Activation of the Nrf2 Pathway by Inorganic Arsenic in Human Hepatocytes and the Role of Transcriptional Repressor Bach1

Abstract

Previous studies have proved that the environmental toxicant, inorganic arsenic, activates nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in many different cell types. This study tried to explore the hepatic Nrf2 pathway upon arsenic treatment comprehensively, since liver is one of the major target organs of arsenical toxicity. Our results showed that inorganic arsenic significantly induced Nrf2 protein and mRNA expression in Chang human hepatocytes. We also observed a dose-dependent increase of antioxidant response element- (ARE-) luciferase activity. Both the mRNA and protein levels of NAD(P)H:quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1) were all upregulated dramatically. On the other hand, entry and accumulation of Nrf2 protein in the nucleus, while exportting the transcriptional repressor BTB and CNC homology 1 (Bach1) from nucleus to cytoplasm, were also confirmed by western blot and immunofluorescence assay. Our results therefore confirmed the arsenic-induced Nrf2 pathway activation in hepatocytes and also suggested that the translocation of Bach1 was associated with the regulation of Nrf2 pathway by arsenic. Hepatic Nrf2 pathway plays indispensable roles for cellular defenses against arsenic hepatotoxicity, and the interplay of Bach1 and Nrf2 may be helpful to understand the self-defensive responses and the diverse biological effects of arsenicals.

Frequent concerted genetic mechanisms disrupt multiple components of the NRF2 inhibitor KEAP1/CUL3/RBX1 E3-ubiquitin ligase complex in thyroid cancer

“…Bach1, competes with NRF2 protein for binding to target genes, leading to inhibited expression of NQO1 and HO-1 for example [17-19]. These results indicate that in addition to DNA disruption to NRF2 complex inhibitors, the NRF2 pathway is also likely activated by increased activity of NRF2 activators and decreased activity of NRF2 gene target transcriptional repressors in PTCs.…”

The Relationship between Liver Dysfunction and Arsenic Methylation in Mice

“…The control group received normal saline (NS). A previous report showed that both 1 mg/Kg/day and 5 mg/kg ATO induced liver injury but did not affect survival in mice [18,19]. Therefore, the ATO group received 1 mg/Kg/day, 2 mg/Kg/day, and 4 mg/kg/day ATO in our study.…”

Utility of Checklist to Describe Experimental Methods for Investigating Molecular Biomarkers

“…Citation of the gene accession number was found in approximately the 29% of publications ( Figure 1A & Supplementary Tables 1 & 2), whereas details on PCR conditions (time, temperature and number of cycles) were reported only in the 33.3% (7/21) of NRF2 and 50% (7/14) of Keap1 research articles ( Figure 1A & Supplementary Tables 1 & 2) [14][15][16][17][18][19]21,22]. The values, shown as percentage of articles in Figure 1A, were obtained from the analysis of data reported in Supplementary Tables 1 & 2. Since protein immunodetection was not performed in all the examined publications, we analyzed only those articles reporting IB, IC-IHC techniques (21 articles for NRF2 and 14 for Keap1) [15-3436-42,44,45].…”

“…The values, shown as percentage of articles in Figure 1A, were obtained from the analysis of data reported in Supplementary Tables 1 & 2. Since protein immunodetection was not performed in all the examined publications, we analyzed only those articles reporting IB, IC-IHC techniques (21 articles for NRF2 and 14 for Keap1) [15-3436-42,44,45]. Surprisingly, except for the antibody manufacturer, information on the primary antibody clonality, catalogue number and working dilution was found in half or in less than half of the published manuscripts, as shown in Figure However, while in certain articles the antibody details were included [17][18]30,34,37], in others it was possible to infer the missing information from the technical datasheet on the manufacturer website and from the knowledge of the secondary antibody idiotype [16,31,33,36,42].…”

“…Some of the papers analyzed here contributed to define the antioxidant and detoxification properties of the transcription factor NRF2 and its cofactor Keap1, in cells exposed to environmental and work-related toxic agents. Among these we note heavy metals [17] and aromatic polycyclic hydrocarbons [19], the toxicity of which is known, and new emerging materials such as nanoparticles, whose potential toxicity is under investigation [15,28]. Due to the emerging role of the NRF2/Keap1 pathway in the protection of human body against drug toxicity and stress-induced diseases, we believe the publication of primer sequences to quantify the mRNA levels as well as of validated antibodies to detect and localize the relative proteins, should be reliable.…”