Inhibition of Insulin Sensitivity by Free Fatty Acids Requires Activation of Multiple Serine Kinases in 3T3-L1 Adipocytes

Abstract

Insulin receptor substrate (IRS) has been suggested as a molecular target of free fatty acids (FFAs) for insulin resistance. However, the signaling pathways by which FFAs lead to the inhibition of IRS function remain to be established. In this study, we explored the FFA-signaling pathway that contributes to serine phosphorylation and degradation of IRS-1 in adipocytes and in dietary obese mice. Linoleic acid, an FFA used in this study, resulted in a reduction in insulin-induced glucose uptake in 3T3-L1 adipocytes. This mimics insulin resistance induced by high-fat diet in C57BL/6J mice. The reduction in glucose uptake is associated with a decrease in IRS-1, but not IRS-2 or GLUT4 protein abundance. Decrease in IRS-1 protein was proceeded by IRS-1 (serine 307) phosphorylation that was catalyzed by serine kinases inhibitor kappaB kinase (IKK) and c-JUN NH2-terminal kinase (JNK). IKK and JNK were activated by linoleic acid and inhibition of the two kinases led to prevention of IRS-1 reduction. We demonstrate that protein kinase C (PKC) theta is expressed in adipocytes. In 3T3-L1 adipocytes and fat tissue, PKCtheta was activated by fatty acids as indicated by its phosphorylation status, and by its protein level, respectively. Activation of PKCtheta contributes to IKK and JNK activation as inhibition of PKCtheta by calphostin C blocked activation of the latter kinases. Inhibition of either PKCtheta or IKK plus JNK by chemical inhibitors resulted in protection of IRS-1 function and insulin sensitivity in 3T3-L1 adipocytes. These data suggest that: 1) activation of PKCtheta contributes to IKK and JNK activation by FFAs; 2) IKK and JNK mediate PKCtheta signals for IRS-1 serine phosphorylation and degradation; and 3) this molecular mechanism may be responsible for insulin resistance associated with hyperlipidemia.

Molecular Mechanisms of Insulin Resistance: Serine Phosphorylation of Insulin Receptor Substrate-1 and Increased Expression of p85α

“…Blocking JNK activation rescued the cellular and molecular defects induced by free fatty acids (56). Furthermore, JNK-1 knockout mice were found to be resistant to dietinduced obesity and insulin resistance (55).…”

Specific protein kinase C isoforms as transducers and modulators of insulin signaling

“…There appear to be several mechanisms by which PKCθ could be inducing insulin resistance. In adipocytes, FFA activation of the isoform was believed to activate IKK and JNK which mediate IRS-1 serine phosphorylation and degradation [138]. This was demonstrated using a PKC inhibitor, however, and was inferred indirectly in rats fed high fat diets.…”

Effects of over-expressing resistin on glucose and lipid metabolism in mice

“…As a result of insulin resistance in mice, the serum GLU and TG levels increased, and the serum FFA, HDL cholesterol and TC concentrations decreased significantly at different stages of the experiment. It has been reported that glucose and lipid metabolism can be influenced by several adipose tissue-derived factors such as FFA (Gao et al, 2004), TNF-α (Borst et al, 2004), leptin (Friedman, 2000) and adiponectin (Yamauchi et al, 2003). More recently, several studies suggested that insulin resistance is related to pro-inflammatory condition that promotes the production of inflammatory factors such as TNF-α and IL-6 (Bastard et al, 2006).…”