e Transcription factor Nrf2 (NF-E2-related factor 2) regulates a broad cytoprotective response to environmental stresses. Keap1 (Kelch-like ECH-associated protein 1) is an adaptor protein for cullin3-based ubiquitin E3 ligase and negatively regulates Nrf2. Whereas the Keap1-Nrf2 system plays important roles in oxidative stress response and metabolism, the roles Nrf2 plays in the prevention of diabetes mellitus remain elusive. Here we show that genetic activation of Nrf2 signaling by Keap1 gene hypomorphic knockdown (Keap1 flox/؊ ) markedly suppresses the onset of diabetes. When Keap1 flox/؊ mice were crossed with diabetic db/db mice, blood glucose levels became lower through improvement of both insulin secretion and insulin resistance. Keap1 flox/؊ also prevented high-calorie-diet-induced diabetes. Oral administration of the Nrf2 inducer CDDO-Im {oleanolic acid 1-[2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oyl] imidazole} also attenuated diabetes in db/db mice. Nrf2 induction altered antioxidant-, energy consumption-, and gluconeogenesis-related gene expression in metabolic tissues. Thus, the Keap1-Nrf2 system is a critical target for preventing the onset of diabetes mellitus.

Nrf2 (NF-E2-related factor 2) plays a key role in the protection of vertebrates against environmental stress by contributing to the inducible expression of detoxification and antioxidant enzymes. Keap1 (Kelch-like ECH-associated protein 1) is a sensor for oxidative and electrophilic stresses. Keap1 also acts as an E3 ubiquitin ligase substrate-recognition subunit that specifically targets Nrf2. Keap1 causes Nrf2 to be degraded through the ubiquitin-proteasome pathway and thus ensures that Nrf2 is constitutively suppressed under unstressed conditions. Upon exposure to oxidative or electrophilic stress, Keap1 loses its ability to ubiquitinate Nrf2. Many lines of evidence have recently clarified that the Keap1-Nrf2 system also plays critical roles in the maintenance of cellular homeostasis. One of the most salient examples is the contribution of Keap1-Nrf2 to metabolic and energy-balance regulation. In particular, how the Keap1-Nrf2 system protects the body against diabetes mellitus and how perturbations in this system provoke the disease condition are now under intense investigation. This review will summarize the recent progress made in this area.

Transcription factor Nrf2 (NF-E2-related factor 2) regulates wide-ranging cytoprotective genes in response to environmental stress. Keap1 (Kelch-like ECH-associated protein 1) is an adaptor protein for Cullin3-based ubiquitin E3 ligase and negatively regulates Nrf2. The Keap1-Nrf2 system plays important roles in the oxidative stress response and metabolism. However, the roles Nrf2 plays in prevention of pancreatic b-cell damage remain elusive. To demonstrate the roles of Nrf2 in pancreatic b-cells, we used four genetically engineered mouse models: 1) b-cell-specific Keap1-conditional knockout mice, 2) b-cell-specific Nos2 transgenic mice, 3) conventional Nrf2-heterozygous knockout mice, and 4) b-cell-specific Nrf2-conditional knockout mice. We found that Nrf2 induction suppressed the oxidative DNA-adduct formation in pancreatic islets of iNOS-Tg mice and strongly restored insulin secretion from pancreatic b-cells in the context of reactive species (RS) damage. Consistently, Nrf2 suppressed accumulation of intracellular RS in isolated islets and pancreatic b-cell lines and also decreased nitrotyrosine levels. Nrf2 induced glutathione-related genes and reduced pancreatic b-cell apoptosis mediated by nitric oxide. In contrast, Nrf2 depletion in Nrf2-heterozygous knockout and b-cell-specific Nrf2-conditional knockout mice strongly aggravated pancreatic b-cell damage. These results demonstrate that Nrf2 induction prevents RS damage in pancreatic b-cells and that the Keap1-Nrf2 system is the crucial defense pathway for the physiological and pathological protection of pancreatic b-cells.

p62/SQSTM1 is a selective substrate of autophagy, and aberrant accumulation of p62 has been observed in various pathological conditions. To understand the roles p62 plays in non-small-cell lung cancer (NSCLC), we carried out immunohistochemical analyses of p62 expression in a cohort of patients with annotated clinicopathological data. As analyses of murine and human hepatocellular carcinomas suggested a correlation between p62 and Nrf2 accumulations, we also examined NRF2 expression in the same cohort. The expression of NRF2 and p62 was examined by immunohistochemical methods in 109 NSCLC cases, which included patients with adenocarcinoma (n = 72), squamous cell carcinoma (n = 31), and large cell carcinoma (n = 6). Accumulation of NRF2 and p62 was detected in 34% and 37% of NSCLC patients, respectively. The accumulations of p62 and NRF2 did not correlate with each other, but both were associated with worse lung cancer-specific survival (P = 0.0003 for NRF2; P = 0.0130 for p62). NRF2 status had an impact on NSCLC prognosis irrespective of histology types, but p62 status did so particularly in adenocarcinoma (P = 0.037). Multivariate analysis indicated that positive immunoreactivities of NRF2 and p62 were both independent factors predicting worse lung cancer-specific survival (P < 0.0001 for NRF2 and P = 0.04 for p62). This study revealed that both NRF2 and p62 are independent prognostic factors for NSCLC. The prognostic impact of p62 status was pronounced in adenocarcinoma patients, suggesting that molecular mechanisms underlying cancer evolution differ between adenocarcinoma and squamous cell carcinoma. (Cancer Sci 2012; 103: 760-766) N on-small-cell lung cancer (NSCLC), which comprises mainly adenocarcinoma and squamous cell carcinoma, is one of the most common human cancers.(1) Despite rigorous endeavors to develop anticancer therapies, the prognosis of lung cancer patients still remains poor. To predict recurrence after surgery and to judge indications of additional therapies appropriately, clinical and biological markers have long been sought.The Keap1-Nrf2 system plays a central role in protecting cells from electrophilic and oxidative stresses.(2) Under unstressed conditions, Keap1 ubiquitinates Nrf2, and Nrf2 is degraded by the proteasome. Upon exposure to the stimuli, Keap1 is inactivated, and stabilized Nrf2 induces the transcription of many cytoprotective genes. Somatic mutations in the NRF2 or KEAP1 gene that cause constitutive stabilization of NRF2 have been found in many human cancers.(3-9) Reduced expression of KEAP1 due to KEAP1 methylation has been suggested as another mechanism for NRF2 stabilization.(10,11) NRF2 stabilization and subsequent accumulation contribute to the poor prognosis of NSCLC patients. (6,7) This is because NRF2 confers resistance to anticancer therapies and an aggressive proliferative tendency on cancer cells. (10,12) Recently, still another cause for the constitutive stabilization of NRF2 in cancer cells was reported. One of the selective substrates for autophagy, (13,1...

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f Nrf2 (NF-E2-related factor 2) contributes to the maintenance of glucose homeostasis in vivo. Nrf2 suppresses blood glucose levels by protecting pancreatic ␤ cells from oxidative stress and improving peripheral tissue glucose utilization. To elucidate the molecular mechanisms by which Nrf2 contributes to the maintenance of glucose homeostasis, we generated skeletal muscle (SkM)-specific Keap1 knockout (Keap1MuKO) mice that express abundant Nrf2 in their SkM and then examined Nrf2 target gene expression in that tissue. In Keap1MuKO mice, blood glucose levels were significantly downregulated and the levels of the glycogen branching enzyme (Gbe1) and muscle-type PhK␣ subunit (Phka1) mRNAs, along with those of the glycogen branching enzyme (GBE) and the phosphorylase b kinase ␣ subunit (PhK␣) protein, were significantly upregulated in mouse SkM. Consistent with this result, chemical Nrf2 inducers promoted Gbe1 and Phka1 mRNA expression in both mouse SkM and C2C12 myotubes. Chromatin immunoprecipitation analysis demonstrated that Nrf2 binds the Gbe1 and Phka1 upstream promoter regions. In Keap1MuKO mice, muscle glycogen content was strongly reduced and forced GBE expression in C2C12 myotubes promoted glucose uptake. Therefore, our results demonstrate that Nrf2 induction in SkM increases GBE and PhK␣ expression and reduces muscle glycogen content, resulting in improved glucose tolerance. Our results also indicate that Nrf2 differentially regulates glycogen metabolism in SkM and the liver.T he tight regulation of glucose homeostasis is essential for the maintenance of biological functions. As glucose metabolites are major energy sources for skeletal muscle (SkM) contraction (1-3), SkM requires an efficient supply of glucose metabolites during exercise. Perturbations in SkM glucose metabolism often provoke metabolic disorders; e.g., impaired glucose tolerance and diabetes mellitus. SkM and liver store glucose as glycogen (4), which is used to generate glucose metabolites when energy is required; consequently, efficient SkM glycogen utilization is an important factor in exercise and the maintenance of glucose homeostasis (4).Our bodies are continuously exposed to toxic chemicals (often electrophiles) and oxidative stress from the environment; these stresses are termed environmental stresses (5). While these environmental stresses are known to provoke tissue damage, they also affect cellular metabolism and energy production (5). The Keap1-Nrf2 system protects our bodies against these environmental stresses (6, 7). Nrf2 (NF-E2-related factor 2) belongs to the cap 'n' collar subfamily of basic region-leucine zipper-type transcription factors (8). Under unstressed conditions, Keap1 (Kelch-like ECHassociated protein 1) constitutively represses Nrf2 activity (9) by acting as an adaptor subunit for cullin-3-based ubiquitin E3 ligase (7). This E3 ligase complex efficiently ubiquitinates Nrf2, leading to its rapid proteasomal degradation (10). When cells are exposed to either electrophilic toxic chemicals or reactive oxygen spec...

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