Isolation and enrichment of preneoplastic hepatocytes during malignant cell transformation.
Ashley-A; Lohitnavy-M; Lu-Y; Chubb-L; Billings-R; Campain-J; Yang-R
Toxicologist 2004 Mar; 78(S-1):225-226
We modified Ito's medium-term liver foci bioassay by incorporating time-course sacrifices to study pharmacokinetics/pharmacodynamics related to the initiation-promotion process. In our earlier clonal growth modeling studies on a series of chlorobenzenes, the probable existence of two cell types in glutathione-S-trasferase (GSTP) foci became apparent. As suggested by other researchers, these two cell types are either susceptible or resistant to mitoinhibitory effects as a response to external chemical insults, and those resistant to mitoinhibitory effect would have a growth advantage serving as basis for malignant transformation. Thus, as preneo-plastic GSTP foci form, isolation of GSTP positive (GSTP+) cells from liver across time allows capturing changing characteristics of an emerging malignant population. This study describes an attempt to isolate primary GSTP+ hepatocytes using Ito's bioassay protocol with diethylnitrosamine (DEN) as an initiating agent and a 2/3 partial hepatectomy with concomitant dosing of hexachlorobenzene (HCB; 28.4 mg/kg/day) with and without coexposure to PCB 126 (9.8 u/kg/day) as promoting agents. Male F344 rats were sacrificed at 20, 24, 28, 47 and 56 days, and 3, 6, and 9 months following DEN injection. By day 56, a significant increase in liver weight was observed in rats coexposed to HCB and PCB 126. Either primary hepatocytes or liver sections were taken to analyze formation of GSTP+ foci. By treating cell suspension and/or culture with ethacrynic acid (EA) (Stenius et al., Carcinogenesis 15:1561-1566, 1994), GSTP+ hepatocytes were enriched by inducing toxicity to non-GSTP+ cells. In so doing, progressively more malignant cell populations can be subjected to cell cycle kinetic studies, genomic analyses, and clonal growth modeling to aid our insight into the carcinogenic process.
Hepatocytes; Cell-transformation; Bioassays; Pharmacodynamics; Models; Laboratory-animals; Animals; Animal-studies; Exposure-levels; Carcinogenesis
Research Tools and Approaches: Risk Assessment Methods
The Toxicologist. Society of Toxicology 43nd Annual Meeting and ToxExpo, March 21-25, 2004, Baltimore, Maryland
Colorado State University - Fort Collins