An investigation into the cellular expression of genes involved in resistance to aflatoxin B1, and the mode of cell death elicited by this carcinogen, in a model system using AFB1-resistant and AFB1-sensitive rat hepatocyte cell lines
thesisposted on 15.12.2014, 10:29 by Donna. Richardson
The toxicity of aflatoxin B1 (AFB1), has been extensively studied in the rat, and its carcinogenic effects have been well documented. Binding of the cytochrome P-450-activated exo-8,9-epoxide form of the molecule to DNA and protein to form adducts, forms the basis of the xenobiotic's cytotoxicity and genotoxicity.;In the rat, the major detoxification pathway is its glutathione-S-transferase (GST)-mediated conjugation with glutathione. An alternative route for the AFB1-8,9-epoxide is hydrolysis to give the cytotoxic AFB1-dihydrodial (AFB1-dhd). The rat GST A5 subunit, exhibits a high catalytic activity for the AFB1-exo-8,9-epoxide, and its protective effect towards AFB1 has been shown. In addition, the expression of a novel AFB1-aldehyde reductase (rAFAR1) gene has been investigated and found to be expressed in the AFB1-sensitive RLE cell line as well as in the AFB1-resistant JB1 cell line, along with the GST A3 and the p-glycoprotein (mdr1b) gene. A model system was devised for the study of AFB1 toxicity and the need for optimisation of all assay conditions is stressed, in particular the concentration of quail liver microsomes (an extrinsic AFB1 activating system). The toxic effect of the microsomes on the RLE cell line has been demonstrated and a concentration of 1% (v/v) in unsupplemented medium (as an AFB1 diluent) is recommended.;Cell death in response to AFB1 toxicity is important since the elimination of individual initiated cells which are induced to undergo apoptosis (programmed cell death) will prevent their clonal expansion and ultimate promotion and progression to tumours.