Isoniazid induces apoptosis: Role of oxidative stress and inhibition of nuclear translocation of nuclear factor (erythroid-derived 2)-like 2 (Nrf2)

Abstract

AimsLong-term treatment of Isoniazid (INH) in tuberculosis (TB) patients can lead to anti-tuberculosis drug-induced hepatotoxicity. To understand the mechanism of hepatotoxicity, an attempt has been made to elucidate the role of Nrf2, a transcription factor induced by oxidative stress, in INH induced apoptosis liver cancer cell lines. Materials and methodsCytotoxicity was evaluated by MTT assay. Apoptosis and reactive oxygen species (ROS) generation was performed by flow cytometry. mRNA and protein expression of various genes involves in INH induced toxicity was evaluated via Real-time PCR and western blot analysis respectively. Differential protein expression was performed by two-dimensional gel electrophoresis followed by identification using MALDI TOF/TOF. Key findingsINH induced ROS and apoptosis in HepG2 as well as THLE-2 cells. Nuclear damage was also observed by INH treatment in HepG2 cells. Expression of apoptotic (Cytochrome C and Caspase 9) and antioxidative (Keap1 and Nrf2) genes were observed to increase. INH induced PKCδ phosphorylation and released Nrf2 from its inhibitor Keap1 in the cytoplasm of HepG2 cells. However, over-expression of Nrf2 did not affect nuclear Nrf2 protein level as well as its downstream target NQO1. Nrf2 importer, Karyopherin β1 level was observed to decrease in HepG2 as well as THLE-2 cells following INH treatment. SignificanceThese findings suggest that INH prevented Nrf2 translocation into the nucleus by inhibiting its importer Karyopherin β1. Therefore Nrf2 might not able to bind ARE sequences from inducing antioxidative response for protecting the cells undergoing apoptosis.