and we found both high-fat diet mouse group and TNF-α treated mouse group present significant insulin resistance. More importantly, the two groups exhibited comparable decrease of adiponectin multierization and secretion in circulation.2. We then treated differentiated 3T3-L1 cell with TNF-α and found TNF-αdecreased the adiponectin multierization and secretion in culture media of 3T3-L1.Furthermore, TNF-α treatment depressed the m RNA and protein levels of ERO1-Lα and Dsb A-L.3. Treatment with PPARγ agonists and inhibitors, and PPARγ knockdown confirmed that PPARγ is involved in the transcriptional regulation of ERO1-Lα and Dsb A-L. Based on the cloning of ERO1-Lα and Dsb A-L promoter sequences, bioinformatic prediction and promoter luciferase activity assay further confirmed that PPARγ is a transcription factor for ERO1-Lα and Dsb A-L. Chromatin immunoprecipitation was used to confirm that PPARγ binds to the-1685/-1533 PPRE element of ERO1-Lα promoter, and the-939/-815 PPRE element of Dsb A-L promoter, and regulate the transcription of ERO1-Lαand Dsb A-L expression.4. TNF-α treatment leads to decrease of ERp44 transcription, overexpression of PPARγ present no significantly rescuing effect on this decrease. Co-IP was used to confirm that TNF-α treatment enhanced the protein interaction between ERp44 and adiponectin.In summary, TNF-α decrease the expression of PPARγ that leads to the downregulation of ERO1-Lα and Dsb A-L transcription, and TNF-α increase the retention of adiponectin via downregulating ERp44 transcription, which collectively result in the decrease of adiponectin multimerization and secretion in the circulation of obese subjects.