Analyzing the Effects of HDAC Inhibitors on DNA Damage and Associated Cytotoxicity in Primary Hepatocytes.

Primary hepatocytes are the gold standard in pharmaco- and toxicokinetic studies during preclinical development of drug candidates. Such cells are a valuable tool to identify potential hepatotoxicity, an important adverse drug reaction. Primary hepatocytes can be obtained not only from wild-type mice but also from genetically engineered knockout mouse strains. Liver perfusion yields murine primary hepatocytes (mpH) with high vitality, expressing an array of metabolic enzymes and transporters that are impaired or even absent in established liver cell lines. Furthermore, mpH display no genetic alterations and are proficient in the DNA damage response pathway. This makes mpH a suitable model to analyze the effects of histone deacetylase inhibitors on DNA damage and cell viability. Here, we report an efficient and fast protocol for the isolation of mpH by liver perfusion. These mpH can be used for downstream applications such as the detection of the DNA damage marker γH2AX by confocal laser scanning microscopy.