Browsing by Author "Habegger, Kirk M."

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    An early, reversible cholesterolgenic etiology of diet-induced insulin resistance
    (Elsevier, 2023) Covert, Jacob D.; Grice, Brian A.; Thornburg, Matthew G.; Kaur, Manpreet; Ryan, Andrew P.; Tackett, Lixuan; Bhamidipati, Theja; Stull, Natalie D.; Kim, Teayoun; Habegger, Kirk M.; McClain, Donald A.; Brozinick, Joseph T.; Elmendorf, Jeffrey S.; Anatomy, Cell Biology and Physiology, School of Medicine
    Objective: A buildup of skeletal muscle plasma membrane (PM) cholesterol content in mice occurs within 1 week of a Western-style high-fat diet and causes insulin resistance. The mechanism driving this cholesterol accumulation and insulin resistance is not known. Promising cell data implicate that the hexosamine biosynthesis pathway (HBP) triggers a cholesterolgenic response via increasing the transcriptional activity of Sp1. In this study we aimed to determine whether increased HBP/Sp1 activity represented a preventable cause of insulin resistance. Methods: C57BL/6NJ mice were fed either a low-fat (LF, 10% kcal) or high-fat (HF, 45% kcal) diet for 1 week. During this 1-week diet the mice were treated daily with either saline or mithramycin-A (MTM), a specific Sp1/DNA-binding inhibitor. A series of metabolic and tissue analyses were then performed on these mice, as well as on mice with targeted skeletal muscle overexpression of the rate-limiting HBP enzyme glutamine-fructose-6-phosphate-amidotransferase (GFAT) that were maintained on a regular chow diet. Results: Saline-treated mice fed this HF diet for 1 week did not have an increase in adiposity, lean mass, or body mass while displaying early insulin resistance. Consistent with an HBP/Sp1 cholesterolgenic response, Sp1 displayed increased O-GlcNAcylation and binding to the HMGCR promoter that increased HMGCR expression in skeletal muscle from saline-treated HF-fed mice. Skeletal muscle from these saline-treated HF-fed mice also showed a resultant elevation of PM cholesterol with an accompanying loss of cortical filamentous actin (F-actin) that is essential for insulin-stimulated glucose transport. Treating these mice daily with MTM during the 1-week HF diet fully prevented the diet-induced Sp1 cholesterolgenic response, loss of cortical F-actin, and development of insulin resistance. Similarly, increases in HMGCR expression and cholesterol were measured in muscle from GFAT transgenic mice compared to age- and weight-match wildtype littermate control mice. In the GFAT Tg mice we found that these increases were alleviated by MTM. Conclusions: These data identify increased HBP/Sp1 activity as an early mechanism of diet-induced insulin resistance. Therapies targeting this mechanism may decelerate T2D development.
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    Chromium Enhances Insulin Responsiveness via AMPK
    (Elsevier, 2014-05) Hoffman, Nolan J.; Penque, Brent A.; Habegger, Kirk M.; Sealls, Whitney; Tackett, Lixuan; Elmendorf, Jeffrey S.; Department of Cellular & Integrative Physiology, IU School of Medicine
    Trivalent chromium (Cr3+) is known to improve glucose homeostasis. Cr3+ has been shown to improve plasma membrane-based aspects of glucose transporter GLUT4 regulation and increase activity of the cellular energy sensor 5′ AMP-activated protein kinase (AMPK). However, the mechanism(s) by which Cr3+ improves insulin responsiveness and whether AMPK mediates this action is not known. In this study we tested if Cr3+ protected against physiological hyperinsulinemia-induced plasma membrane cholesterol accumulation, cortical filamentous actin (F-actin) loss and insulin resistance in L6 skeletal muscle myotubes. In addition, we performed mechanistic studies to test our hypothesis that AMPK mediates the effects of Cr3+ on GLUT4 and glucose transport regulation. Hyperinsulinemia-induced insulin-resistant L6 myotubes displayed excess membrane cholesterol and diminished cortical F-actin essential for effective glucose transport regulation. These membrane and cytoskeletal abnormalities were associated with defects in insulin-stimulated GLUT4 translocation and glucose transport. Supplementing the culture medium with pharmacologically relevant doses of Cr3+ in the picolinate form (CrPic) protected against membrane cholesterol accumulation, F-actin loss, GLUT4 dysregulation and glucose transport dysfunction. Insulin signaling was neither impaired by hyperinsulinemic conditions nor enhanced by CrPic, whereas CrPic increased AMPK signaling. Mechanistically, siRNA-mediated depletion of AMPK abolished the protective effects of CrPic against GLUT4 and glucose transport dysregulation. Together these findings suggest that the micronutrient Cr3+, via increasing AMPK activity, positively impacts skeletal muscle cell insulin sensitivity and glucose transport regulation.
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    Identification of an Actin-Based Antidiabetic Action of Chromium in Skeletal Muscle
    (Office of the Vice Chancellor for Research, 2010-04-09) Hoffman, Nolan J.; Habegger, Kirk M.; Elmendorf, Jeffrey S.
    We recently demonstrated that cortical filamentous actin (F-actin) loss contributes to cellular insulin resistance induced by hyperinsulinemia. New animal and human analyses suggest a similar loss of F-actin is present in insulin-resistant skeletal muscle and results from cellular cholesterol accrual. Interestingly, we found that chromium picolinate (CrPic), a dietary supplement recognized to improve insulin action, lowers plasma membrane cholesterol in cultured adipocytes. Understanding whether CrPic can improve F-actin structure in insulinresistant skeletal muscle via lowering membrane cholesterol is not known, yet significant, as skeletal muscle is responsible for a large majority of insulin-stimulated glucose transport. In L6 myotubes stably expressing the insulin-responsive glucose transporter GLUT4 carrying an exofacial myc-epitope tag, acute insulin stimulation (20 min, 100 nM) increased myc-epitope labeling at the surface of intact cells by ~2-fold (P<0.05). In contrast, the ability of insulin to stimulate this process was inhibited 25% (P<0.05) by sustained exposure of L6 myotubes to insulin (12 h, 5 nM). Defects in insulin signaling did not readily account for the observed disruption. However, we found that insulin-induced insulin-resistant myotubes displayed a 28% elevation (P<0.05) in membrane cholesterol with a reciprocal 14% loss (P<0.05) in F-actin. This cholesterol/actin imbalance and insulin/GLUT4 dysfunction was corrected by the cholesterollowering action of CrPic. Mechanistically, CrPic increased the activity of the AMP-activated protein kinase (AMPK). Tests also revealed that other well-recognized activators of AMPK (e.g., AICAR, DNP) lowered membrane cholesterol and that, in a fashion similar to that witnessed for CrPic, improved regulation of GLUT4 in insulin-induced insulin-resistant myotubes. These data, as well as findings from ongoing siRNA-mediated AMPK knockdown experiments, are consistent with AMPK mediating its antidiabetic action by lowering cellular cholesterol. We predict that chromium, via AMPK activation, protects against cholesterol accrual that induces skeletal muscle F-actin loss and insulin resistance.
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    Membrane cholesterol balance in exercise and insulin resistance
    (2009-10) Habegger, Kirk M.; Elmendorf, Jeffrey S.; Roach, Peter J.; Brozinick, Joseph T.; Sturek, Michael S.; Considine, Robert V.
    Study has shown that plasma membrane (PM) cholesterol and cortical filamentous actin (F-actin) influence skeletal muscle glucose transport. Of fundamental and clinical interest is whether diabetogenic insults promote membrane/cytoskeletal dysfunction amendable for therapy. As exposure to excess fatty acid (FA)s induce glucose intolerance by mechanisms imperfectly understood, we tested if PM cholesterol/F-actin changes could contribute to FA-induced glucose transporter GLUT4 dysregulation in skeletal muscle. High-fat fed, insulin-resistant animals displayed elevated levels of skeletal muscle PM cholesterol and a loss in cortical F-actin, compared to normal-chow fed animals. Consistent with a PM cholesterol component of glucose intolerance, human skeletal muscle biopsies revealed an inverse correlation between PM cholesterol and whole-body glucose disposal. Mechanistically, exposure of L6 myotubes to the saturated FA palmitate induced an increase in PM cholesterol that destabilized actin filaments and decreased insulin-stimulated PM GLUT4 and glucose transport, which could be reversed with cholesterol lowering. Next, study tested if the lipid-lowering action of the antidiabetic AMP-activated protein kinase (AMPK) had a beneficial influence on PM cholesterol balance. Consistent with AMPK inhibition of 3-hydroxy-3-methylglutaryl CoA reductase, a rate-limiting enzyme of cholesterol synthesis, we found that AMPK activation promoted a significant reduction in PM cholesterol and amplified basal and insulin-stimulated GLUT4 translocation. A similar loss of PM cholesterol induced by β-cyclodextrin caused an analogous enhancement of GLUT4 regulation. Interestingly, PM cholesterol replenishment abrogated the AMPK effect on insulin, but not basal, regulation of GLUT4 translocation. Conversely, AMPK knockdown prevented the enhancement of both basal and insulin-stimulated GLUT4 translocation. As a whole these studies show PM cholesterol accrual and cortical F-actin loss uniformly in skeletal muscle from glucose-intolerant mice, swine, and humans. In vivo and in vitro dissection demonstrated this membrane/cytoskeletal derangement induces insulin resistance and is promoted by excess FAs. Parallel studies unveiled that the action of AMPK entailed lowering PM cholesterol that enhanced the regulation of GLUT4/glucose transport by insulin. In conclusion, these data are consistent with PM cholesterol regulation being an unappreciated aspect of AMPK signaling that benefits insulin-stimulated GLUT4 translocation during states of nutrient excess promoting PM cholesterol accrual.