SUN-589 - The Effect of High-Fat Diet on Insulin Signaling in White Adipose Tissue of Protein Kinase C Heterozygous Male Mice

Background:
Obesity-associated insulin resistance is a central driver of metabolic diseases such as type 2 diabetes. White adipose tissue (WAT) plays a critical role in systemic insulin sensitivity and is a primary site of lipid storage and insulin action. Lipid-induced activation of protein kinase C epsilon (PKCε) has been implicated in impaired insulin signaling in metabolic tissues; however, its contribution to adipose tissue insulin responsiveness remains incompletely defined. Because WAT is highly susceptible to lipid overload and shares canonical insulin signaling pathways with other metabolic tissues, PKCε represents a biologically plausible regulator of adipose insulin signaling under conditions of dietary fat excess.

Methods:
Male PKCε heterozygous (PKCε⁺/–) mice and littermate controls were randomly assigned to a control diet (Research Diets Inc., D12450J) or a high-fat diet (HFD; Research Diets Inc., D12492) for 1 month (n = 4–6 per condition). Following a 6–7 hour fast, mice received 0.5 U/kg insulin or saline, and WAT was harvested 10 minutes post-injection. Insulin and saline injections, tissue processing, and densitometric analyses were performed in a blinded manner. Insulin signaling was assessed by Western blot analysis of total insulin receptor (IR), total AKT, phosphorylated AKT (p-AKT), and total FOXO1 protein expression, with actin used as a loading control.

Results:
In control diet–fed mice, insulin stimulation robustly increased p-AKT abundance in WAT (p < 0.05), with modest, non-significant effects on total IR and total FOXO1 protein levels. In contrast, HFD feeding significantly reduced total IR and total AKT protein abundance (p < 0.05) and abolished insulin-stimulated increases in p-AKT compared with control diet–fed mice. Although PKCε heterozygous mice exhibited preserved basal insulin signaling under control diet conditions, HFD feeding eliminated insulin-induced increases in total AKT and total FOXO1 expression. Notably, reductions in total IR and total AKT following HFD exposure were not rescued by partial PKCε deletion.

Conclusion:
Short-term high-fat diet feeding profoundly impairs insulin signaling protein abundance in white adipose tissue. Partial loss of PKCε is insufficient to rescue HFD-induced reductions in total IR and total AKT expression in WAT. Collectively, these findings suggest that dietary fat–induced adipose insulin resistance occurs upstream of, or independently from, PKCε dosage and underscore the complexity of tissue-specific mechanisms underlying lipid-induced insulin resistance.

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