Abstract 1076: Reversible effect of all-trans-retinoic acid on AML12 hepatocyte proliferation and cell cycle progression

Abstract

Abstract The role of all-trans-retinoic acid (atRA) in the regulation of cellular proliferation and differentiation is well documented. Numerous studies have established the cancer preventive properties of atRA which functions to regulate levels of cell cycle proteins essential for the G1/S transition and progression through S phase. Using AML12 cells, a non-transformed immortalized mouse hepatocyte cell line, the goals of this study were to first determine if changes in AML12 cell proliferation could be observed in response to atRA treatment and establish whether this was a suitable system to investigate molecular mechanisms altered by hepatocarcinogens. Cells were treated with 1µM and 5 µM atRA for 2 to 10 days. At various time points, control (DMSO) and treated cells were enumerated to determine the effect of atRA on AML12 cell proliferation. At 5 days of atRA treatment, there was a 56% decrease in the number of atRA treated cells when compared to control cells. This effect was reversible as an increase in cell number was observed two days after removal of atRA from the treatment media. Cell cycle analysis by flow cytometry using propidium iodide stained AML12 cells showed a significant reduction (p<0.05) in S phase cells and a significant increase (p<0.05) in G1 phase cells with 48 hours of atRA treatment when compared to control cells indicating a block at the G1/S checkpoint. Western analysis of control and treated whole cell lysates revealed a decrease in phosphorylated (Serine807/Serine811) retinoblastoma protein (pRb). Taken together, these results indicate that proliferation in AML12 hepatocytes can be reversibly modified by atRA by regulating these cells at the G1/S checkpoint with a reduction in pRb. Previous studies of liver tissues collected from mice exposed to the hepatocarcinogenic fungicide, propiconazole, have revealed an increase in cellular proliferation, increases in atRA metabolism as well as decreases in the levels of hepatic atRA. Based on the results of the present experiments, AML12 hepatocytes provide an optimal in vitro system to elucidate the molecular mechanism by which propiconazole induces hepatotumorigenesis. This work was reviewed by EPA and approved for publication but does not necessarily reflect official Agency policy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1076.