Human Hepatocytes as a Tool for Studying Toxicity and Drug Metabolism

Abstract

Drugs are usually biotransformed into new chemical species that may have either toxic or therapeutic effects. Drug metabolism studies are routinely performed in laboratory animals but, due to metabolic interspecies differences when compared to man, they are not accurate enough to anticipate the metabolic profile of a drug in humans. Human hepatocytes in primary culture provide the closest in vitro model to human liver and the only model that can produce a metabolic profile of a given drug that is very similar to that found in vivo. However their availability is limited due to the restricted access to suitable tissue samples. The scarcity of human liver has led to optimising the cryopreservation of adult hepatocytes for long-term storage and regular supply. Human hepatocytes in primary culture express typical hepatic functions and express drug metabolising enzymes. Moreover, qualitative and quantitative similarities between in vitro and in vivo metabolism of drugs were observed. Different strategies have been envisaged to prolong cell survival and delay the spontaneous decay of the differentiated phenotype during culture. Thus, hepatocytes represent the most appropriate model for the evaluation of integrated drug metabolism, toxicity/metabolism correlations, mechanisms of hepatotoxicity, and the interactions (inhibition and induction) of xenobiotics and drug-metabolising enzymes. However, in view of limitations of primary hepatocytes, efforts are made to develop alternative cellular models (i.e. metabolic competent CYP-engineered cells stably expressing individual CYPs and transient expression of CYPs by transduction of hepatoma cells with recombinant adenoviruses). In summary, several cellular tools are available to address key issues at the earliest stages of drug development for a better candidate selection and hepatotoxicity risk assessment.

Similarities and Differences in the Expression of Drug-Metabolizing Enzymes between Human Hepatic Cell Lines and Primary Human Hepatocytes

“…For the 251 drug-metabolizing genes and transporter genes, the relative contribution of the variance is shown by three major principal components plotted in three dimensions. dmd.aspetjournals.org their expression of DMETs have also been discussed (Pfeifer et al, 1993;Gómez-Lechón et al, 2003;Wilkening et al, 2003;Knasmüller et al, 2004;Aninat et al, 2006;Donato et al, 2008). All of these in vitro models exhibit advantages and disadvantages.…”

“…However, the expression levels of liver-enriched transcription factors are quite different between primary hepatocytes and hepatic cell lines. For example, most of these transcription factors were found to be weakly expressed in hepatoma cell lines, with the exception of HNF4, which is expressed at a similar level in the hepatoma cell lines and primary hepatocytes (Gómez-Lechón et al, 2003). The fact that P450 enzymes are usually expressed at low levels or are undetectable in hepatoma cells may be largely due to the decreased expression levels of key transcription factors in those cell lines.…”

Preapoptotic cell stress response of primary hepatocytes

“…Expression of cytochromes P450 (CYP) is especially well studied, because of their involvement in drug metabolism. The CYP messenger RNA (mRNA) levels of isolated hepatocytes are similar to those of liver 2 ; however, they decline progressively during the first days in culture. 3 Also, there are significant perturbations of genes encoding for antioxidant enzymes, heat shock proteins, nitric oxide synthase, and methionine adenosyltransferase following the isolation and culture of hepatocytes.…”

“…3 Also, there are significant perturbations of genes encoding for antioxidant enzymes, heat shock proteins, nitric oxide synthase, and methionine adenosyltransferase following the isolation and culture of hepatocytes. 2 As a consequence, the use of primary cultured hepatocytes in drug metabolism studies is confined to the first days in culture. 4 Although the metabolic state of isolated hepatocytes is extensively studied, there are no data on modulation of apoptotic machinery after isolation.…”

The use of bioreactors as in vitro models in pharmaceutical research

“…Hepatocytes are the primary choice for use in BAL systems as they are recognized to be the closest cellular model to the liver [30] [31], and carry out most of the in vivo metabolic processes including the synthesis and excretion of albumin, metabolism of amino acids, urea production and the processing and elimination of drugs and toxins. Phase I and phase II drug metabolising enzymes are crucial in the processing of drugs within the liver, so preservation of these enzymes within a model is essential to reproduce cellular environments in vivo.…”