BACKGROUND/AIMS: Apoptosis is a key event in the pathophysiology of many liver diseases. Primary human hepatocytes (PHH) provide a useful model to study physiological and pathophysiological processes in the liver. Our aim was to optimize PHH cultures to allow studies on induction of apoptosis and of hepatitis B virus (HBV) infection. METHODS: PHH were isolated from human liver tissue by two-step collagenase perfusion. PHH and hepatoma cells were treated with different apoptosis-inducing agents in parallel. PHH cultures were infected with wild type HBV and transduced with HBV genomes using adenoviral vectors. RESULTS: PHH were successfully isolated from 40 different tissue samples with high viability and purity. Perfusion time and seeding density turned out to be critical parameters for optimal cell yield and culture conditions, respectively. Serum addition to the medium reduced viability of PHH. PHH allowed reproducible studies of CD95-dependent and -independent apoptosis. Sensitivity towards CD95-mediated apoptosis was markedly higher than in hepatoma cells. PHH could efficiently be infected with HBV, but infection did neither induce apoptosis nor prevent CD95-induced cell death. CONCLUSIONS: Our data show that PHH provide an excellent tool for the investigation of apoptosis induced by agents like death receptor-ligands and hepatotropic viruses.
BACKGROUND/AIMS: Apoptosis is a key event in the pathophysiology of many liver diseases. Primary human hepatocytes (PHH) provide a useful model to study physiological and pathophysiological processes in the liver. Our aim was to optimize PHH cultures to allow studies on induction of apoptosis and of hepatitis B virus (HBV) infection. METHODS:PHH were isolated from human liver tissue by two-step collagenase perfusion. PHH and hepatoma cells were treated with different apoptosis-inducing agents in parallel. PHH cultures were infected with wild type HBV and transduced with HBV genomes using adenoviral vectors. RESULTS:PHH were successfully isolated from 40 different tissue samples with high viability and purity. Perfusion time and seeding density turned out to be critical parameters for optimal cell yield and culture conditions, respectively. Serum addition to the medium reduced viability of PHH. PHH allowed reproducible studies of CD95-dependent and -independent apoptosis. Sensitivity towards CD95-mediated apoptosis was markedly higher than in hepatoma cells. PHH could efficiently be infected with HBV, but infection did neither induce apoptosis nor prevent CD95-induced cell death. CONCLUSIONS: Our data show that PHH provide an excellent tool for the investigation of apoptosis induced by agents like death receptor-ligands and hepatotropic viruses.
Authors: Dawei Cai; Courtney Mills; Wenquan Yu; Ran Yan; Carol E Aldrich; Jeffry R Saputelli; William S Mason; Xiaodong Xu; Ju-Tao Guo; Timothy M Block; Andrea Cuconati; Haitao Guo Journal: Antimicrob Agents Chemother Date: 2012-05-29 Impact factor: 5.191
Authors: Vyas Ramanan; Margaret A Scull; Timothy P Sheahan; Charles M Rice; Sangeeta N Bhatia Journal: Annu Rev Virol Date: 2014-11 Impact factor: 10.431