Effects of high fructose diet on insulin signaling in Skeletal muscle tissue of Protein Kinase C Epsilon knockout (PKCεKO) heterozygous mice

Background: Protein kinase C epsilon (PKCε) plays an important role in glucose metabolism by regulating insulin signaling pathways in hepatic and skeletal muscle tissues. Insulin resistance as a consequence of impaired insulin signaling is one of the key features of metabolic diseases such as type 2 diabetes.Evidence suggests that impaired insulin signaling occurs within the Akt pathway, which regulates glucose uptake, glycogen synthesis, and lipid metabolism in tissue specific manner. Insulin typically increases phosphorylated Akt (p-Akt) levels to promote glucose uptake; however, conditions such as diet-induced obesity, chronic inflammation, glucocorticoid excess, nutrient imbalances, and chronic hyperinsulinemia disrupt this effect. Mechanisms including increased protein phosphatase activity, inflammatory cytokines like TNF-α, elevated free fatty acids, and genetic mutations impair p-Akt activation which lead to elevated hepatic glucose production, impaired glycogen synthesis, and lipid accumulation. However, the precise mechanism is still poorly understood. Methods: This study investigates the effects of PKCε heterozygosity (Liv-PKCε-Het) in a high-fructose diet (HFrD) model, focusing on Akt and Foxo1 phosphorylation as markers of insulin signaling. We also aim to investigate the molecular mechanism of high-fructose diet (HFrD) induced insulin resistance mice liver protein kinase C epsilon knockout (Liv-PKCε) mice. The heterozygous LivPKCε (PKCε^fl/Δ) mice were fed either a control diet (Cntrl; Research Diets Inc., D12450J) or a high fructose diet (HFrD; Research Diets Inc., D02022704) for one month. Following this period, a subset of mice received an injection of 0.5 U/kg of insulin prior to euthanasia, after which skeletal muscle tissues were harvested. Protein expression in the skeletal muscle tissue was analyzed via western blot, with protein concentrations using BCA assays. Insulin signaling was evaluated by measuring p-AKT (Ser473), using a specific antibody (Santa Cruz Biotechnology, sc-514032). Insulin tolerance tests (ITT) and glucose tolerance tests (GTT) were performed. Results: Western blot analysis reveals that under basal HFrD conditions, phosphorylated Akt (p-Akt, S473) levels are elevated; however, insulin stimulation paradoxically reduces p-Akt levels, indicative of impaired hepatic insulin signaling. Similarly, phosphorylated Foxo1 (p-Foxo1, S256) levels decrease significantly following insulin exposure. This result suggest a disruptions in insulin-mediated regulation of gluconeogenic transcription factors.Using tissue-specific knockout models, we explored PKCε’s influence on signaling disruptions in skeletal muscle under various dietary conditions, including a low-fat control diet.PKCε deficiency enhances insulin sensitivity, evidenced by increased p-Akt levels and improved glucose uptake in skeletal muscle under low-fat diet conditions. Conclusion: Our results suggest that HFrD-induced metabolic stress compromises Akt activation and downstream signaling in the Liv-PKCε-Het model, contributing to hepatic and skeletal muscle insulin resistance. Understanding these conditions PKCε is highlighted as a potential therapeutic target for addressing insulin resistance and restoring Akt pathway function across metabolic tissues. We would like to thank the National Science Foundation for award NSF1931045 and the National Institutes of Health for award 1R01DK126892-01A1. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.