Arf6 Heterozygosity Results in Impaired Glucose Tolerance Due to Abberant Hepatic Gluconeogenesis

Abstract

Adenosine diphosphate (ADP) ribosylation factor 6 (Arf6) is a small GTPase that plays a critical role in numerous cellular processes including proliferation and membrane trafficking. Arf6 has been found to be dysregulated in diseases such as diabetic retinopathy and tumor metastasis. Furthermore, genetic insufficiency or pharmacological inhibition of Arf6 is protective in the aforementioned diseases, suggesting that Arf6 may be a potential therapeutic target. Here, we assessed the metabolic consequences of a reduction in Arf6 expression by performing glucose (GTT: 2 g/kg, ip), insulin (ITT: 1 U/kg, ip) and pyruvate (PTT: 2 g/kg, ip) tolerance tests in 3–4 mo old whole body Arf6 heterozygote (Arf6+/−) and wildtype (Arf6+/+) littermate control mice. Both protein and gene expression of Arf6 in the liver were 40–50% lower (p<0.05) in Arf6+/− mice compared to Arf6+/+. Body mass was higher in normal chow fed Arf6+/− compared to Arf6+/+ mice (30.3±0.9 vs 36.4±2.0 g, p<0.05). Blood glucose at 15, 30, and 60 min after injection during the GTT were 20–30% higher in Arf6+/− mice compared to Arf6+/+ (each p<0.05). Likewise, the area under the GTT time response curve was ~15% higher in Arf6+/− compared to Arf6+/+ (p<0.05), indicating impaired glucose metabolism. To examine the underlying cause of this glucose intolerance, we performed an ITT and assessed insulin‐stimulated suppression of plasma free fatty acids 15, and 30 min after insulin injection. We found no groups differences between the ITT time response curves (p>0.05) and no difference in the percent decrease in plasma FFAs from fasted between Arf6+/− and Arf6+/+ mice (~50% vs ~50% p>0.05). Likewise, fasted and glucose‐stimulated (2g/kg, ip) serum insulin were not different between the groups (2.44±0.58 vs 1.73±0.28, fasted; 4.54±1.74 vs 4.03±1.68 insulin stimulated; both p>0.05), suggesting that reductions in Arf6 expression do not impact pancreatic beta cell insulin secretion. Collectively, these measures suggest that neither peripheral insulin resistance nor impaired pancreatic beta cell function underlie glucose intolerance in Arf6+/− mice. To determine the contribution of hepatic gluconeogenesis to the observed glucose intolerance in the Arf6+/− mice, we performed a PTT and found that Arf6+/− mice exhibited a 15–25% increase in blood glucose over time compared to Arf6+/+ mice (p<0.05), suggesting a role for elevated hepatic gluconeogenesis. Taken together, this study demonstrates that Arf6 heterozygosity impairs glucose metabolism that is associated with aberrant hepatic gluconeogenesis but is independent of peripheral insulin resistance or pancreatic beta cell dysfunction.Support or Funding InformationNIA R01 AG048366, R01 AG050238 and K02 AG045339, R01 AG060395 and US Department of Veterans Affairs I01 BX002151, I01 BX004492.