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Proteomics analyses of subcutaneous adipocytes reveal novel abnormalities in human insulin resistance.

ABSTRACT: To provide a more global view of adipocyte changes in human insulin resistance by proteomics analyses.Baseline biopsies of abdominal subcutaneous adipose tissue were obtained from 23 subjects without diabetes. Euglycemic clamps were used to divide subjects into an insulin-resistant group (IR, N = 10) and an insulin-sensitive (IS, N = 13) group, which were of similar age and gender but unequal adiposity (greater in IR). Proteins of isolated adipocytes were quantified by mass spectrometry using normalized spectral abundance factors.Of 1,245 proteins assigned, 30 were detected in at least 12 of the 23 subjects that differed significantly in abundance ?1.5-fold between IR and IS. IR displayed a pattern of increased cytoskeletal proteins and decreased mitochondrial proteins and FABP4 and FABP5. In subgroup analyses of adiposity-matched subjects, several of these changes were less pronounced in IR, but the abundance of proteins related to lipid metabolism and the unfolded/misfolded protein response were significantly and unfavorably altered.These results confirm lower abundance of mitochondrial proteins and suggest increased cytoskeletal proteins and decreased FABP4 and FABP5 in subcutaneous adipocytes of typical IR individuals. Changes in proteins related to lipid metabolism and the unfolded/misfolded protein may discriminate IR and IS individuals of equal adiposity.

SUBMITTER: Xie X

PROVIDER: S-EPMC4926648 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Proteomics analyses of subcutaneous adipocytes reveal novel abnormalities in human insulin resistance.

Xie Xitao X Yi Zhengping Z Sinha Sandeep S Madan Meenu M Bowen Benjamin P BP Langlais Paul P Ma Danjun D Mandarino Lawrence L Meyer Christian C

Obesity (Silver Spring, Md.) 20160701 7

<h4>Objective</h4>To provide a more global view of adipocyte changes in human insulin resistance by proteomics analyses.<h4>Methods</h4>Baseline biopsies of abdominal subcutaneous adipose tissue were obtained from 23 subjects without diabetes. Euglycemic clamps were used to divide subjects into an insulin-resistant group (IR, N = 10) and an insulin-sensitive (IS, N = 13) group, which were of similar age and gender but unequal adiposity (greater in IR). Proteins of isolated adipocytes were quanti ...[more]

PMID: 27345962

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Project description:: Insulin resistance is present in the majority of patients with non-insulin-dependent diabetes mellitus (NIDDM) and obesity. In this study, we aimed to investigate the key genes and potential molecular mechanism in insulin resistance. Expression profiles of the genes were extracted from the Gene Expression Omnibus (GEO) database. Pathway and Gene Ontology (GO) enrichment analyses were conducted at Enrichr. The protein⁻protein interaction (PPI) network was settled and analyzed using the Search Tool for the Retrieval of Interacting Genes (STRING) database constructed by Cytoscape software. Modules were extracted and identified by the PEWCC1 plugin. The microRNAs (miRNAs) and transcription factors (TFs) which control the expression of differentially expressed genes (DEGs) were analyzed using the NetworkAnalyst algorithm. A database (GSE73108) was downloaded from the GEO databases. Our results identified 873 DEGs (435 up-regulated and 438 down-regulated) genetically associated with insulin resistance. The pathways which were enriched were pathways in complement and coagulation cascades and complement activation for up-regulated DEGs, while biosynthesis of amino acids and the Notch signaling pathway were among the down-regulated DEGs. Showing GO enrichment were cardiac muscle cell⁻cardiac muscle cell adhesion and microvillus membrane for up-regulated DEGs and negative regulation of osteoblast differentiation and dendrites for down-regulated DEGs. Subsequently, myosin VB (MYO5B), discs, large homolog 2(DLG2), axin 2 (AXIN2), protein tyrosine kinase 7 (PTK7), Notch homolog 1 (NOTCH1), androgen receptor (AR), cyclin D1 (CCND1) and Rho family GTPase 3 (RND3) were diagnosed as the top hub genes in the up- and down-regulated PPI network and modules. In addition, GATA binding protein 6 (GATA6) , ectonucleotide pyrophosphatase/phosphodiesterase 5 (ENPP5), cyclin D1 (CCND1) and tubulin, beta 2A (TUBB2A) were diagnosed as the top hub genes in the up- and down-regulated target gene⁻miRNA network, while tubulin, beta 2A (TUBB2A), olfactomedin-like 1 (OLFML1), prostate adrogen-regulated mucin-like protein 1 (PARM1) and aldehyde dehydrogenase 4 family, member A1 (ALDH4A1)were diagnosed as the top hub genes in the up- and down-regulated target gene⁻TF network. The current study based on the GEO database provides a novel understanding regarding the mechanism of insulin resistance and may provide novel therapeutic targets.

Project description:Adipose tissue is both an energy storage depot and an endocrine organ. The impaired regulation of the secreted proteins of adipose tissue, known as adipocytokines, observed during obesity contributes to the onset of whole-body insulin resistance and the pathobiology of type 2 diabetes mellitus (T2DM). In addition, the global elevation of the intracellular glycosylation of proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) via either genetic or pharmacological methods is sufficient to induce insulin resistance in both cultured cells and animal models. The elevation of global O-GlcNAc levels is associated with the altered expression of many adipocytokines. We have previously characterized the rodent adipocyte secretome during insulin sensitive and insulin resistant conditions. Here, we characterize and quantify the secretome and glycome of primary human adipocytes during insulin responsive and insulin resistant conditions generated by the classical method of hyperglycemia and hyperinsulinemia or by the pharmacological manipulation of O-GlcNAc levels. Using a proteomic approach, we identify 190 secreted proteins and report a total of 20 up-regulated and 6 down-regulated proteins that are detected in both insulin resistant conditions. Moreover, we apply glycomic techniques to examine (1) the sites of N-glycosylation on secreted proteins, (2) the structures of complex N- and O-glycans, and (3) the relative abundance of complex N- and O-glycans structures in insulin responsive and insulin resistant conditions. We identify 91 N-glycosylation sites derived from 51 secreted proteins, as well as 155 and 29 released N- and O-glycans respectively. We go on to quantify many of the N- and O-glycan structures between insulin responsive and insulin resistance conditions demonstrating no significant changes in complex glycosylation in the time frame for the induction of insulin resistance. Thus, our data support that the O-GlcNAc modification is involved in the regulation of adipocytokine secretion upon the induction of insulin resistance in human adipocytes.

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Proteomics analyses of subcutaneous adipocytes reveal novel abnormalities in human insulin resistance.

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Proteomics analyses of subcutaneous adipocytes reveal novel abnormalities in human insulin resistance.

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