Increased Rate of Apoptosis Correlates with Hepatocellular Proliferation in Fischer-344 Rats Following Long-Term Exposure to a Mixture of Groundwater Contaminants

Abstract

Apoptosis was evaluated in the livers of Fischer-344 rats following observations of increased hepatocellular proliferation from exposures, at low parts per million (ppm) levels, to a drinking water mixture of 7 groundwater contaminants during a 6-mo timecourse study. The 7 chemicals used are among the most frequently detected contaminants associated with hazardous waste sites: arsenic, benzene, chloroform, chromium, lead, phenol, and trichloroethylene. Significant increases in 5-bromo-2'-deoxyuridine hepatocellular labeling were present in a unique pattern surrounding large hepatic veins (0.5-2.0 mm). This did not appear to be a regenerative response due to cytotoxicity, as assessed by the absence of increased plasma enzyme activity and the absence of hepatocellular lesions. Immunohistochemical staining for apoptosis, using the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method showed patterns of labeling in treated animals that directly correlated to areas of increased hepatocyte proliferation. Apoptotic activity was maximum at the 1-mo exposure time point, whereas proliferating hepatocytes reached a maximum rate at the 10-day time point. This may have been triggered as a compensatory response to the increased cell proliferation or as a protective response to remove cells with altered DNA due to chemical mixture exposure. The principal findings of this paper are that (a) apoptosis directly correlated with changes in cell proliferation; (b) observed effects were produced by repeated exposures to a relatively low-level chemical mixture; and (c) the TUNEL method detected apoptotic cells at very early and late stages, potentially increasing the observable time period for apoptosis.

Alteration in Cell Kinetics in Control B6C3F1 Mice Infected with Helicobacter hepaticus

“…Increased cell proliferation, as indicated by proliferating cell nuclear antigen (PCNA) immunohistochemistry, was confirmed in the gastric mucosa of ferrets experimentally infected with H. mustelae, which mimics H. pylori gastritis in humans (31). (14), and it has been suggested that a process favoring cell proliferation and survival over cell death may drive the carcinogenic process (2 (Table I). The incidences of disease (studies with H. hepaticus-related hepatitis) in Studies A-E are given in Table I.…”

“…The actual number of mice with and without hepatitis that was selected from these 3 studies was based on presence or absence of lesions and tumors, tissue condition, Positive staining for PCNA was categorized based on cellular distribution and intensity of the brown to black reaction product that correlated with the different phases of the cell cycle as previously reported (6). G1 cells was based on morphologic criteria (9, 11), confirmed immunohistochemically by the TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling) procedure (10), and was evaluated in Studies A, B, and C. The apoptotic index was determined from the TUNEL-stained slides by counting a total of 5,000 randomly selected hepatocytes as previously described (2). Contiguous groups of apoptotic bodies were considered to have originated from a single cell.…”

“…It was suggested that cell death occurred more frequently as the neoplastic process advanced, with more phenotypic diversity and instability. Constan et al (2) proposed that the increase in apoptosis was either a secondary phenomenon, induced in order to regulate the cell number, or the initial response, acting as a protective mechanism by removing DNA-damaged hepatocytes from the cell population and thereby inducing cell proliferation to replace lost cells. In the NTP studies, it may be that the chronic H. hepaticus infection was associated with the release of chemically reactive, potentially genotoxic substances (i.e., nitric oxide and reactive oxygen species), which triggered apoptosis and subsequent cell proliferation (22).…”

Upregulation of Apoptosis with Dietary Restriction: Implications for Carcinogenesis and Aging

“…These include hypomethylation and overexpression of apoptosis-regulating genes such as bcl-2 (21) and/or loss of gap junctional communication (22). Although most experimental evidence points to increased proliferation as the primary stimulus for the compensatory increase in apoptosis, recent reports suggest that a primary increase in apoptosis can also stimulate compensatory proliferation (23,24). These results would suggest a bidirectional responsiveness to either increased apoptosis or to increased mitosis.…”

Specificity in the Metabolic Activation of Chlorinated Ethylenes by Cytochromes P450 in Primary Rat Hepatocytes.

“…Constan et al reported that apoptosis caused by environmental pollutants, including TCE, were directly associated with the enhanced proliferation of hepatocytes. 9) In the case of cocaine treatment, the preceding phenobarbital (PB)-treatment (or the induction of CYP2B1/2) enhanced lactate dehydrogenase (LDH) leakage, and decreased both the glutathione level and the production of lipid peroxide in rat hepatocytes. 10) Although CEs are metabolized by CYP1A1/2, CYP2B1/2, CYP2E1 and CYP2C11, it remains to be determined which forms of CYP are mainly associated with the metabolic activation of individual CEs differing in the number of chlorine substitutions.…”