|Fetuin-A, a liver secreted phosphoprotein and rat homologue of human a2-HS-glycoprotein (AHSG), is a physiological inhibitor of insulin action. Fetuin-A-null mice exhibit improved glucose tolerance and resistance to weight gain on a high-fat diet. Recent human data demonstrate a positive correlation of circulating fetuin-A levels with BMI, insulin resistance, and fatty liver. Since phosphorylation is critical for its inhibitory activity, circulating fetuin-A phosphorylation was assayed in animal models of insulin resistance, diabetes, and obesity. Using a phospho-specific (312Ser) fetuin-A antibody, fetuin-A phosphorylation status was assayed in Zucker diabetic fatty (ZDF) rats, ob/ob and db/db mice, streptozotocin-treated rats, and in a rat model of diet-induced obesity (DIO). The leptin receptor-deficient, insulin resistant ZDF rats had significantly higher levels of phosphorylated fetuin-A compared to their lean counterparts. Likewise, the
leptin deficient, obese ob/ob mice had significantly higher levels of phosphorylated fetuin-A. Consistent with these results, central leptin administration to control and STZ-
treated rats, as well as leptin treatment to human hepatoma cell lines, significantly decreased fetuin-A phosphorylation. To further characterize the regulation of fetuin-A phosphorylation, we demonstrate that insulin treatment downregulates phosphorylation in Hep3B and HepG2 cell lines. Similarly, DIO rats that exhibit elevated levels of insulin, albeit not to a significant degree, demonstrate significantly decreased levels of fetuin-A phosphorylation. We demonstrate that insulin and leptin decrease phosphorylation of fetuin-A. Accordingly, in conditions of mild to moderate elevation of insulin (as in the DIO model), fetuin-A phosphorylation is decreased, mitigating the inhibitory effect of fetuin-A on IR and allowing for increased insulin signaling. In conditions of extreme insulin resistance (ZDF and ob/ob models), where insulin is unable to exert its actions on insulin receptors, fetuin-A phosphorylation is increased, potentially exacerbating the diabetic phenotype. These findings suggest that fetuin-A may play a significant role in the regulation of insulin action.