Stable Transfection of LLC-PK1 Cells with Human Microsomal GlutathioneS-Transferase Gene Increases Haloalkene GlutathioneS-conjugate Formation and Cytotoxicity

Abstract

Nephrotoxic haloalkenes undergo glutathione- and cysteine conjugate beta-lyase-dependent bioactivation, and glutathione S-conjugate formation with haloalkenes as substrates is preferentially catalyzed by the hepatic microsomal glutathione S-transferase (mGST). Porcine kidney-derived LLC-PK1 cells, which are competent to bioactivate glutathione and cysteine S-conjugates of haloalkenes, show low mGST activity. Stable transfection of LLC-PK1 cells with the gene encoding mGST would be expected to increase glutathione S-conjugate formation and, therefore, to increase haloalkene cytotoxicity. Transfection of LLC-PK1 cells with human mGST genes resulted in increased expression of mGST protein in microsomal fractions, in increased glutathione S-conjugate formation with hexachloro-1,3-butadiene and 1-chloro-2,4-dinitrobenzene as the substrates, and in increased cytotoxicity of hexachloro-1,3-butadiene. In addition, transfection with mGST gene also increased the activity of cytosolic glutathione S-transferases.