Intact glucose uptake despite deteriorating signaling in adipocytes with high-fat feeding

Abstract

To capture immediate cellular changes during diet-induced expansion of adipocyte cell volume and number, we characterized mature adipocytes during a short-term high-fat diet (HFD) intervention. Male C57BL6/J mice were fed chow diet, and then switched to HFD for 2, 4, 6 or 14 days. Systemic glucose clearance was assessed by glucose tolerance test. Adipose tissue was dissected for RNA-seq and cell size distribution analysis using coulter counting. Insulin response in isolated adipocytes was monitored by glucose uptake assay and Western blotting, and confocal microscopy was used to assess autophagic activity. Switching to HFD was accompanied by an immediate adipocyte size expansion and onset of systemic insulin resistance already after two days, followed by recruitment of new adipocytes. Despite an initially increased non-stimulated and preserved insulin-stimulated glucose uptake, we observed a decreased phosphorylation of insulin receptor substrate-1 (IRS-1) and protein kinase B (PKB). After 14 days of HFD, both the insulin-stimulated phosphorylation of Akt substrate of 160 kDa (AS160) and glucose uptake was blunted. RNA-seq analysis of adipose tissue revealed transient changes in gene expression at day four, including highly significant upregulation of , previously demonstrated to be involved in autophagy. We confirmed increased autophagy, measured as an increased density of LC3-positive puncta and decreased p62 expression after 14 days of HFD. In conclusion, HFD rapidly induced systemic insulin resistance, whereas insulin-stimulated glucose uptake remained intact throughout 6 days of HFD feeding. We also identified autophagy as an early cellular process that potentially influences adipocyte function upon switching to HFD.

EHD2 regulates adipocyte function and is enriched at cell surface–associated lipid droplets in primary human adipocytes

“…Western blot analysis demonstrated that the protein levels of caveolin-1 and EHD2 had increased three- and twofold, respectively, after 14 d of high-fat diet (HFD) (Figure 1B). While the protein level of caveolin-1 rapidly increased after 2 d, the EHD2 level increased after 14 d, a time point that we previously (Hansson et al , 2018) demonstrated to coincide with considerable adipocyte differentiation. Next, by comparing the EHD2 levels in adipose tissue versus isolated adipocytes from chow- or HFD-fed mice, we found that the increase of EHD2 in adipose tissue following HFD was not due to an increase in the isolated adipocytes (Figure 1C).…”

“…First, we examined the relationship between caveolin-1 and EHD2 expression in adipose tissue using RNA sequence data previously obtained from adipose tissue during a short-term (14 d) overfeeding study in mice (Hansson et al , 2018). By analyzing that data set, we found a positive correlation between EHD2 and caveolin-1 mRNA expression (Figure 1A).…”

“…Impaired energy storage capacity in adipocytes is associated with increased levels of circulating FFA and with ectopic lipid accumulation in liver and muscle, which is a major risk factor for developing insulin resistance and type 2 diabetes (Schinner et al , 2005). We and others have examined the dynamics of adipose tissue expansion (McLaughlin et al , 2007; Li et al , 2016; Hansson et al , 2018), which involves increased adipocyte size (hypertrophy) as well as increased cell number (hyperplasia). The latter occurs through differentiation of preadipocytes (adipogenesis) (Salans et al , 1968), a process tightly regulated by various transcription factors, including peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer-binding proteins alpha and beta (C/EBPα and C/EBPβ).…”

“…Recently, we identified EH domain–containing 2 (EHD2) as one of the most highly up-regulated genes in adipose tissue during short-term overfeeding in mice (Hansson et al , 2018). EHD2 belongs to a family of well-conserved dynamin-related ATPase proteins (EHD1–4) that are implicated in several cellular processes, including membrane recycling, receptor internalization, and GLUT4 endocytosis (Guilherme et al , 2004; Naslavsky et al , 2004; Park et al , 2004).…”

Adipose Morphology: a Critical Factor in Regulation of Human Metabolic Diseases and Adipose Tissue Dysfunction

“…Apart from the intrinsic regulation, the architecture of adipocytes was also influenced by extracellular factors. Overnutrition induced by short-term overfeeding in mice rapidly causes onset of cellular insulin resistance in adipocytes [134], marked by impaired insulin signaling at the level of insulin receptor substrate (IRS)-1 [135,136] and Akt [137]. Besides, IQ motif containing GTPase activating protein 1 (IQGAP-1) is reduced.…”