title: Establishment of a human neonatal hepatocyte cell line.
abstract: Hepatocytes are routinely used to generate and identify drug metabolites and hepatic toxicity. Primary cultures of human hepatocytes are the model cell of choice for most of these pharmacological and toxicological studies. However, major problems are encountered with primary liver cell cultures: the dwindling availability of viable livers, hepatocytes having a limited life span, the loss of liver-specific functions in culture, and the donor to donor variability. These limitations have created a significant need for an in vitro hepatocyte system, which has both the potential for use in toxicological and pharmaceutical studies as well as clinical applications. Ectopic expression of human telomerase reverse transcriptase (hTERT) is one of the major strategies used to develop immortalized cells. Immortalization of primary cells using hTERT allows retention of the original cellular characteristics and functions and avoids some of the genetic and phenotypic instabilities associated with using known oncogenes. In the present study, we developed a cell line from human neonatal hepatocytes by transduction with a recombinant retrovirus expressing the hTERT gene. Induction of stable expression of hTERT in the neonatal cells led to immortalization of these cells. The cell line was cultured continuously for more than 25 passages, equivalent to >25 population doublings, whereas the parental cells senesced within five passages. Analysis of telomerase activity as measured by telomeric repeat amplification protocol assay indicated elevated levels of telomerase activity in immortalized cells compared to the parental cells. These immortalized human hepatocytes cells maintained a normal diploid karyotype as well as the gene expression profile similar to that of human normal neonatal hepatocytes. The data suggest that these immortalized cells preserved some of the biological characteristics of hepatic progenitor cells and might be useful as an in vitro model for pharmacological and toxicity studies.
authors: Cell Biology, American Type Culture Collection, Manassas, VA 20110-2209, USA. yreid@atcc.org
authors: Yvonne
authors: Y
authors: Reid
authors: Jaya P
authors: JP
authors: Gaddipati
authors: Deepmala
authors: D
authors: Yadav
authors: Judy
authors: J
authors: Kantor
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D002461:Cell Line, Transformed; D049109:Cell Proliferation; D048430:Cell Shape; D004247:DNA / Q000032:analysis; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D005838:Genotype; D022781:Hepatocytes / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007231:Infant, Newborn; D007621:Karyotyping; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D019098:Telomerase / Q000378:metabolism
chemical_list: D015415:Biomarkers; D012333:RNA, Messenger; D004247:DNA; C509186:TERT protein, human; D019098:Telomerase