Toll‐like receptor 4 (TLR4) mediates the induction of cPEPCK expression by palmitate in HepG2 cells

Saturated FFA can activate inflammatory cascades including the toll‐like receptor 4 (TLR4) pathway. TLR4 is expressed by hepatocytes and it may be involved in mechanisms linking FFA to altered liver function, such as gluconeogenesis, in insulin resistance. We tested the role of TLR4 in mediating palmitate effects on promoter activity of cPEPCK, a rate‐controlling gluconeogenic enzyme. HepG2 cells were cotransfected with a reporter gene plasmid encoding the human cPEPCK promoter and siRNA targeting TLR4 or a scrambled sequence. Cotransfected HepG2 cells were incubated in DMEM ± 1 mM palmitate for 24 h with and without a TLR4 decoy peptide (1 μM) which blocks TLR4 signal transduction. Palmitate doubled cPEPCK promoter activity ( P < 0.05 vs. control), and TLR4 knockdown completely ablated this response ( P < 0.05 vs. palmitate alone). We also found that palmitate significantly increased nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) reporter gene activity, which was unaffected by TLR4 decoy treatment or TLR4 knockdown (all P < 0.05 vs. control). Palmitate may have induced cPEPCK promoter activity via TLR4‐dependent, NF‐κB‐independent pathways. Palmitate caused a 20‐fold increase in mRNA of hepatocyte‐specific cyclic AMP response element binding protein (CREBH), another transcriptional regulator of cPEPCK, and the TLR4 decoy peptide significantly decreased this response ( P < 0.05). These results suggest that TLR4 signaling could play a critical role in linking elevated FFA concentrations to increased transcription of gluconeogenic genes.