Quantitative histochemistry of benzaldehyde dehydrogenase in hepatocellular carcinomas of vinyl chloride-treated rats

Hepatocarcinogenesis in rats treated with several chemicals is associated with changes in aldehyde dehydrogenase (AlDH) activity, particularly heterogeneous expression of a "tumor specific" phenotype that is very active with aromatic aldehydes, e.g., benzaldehyde (Bz). Objectives of this study were first, to determine if liver cancers in vinyl chloride-treated rats also expressed this AlDH phenotype, and second, to quantitate the NAD- and NADP-dependent AlDH activity for the substrates Bz and acetaldehyde (Ac) in the cancers and surrounding tissue. Small cubes of tissue containing well-differentiated hepatocellular carcinoma were obtained from five Sprague-Dawley rats exposed to 2500 ppm vinyl chloride for 55 weeks. An optimized procedure was developed for AlDH histochemistry. Frozen sections were preincubated in nitroblue tetrazolium/acetone and then incubated at 20 degrees C in viscous polyvinyl alcohol media containing buffer, phenazine methosulfate, sodium azide, substrate, coenzyme, and nitroblue tetrazolium. Background activity was evaluated by omission of substrate. Activity was quantitated by computer-assisted microscopic photometry. All five carcinomas had heterogeneous staining of NADP- and NAD-dependent BzDH and AcDH activity, with clusters of very high-activity cells. The magnitude of staining in the high-activity neoplastic cells was at least tenfold greater for BzDH-NADP and about twofold greater for BzDH-NAD, AcDH-NADP, and AcDH-NAD than the staining in other liver cells. More neoplastic cells had high BzDH than high AcDH activity. Only BzDH-NADP was localized predominantly to the carcinoma.