Effects of Dichloroacetic Acid (DCA) on Apoptosis in Normal, Preneoplastic,
and Neoplastic Liver in the Male B6C3F1 Mouse
R.D. Snyder*, H.W. Carter*, J.H. Carter*, J.L. Pullman*, A.B. DeAngelo**
*Wood Hudson Cancer Research Laboratory, 931 Isabella St., Newport, KY 41071,
USA and **Health Effects Research Laboratory, United States Environmmental
Protection Agency, Research Triangle Park, NC 27711, USA
ABSTRACT
Dichloroacetic acid (DCA) is a complete hepatocarcinogen and tumor promoter
in the male B6C3F1 mouse. The mechanism(s) of its carcinogenicity is unknown.
The purpose of this study was to examine the effects of DCA on apoptosis
in hepatocytes. Male B6C3F1 mice were given DCA in their drinking water
for 5-30 days (short-term study) or 100 weeks (long term study). Animals
were sacrificed and histopathological analysis was conducted on H & E stained
liver sections. Apoptosis was evaluated by in-situ end-labeling
analysis. This assay employs a standard avidin peroxidase diaminobenzidine
(DAB) procedure to detect biotinylated dUTP molecules specifically inserted
into apoptotic cells by terminal deoxynucleotidyl transferase. In short
term studies, the absolute number of apoptotic (DAB positive) cells in each
section was counted microscopically and total area of tissue was determined
by planimetry using a Zeiss IBAS image analysis system. The percent apoptotic
cells was calculated for each group based on cellularity measurements and
subjected to statistical analysis by the Student's t - test. A highly
significant time and dose related inhibition of spontaneous apoptosis was
found in animals receiving drinking water containing 0.5 or 5.0 g/L DCA
for 5, 10, 15, or 30 days. Hepatocyte apoptotic frequencies of 0.84, 0.53,
and 0.24 were found after 5 days of treatment in mice receiving 0, 0.5,
or 5.0 g/L DCA, respectively, and these frequencies were 0.61, 0.26, and
0.14 after 30 days of treatment. Examination of livers from mice having
received DCA for 100 weeks revealed a similar DCA dose-dependent suppression
of apoptosis in histologically normal liver. DCA induced hyperplastic foci,
adenomas, and carcinomas also exhibited a trend toward decreased apoptosis
with increased DCA dose. This time and dose related downregulation of apoptosis
in animals exposed to DCA suggests that the non-genotoxic hepatocarcinogenicity
of this agent may arise in part through a reduced ability to apoptotically
clear spontaneously initiated cells. (This abstract does not reflect EPA
Policy.)
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