Identification of Apoptotic Drugs: Multiparametric Evaluation in Cultured Hepatocytes

Maria   Jose   Gomez-Lechon; Jose   Enrique   O'Connor; Agustin      Lahoz; Jose   V.   Castell; Maria   Teresa   Donato

doi:10.2174/092986708785132861

Abstract

It is now recognized that necrosis is not the only mechanism responsible for chemically-induced cell death. It is believed that apoptosis could be the major form of cell death induced by toxicants and that necrosis is associated only with circumstances of gross cell injury. The liver is a key target organ for drug toxicity and an important effort in drugdiscovery deals with the identification of molecules with hepatotoxic potential. The importance of apoptosis in toxicology has been underestimated given the difficulty of identifying apoptotic cells in in vitro models when apoptosis normally degenerates to secondary necrosis. Nowadays, the central role played by apoptosis in the toxicity of many xenobiotics and P450-generated metabolites is recognized. The detection of drug-induced apoptosis constitutes one of the highest priorities of the pharmaceutical industry. Different markers aimed at identifying apoptotic compounds irrespectively of the pathway of how cell apoptosis was initiated have been proposed. The aim of the present paper is to review the utility of some available in vitro strategies for studying drug-induced liver apoptosis. The evaluation of apoptotic or anti-apoptotic effects of chemicals in hepatocytes is illustrated by several examples including model apoptotic compounds, pharmaceutical drugs which have been shown to induce apoptosis as an adverse effect; and drugs preventing apoptosis. By combining appropriated markers, apoptosis can be detected in hepatocytes long before cell necrosis, at sub-cytotoxic concentrations of the drugs. The possibility of using small amounts of cells cultured in multiwell formats and automation has notably contributed to develop reproducible, reliable, sensitive, easy-to-handle and rapid multiparametric assays that are ideally amenable to high throughput screening (HTS).

Keywords: Apoptosis, Bcl2, caspases, DNA fragmentation, cytochrome c, flow cytometry, DNA analysis, hepatocytes