Nrf2 and NF-κB Activation in Asbestos Exposure

The presence of nuclear factor E2-related factor 2 (Nrf2) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) p65 subunit was determined in nuclear extracts isolated from human mesothelial cells exposed to asbestos and harvested at 0, 2, 4, and 8 h post asbestos exposure. Cytoplasmic and nuclear extracts were prepared using a commercially available nuclear extraction kit (Cayman Chemical Company, Ann Arbor, MI, USA). Transcription factor assay kits (Cayman Chemical Company, Ann Arbor, MI, USA) were used to detect nuclear Nrf2 and NF-κB transcription factors, as previously described [17 (link),18 (link)]. The Transcription factor assay kits utilize a specific double-stranded DNA sequence containing the Nrf2 and NF-κB response element, respectively. The data are reported as the ratio of the absorbance at 450 nm (OD₄₅₀) to the nuclear extract protein concentration (μg). Absorbance measurements were obtained using a SpectraMax i3x Multi-Mode microplate reader (Molecular Devices, Sunnyvale, CA, USA).