Distinct Mechanisms Are Responsible for Nrf2-Keap1 Pathway Activation at Different Stages of Rat Hepatocarcinogenesis

Abstract

Activation of the Nrf2-Keap1 pathway, the main intracellular defense against environmental stress, has been observed in several human cancers, including hepatocellular carcinoma (HCC). Here, we assessed whether distinct mechanisms of activation may be involved at different stages of hepatocarcinogenesis. We adopted an experimental model consisting of treatment with diethylnitrosamine (DENA) followed by a choline-devoid methionine-deficient (CMD) diet for 4 months. The CMD diet was then replaced with a basal diet, and the animals were killed at 6, 10 or 13 months after DENA injection. Nrf2 activation occurred at early steps of hepatocarcinogenesis and persisted throughout the tumorigenic process. While Nrf2 mutations were extremely frequent at early steps (90%), their incidence diminished with the progression to malignancy (25%). Conversely, while p62 was almost undetectable in early nodules, its accumulation occurred in HCCs, suggesting that Nrf2 pathway activation at late stages is mainly due to Keap1 sequestration by p62. We demonstrate that, in a model of hepatocarcinogenesis resembling human non-alcoholic fatty liver disease, Nrf2 mutations are the earliest molecular changes responsible for the activation of the Nrf2-Keap1 pathway. The progressive loss of mutations associated with a concomitant p62 accumulation implies that distinct mechanisms are responsible for Nrf2-Keap1 pathway activation at different stages of hepatocarcinogenesis.

Kaempferol-3-O-Glucuronide Ameliorates Non-Alcoholic Steatohepatitis in High-Cholesterol-Diet-Induced Larval Zebrafish and HepG2 Cell Models via Regulating Oxidation Stress

“…The Nrf2-Keap1 signaling pathway is a key signaling pathway re-sponsible for the antioxidant defense against oxidation [ 27 ]. It has been widely reported as a potential treatment target of NAFLD in the liver [ 28 ]. Nrf2, as a dominant regulator of the antioxidant response, exists in multiple tissues and protects against oxidation and inflammation [ 29 ].…”

NRF2 and the Ambiguous Consequences of Its Activation during Initiation and the Subsequent Stages of Tumourigenesis

“…The stage of cancer at which mutations in NFE2L2 and KEAP1 arise is currently unknown. However, it has been shown that mutations in NFE2L2 occur in the early stages of HCC, and whilst the prevalence of NFE2L2 mutations decreases as the disease increases in severity, NRF2 activity remains heightened throughout HCC progression [252]. Potentially, this is due to high levels of oxidative stress leading to phosphorylation of p62/SQSTM1, which allows it to compete for binding to KEAP1, resulting in increased NRF2 activity in the later stages of HCC.…”

Nrf2 in Neoplastic and Non-Neoplastic Liver Diseases

“…Animal models allow the dissection of the several steps of hepatocarcinogenesis, such as the rat resistant-hepatocyte (RH) and the choline-methionine deficient (CMD) models [74,115], whose translational value have been established [116,117], have been very useful to address this question. By using these animal models, Columbano et al showed that the dysregulation of the Keap1/Nrf2 pathway takes place in the earliest steps of hepatocarcinogenesis [28,[117][118][119]. In addition, it was shown that the genetic inactivation of Nrf2 fully impaired the clonal expansion of carcinogen-initiated hepatocytes to preneoplastic nodules in rats [28], in spite of the presence of a tumor promoting environment, such as liver damage and compensatory hepatocyte proliferation caused by CMD diet.…”