Cross-talk between the hypoxia sensing pathway and the insulin signaling pathway in the mouse liver
- Vascular endothelial growth factor (VEGF) signaling plays key roles in vascular development and is a therapeutic target in the treatment of malignancies. By blocking VEGF from binding to the VEGF receptor in mice pharmacologically, we show that VEGF inhibition is associated with a decrease in serum glucose levels and a suppression of hepatic glucose production. In adult wild-type mice, chronic VEGF inhibition results in lower fasting and post-prandial glucose levels and enhanced glucose tolerance. Using a hyperinsulinemic euglycemic clamp study, we find that VEGF inhibition leads to suppression of hepatic glucose production via sensitization of hepatic insulin signaling. Furthermore, gene expression analysis of livers from VEGF-inhibitor treated mice shows decreased expression of the glucose-6-phosphatase catalytic subunit (G6pc) and phosphoenolpyruvate carboxykinase (Pepck), both enzymes involved in hepatic gluconeogenesis. We also observe an up-regulation of canonical HIF-2[alpha] target genes in livers from VEGF inhibited mice, and find that HIF-2[alpha] is required for VEGF-inhibitor mediated suppression of hepatic glucose production as mice genetically lacking hepatic Hif2[alpha] (Hif2[alpha]flox/flox Albumin Cre recombinase) do not have the enhanced glucose tolerance and hypoglycemia observed in wild-type mice. Furthermore, mice treated with an adenovirus expressing a constitutively active Hif-2[alpha] allele (Ad-Hif2[alpha]PN) results in a phenocopy of VEGF inhibitor treated mice, indicating that activation of HIF-2[alpha] mediated signaling is not only necessary but also sufficient for suppression of hepatic gluconeogenesis. Western blot analysis of liver extracts from VEGF inhibitor treated mice show increased levels of two key downstream mediators of insulin receptor signaling, insulin receptor substrate 2 (IRS-2) and phosphorylated-Akt (p-Akt) when compared with controls. In fact, Ad-Hif2[alpha]PN requires Irs-2 expression to sensitize insulin signaling in primary hepatocytes, since shRNA mediated depletion of IRS-2 reduces the expected phosphorylation of Akt in response to exogenous insulin stimulation. Furthermore, we show that Hif2[alpha]PN suppresses expression of Srepb-1c, a negative regulator of Irs-2, thereby increasing expression of Irs-2. To address whether modulation of HIF-2[alpha] activity by VEGF-inhibitors may be a potential therapeutic target in diabetes, we treated diabetic, insulin resistant db/db mice with VEGF inhibitors and find significant improvement in glucose tolerance, decreases in fasting as well as post-prandial glucose levels and correction of hyperinsulinemia in a Hif-2[alpha]-dependent manner. In summary, we present evidence of crosstalk between hypoxia and insulin signaling pathways via HIF-2[alpha] modulation and that stimulation of hepatic HIF-2[alpha] may be a therapeutic target for treating type II diabetes. A second focus of this thesis examines the role of HIF-2[alpha] in mediating the ability of VEGF inhibitors to increase hematocrit. We show that VEGF inhibitors increase hematocrit in both hepatic Hif-2[alpha]-independent and --dependent ways. VEGF inhibitors require hepatic Hif-2[alpha] to induce hepatic erythropoietin (EPO)-dependent erythrocytosis and polycythemia. Conditional inactivation of hepatic Hif-2[alpha] suppresses the ability of VEGF inhibitors to increase hepatic Epo expression, reticulocytosis and hematocrit. VEGF inhibitors also increase hematocrit through a contraction in blood plasma volume in an EPO- and Hif-2[alpha]-independent manner. This thesis demonstrates that hepatic HIF-2[alpha] has essential roles both in modulating carbohydrate metabolism and inducing expression of Epo. These activities can be manipulated by treatment with VEGF inhibitors.
|Type of resource
|electronic; electronic resource; remote
|1 online resource.
|McGinnis, Lisa Michelle
|Stanford University, Program in Immunology.
|Kuo, Calvin Jay
|Kuo, Calvin Jay
|Giaccia, Amato J
|Giaccia, Amato J
|Statement of responsibility
|Lisa Michelle McGinnis.
|Submitted to the Program in Immunology.
|Thesis (Ph.D.)--Stanford University, 2013.
- © 2013 by Lisa Michelle McGinnis
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