FFA-induced hepatic insulin resistance in vivo is mediated by PKCδ, NADPH oxidase, and oxidative stress

Abstract

Fat-induced hepatic insulin resistance plays a key role in the pathogenesis of type 2 diabetes in obese individuals. Although PKC and inflammatory pathways have been implicated in fat-induced hepatic insulin resistance, the sequence of events leading to impaired insulin signaling is unknown. We used Wistar rats to investigate whether PKCδ and oxidative stress play causal roles in this process and whether this occurs via IKKβ- and JNK-dependent pathways. Rats received a 7-h infusion of Intralipid plus heparin (IH) to elevate circulating free fatty acids (FFA). During the last 2 h of the infusion, a hyperinsulinemic-euglycemic clamp with tracer was performed to assess hepatic and peripheral insulin sensitivity. An antioxidant, N-acetyl-L-cysteine (NAC), prevented IH-induced hepatic insulin resistance in parallel with prevention of decreased IκBα content, increased JNK phosphorylation (markers of IKKβ and JNK activation, respectively), increased serine phosphorylation of IRS-1 and IRS-2, and impaired insulin signaling in the liver without affecting IH-induced hepatic PKCδ activation. Furthermore, an antisense oligonucleotide against PKCδ prevented IH-induced phosphorylation of p47(phox) (marker of NADPH oxidase activation) and hepatic insulin resistance. Apocynin, an NADPH oxidase inhibitor, prevented IH-induced hepatic and peripheral insulin resistance similarly to NAC. These results demonstrate that PKCδ, NADPH oxidase, and oxidative stress play a causal role in FFA-induced hepatic insulin resistance in vivo and suggest that the pathway of FFA-induced hepatic insulin resistance is FFA → PKCδ → NADPH oxidase and oxidative stress → IKKβ/JNK → impaired hepatic insulin signaling.

Effect of N-acetyl-l-cysteine on insulin resistance caused by prolonged free fatty acid elevation

“…Studies of the effect of antioxidants on FFA-induced insulin resistance have generally not focused on the liver. We have reported that the pathway in the liver through which short-term plasma FFA elevation causes hepatic insulin resistance is as follows: protein kinase C delta (PKCd) activation/NADPH oxidase activation/oxidative stress/blunted insulin signaling (Pereira et al 2014). Taken together with the results shown herein, we can conclude that as the duration of plasma FFA elevation increases from 7 to 48 h, oxidative stress in the liver is normalized.…”

“…The underlying mechanism is unclear, but one possibility is that oxidative stress near the beginning of lipid infusion activates the transcription factor NRF2, which increases antioxidant enzyme expression (Cullinan & Diehl 2006), such that the oxidative stress is overcome by 48 h. Another potential explanation has hepatic PKCd as a key player. Both short-term (Pereira et al 2014) and prolonged (Pereira et al 2013) IH infusions lead to PKCd activation in the liver. Short-term IH infusion activates ERK1/2 in the liver (data unpublished) and it has been reported that PKCd activates ERK (Ueda et al 1996).…”

“…Wang et al (2009) have first shown that the antioxidant N-acetyl-L-cysteine (NAC) improves the whole-body insulin resistance and the reduction of circulating GSH caused by short-term (4 h) lipid infusion. We have recently shown that NAC prevents whole-body, hepatic and peripheral insulin resistance caused by short-term (7 h) IH infusion (Pereira et al 2014). The normalization of hepatic insulin sensitivity by NAC co-infusion was accompanied by prevention of IH-induced increase in hepatic protein carbonyl content.…”

Free Fatty Acids Shift Insulin-induced Hepatocyte Proliferation towards CD95-dependent Apoptosis

“…Saturated FFAs rapidly trigger generation of ROS, which may explain JNK activation. Mitochondria (34,35) and NADPH oxidase were discussed as predominant sources of palmitate-induced ROS generation in the liver (36,37). The FFA-induced JNK signal directs activated EGFR from proliferative signaling toward association with CD95 and proapoptotic signaling.…”

Chinese Purple Yam (<i>Dioscorea alata</i> L.) Extracts Inhibit Diabetes-Related Enzymes and Protect HepG2 Cells Against Oxidative Stress and Insulin Resistance Induced by FFA

“…One of the postulated mechanisms underlying obesityassociated insulin resistance and T2D is oxidative stress, which is defined as an excessive production of reactive oxygen species (ROS) (Matsuda and Shimomura, 2013). Accumulation of lipid within cells is always accompanied with over-production of ROS (Furukawa et al, 2004;Pereira et al, 2014). Treating obese mice with antioxidant agents could attenuate the development of diabetes (Kaneto et al, 1999).…”