Nrf2 is an antioxidant transcription factor. AMPK functions as a central regulator of cell survival in response to stressful stimuli. Nrf2 should be coordinated with the cell survival pathway controlled by AMPK, but so far the mechanistic connections remain undefined. This study investigated the role of AMPK in Nrf2 trafficking and its activity regulation. A sub-network integrating neighbor molecules suggested direct interaction between AMPK and Nrf2. In cells, AMPK activation caused nuclear accumulation of Nrf2. In the in vitro kinase and peptide competition assays, AMPK phosphorylated Nrf2 at Ser558 residue (Ser550 in mouse) located in the canonical nuclear export signal. Nrf2 with S550A mutation failed to be accumulated in the nucleus after AMPK activation. Leptomycin B, a nuclear export inhibitor, did not enhance nuclear accumulation of WT-Nrf2 activated by AMPK or phospho-Ser550 mimetic Nrf2 mutant, corroborating that AMPK facilitated nuclear accumulation of Nrf2 probably by inhibiting nuclear export. Activated GSK3β diminished the basal nuclear level of Myc-S550A-Nrf2. Collectively, AMPK phosphorylates Nrf2 at Ser550 residue, which in conjunction with AMPK-mediated GSK3β inhibition promotes nuclear accumulation of Nrf2 for antioxidant response element (ARE)-driven gene transactivation.

Sauchinone, a lignan isolated from Saururus chinensis (Saururaceae), is a diastereomeric lignan with cytoprotective and antioxidant activities in cultured hepatocytes. The effects of sauchinone on the inducible nitric oxide synthase (iNOS), tumor necrosis factor‐α (TNF‐α) and cyclooxygenase 2 (COX‐2) gene expression and on the activation of transcription factors, nuclear factor‐κB (NF‐κB), CCAAT/enhancer‐binding protein (C/EBP), activator protein‐1 (AP‐1) and cAMP‐response element‐binding protein (CREB) were determined in Raw264.7 cells as part of the studies on its anti‐inflammatory effects. Expression of the iNOS, TNF‐α and COX‐2 genes was assessed by Northern and Western blot analyses. NO production was monitored by chemiluminescence detection using a NO analyzer. To identify the transcriptional factors affected by sauchinone, the extents of NF‐κB, C/EBP, AP‐1 and CREB activation were measured. Activation of the transcription factors was monitored by gel mobility shift assay, whereas p65 and I‐κBα were analyzed by immunocytochemical and immunoblot analyses. Sauchinone inhibited the induction of iNOS, TNF‐α and COX‐2 by lipopolysaccharide (LPS) (IC5010 μM) with suppression of the mRNAs. Sauchinone (1–30 μM) inhibited LPS‐inducible nuclear NF‐κB activation and nuclear translocation of p65, which was accompanied by inhibition of I‐κBα phosphorylation. LPS‐inducible increase in the intensity of C/EBP binding to its consensus sequence was also inhibited by sauchinone. The AP‐1, but not CREB, DNA binding activity was weakly inhibited by sauchinone. These results demonstrate that sauchinone inhibits LPS‐inducible iNOS, TNF‐α and COX‐2 expression in macrophages through suppression of I‐κBα phosphorylation and p65 nuclear translocation and of C/EBP and/or AP‐1 activation, which may constitute anti‐inflammatory effects of the lignan. British Journal of Pharmacology (2003) 139, 11–20. doi:10.1038/sj.bjp.0705231

Endoplasmic reticulum (ER) stress is associated with liver injury and fibrosis, and yet the hepatic factors that regulate ER stress-mediated inflammasome activation remain unknown. Here, we report that farnesoid X receptor (FXR) activation inhibits ER stress-induced NACHT, LRR, and PYD domains-containing protein 3 (NLRP3) inflammasome in hepatocytes. In patients with hepatitis B virus (HBV)-associated hepatic failure or non-alcoholic fatty liver disease, and in mice with liver injury, FXR levels in the liver inversely correlated with the extent of NLRP3 inflammasome activation. Fxr deficiency in mice augmented the ability of ER stress to induce NLRP3 and thioredoxin-interacting protein (TXNIP), whereas FXR ligand activation prevented it, ameliorating liver injury. FXR attenuates CCAAT-enhancer-binding protein homologous protein (CHOP)-dependent NLRP3 overexpression by inhibiting ER stress-mediated protein kinase RNA-like endoplasmic reticulum kinase (PERK) activation. Our findings implicate miR-186 and its target, non-catalytic region of tyrosine kinase adaptor protein 1 (NCK1), in mediating the inhibition of ER stress by FXR. This study provides the insights on how FXR regulation of ER stress ameliorates hepatocyte death and liver injury and on the molecular basis of NLRP3 inflammasome activation.

Protein tyrosine phosphatase 1B (PTP1B) inhibits hepatic insulin signaling by dephosphorylating tyrosine residues in insulin receptor (IR) and insulin receptor substrate (IRS). MicroRNAs may modulate metabolic functions. In view of the lack of understanding of the regulatory mechanism of PTP1B and its chemical inhibitors, this study investigated whether dysregulation of specific microRNA causes PTP1B-mediated hepatic insulin resistance, and if so, what the underlying basis is. In high-fat-diet-fed mice or hepatocyte models with insulin resistance, the expression of microRNA-122 (miR-122), the most abundant microRNA in the liver, was substantially down-regulated among those predicted to interact with the 3′-untranslated region of PTP1B messenger RNA (mRNA). Experiments using miR-122 mimic and its inhibitor indicated that miR-122 repression caused PTP1B induction. Overexpression of c-Jun N-terminal kinase 1 (JNK1) resulted in miR-122 down-regulation with the induction of PTP1B. A dominant-negative mutant of JNK1 had the opposite effect. JNK1 facilitated inactivating phosphorylation of hepatocyte nuclear factor 4α (HNF4α) responsible for miR-122 expression, as verified by the lack of HNF4α binding to the gene promoter. The regulatory role of JNK1 in PTP1B induction by a decrease in miR-122 level was strengthened by cell-based assays using isoliquiritigenin and liquiritigenin (components in Glycyrrhizae radix) as functional JNK inhibitors; JNK inhibition enabled cells to restore IR and IRS1/2 tyrosine phosphorylation and insulin signaling against tumor necrosis factor alpha, and prevented PTP1B induction. Moreover, treatment with each of the agents increased miR-122 levels and abrogated hepatic insulin resistance in mice fed a high-fat diet, causing a glucose-lowering effect. Conclusion: Decreased levels of miR-122 as a consequence of HNF4α phosphorylation by JNK1 lead to hepatic insulin resistance through PTP1B induction, which may be overcome by chemical inhibition of JNK. (HEPATOLOGY 2012;56:2209–2220)