Telomerase reconstitution immortalizes human fetal hepatocytes without disrupting their differentiation potential

Henning Wege, Hai T. Le, Michael S. Chui, Li Liu, Jian Wu, Ranjit Giri, Harmeet Malhi, Baljit S. Sappal, Vinay Kumaran, Sanjeev Gupta, Mark A. Zern

Research output: Contribution to journalArticle

146 Citations (Scopus)

Abstract

Background & Aims: The availability of in vitro expandable human hepatocytes would greatly advance liver-directed cell therapies. Therefore, we examined whether human fetal hepatocytes are amenable to telomerase-mediated immortalization without inducing a transformed phenotype and disrupting their differentiation potential. Telomerase is a ribonucleoprotein that plays a pivotal role in maintaining telomere length and chromosome stability. Human somatic cells, including hepatocytes, exhibit no telomerase activity. Consequently, their telomeres progressively shorten with each cell cycle until critically short telomeres trigger replicative senescence. Methods: The catalytic subunit, telomerase reverse transcriptase, was expressed in human fetal hepatocytes. Transduced cells were characterized for telomerase activity, telomere length, proliferative capacity, hepatocellular functions, oncogenicity, and their in vivo maturation potential. Results: The expression of human telomerase reverse transcriptase restored telomerase activity in human fetal hepatocytes. Telomerase-reconstituted cells were capable of preserving elongated telomeres, propagated in culture beyond replicative senescence for more than 300 cell doublings (to date), and maintained their liver-specific nature, as analyzed by a panel of hepatic growth factors, growth factor receptors, and transcription factors as well as albumin, glucose-6-phosphatase, glycogen synthesis, cytochrome P450 (CYP) expression profiles, and urea production. Moreover, the immortalized cells exhibited no oncogenicity, and no upregulation of c-Myc was detected. The cells engrafted and survived in the liver of immunodeficient mice with hepatocellular gene expression. Conclusions: Reconstitution of telomerase activity induces indefinite replication in human fetal hepatocytes and offers unique opportunities for examining basic biologic mechanisms and for considering development of stable cell lines for liver-directed therapies.

Original languageEnglish (US)
Pages (from-to)432-444
Number of pages13
JournalGastroenterology
Volume124
Issue number2
DOIs
StatePublished - Feb 1 2003

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Telomerase
Hepatocytes
Telomere
Liver
Cell Aging
Telomere Shortening
Glucose-6-Phosphatase
Chromosomal Instability
Ribonucleoproteins
Growth Factor Receptors
Cell- and Tissue-Based Therapy
Glycogen
Human Activities
Cytochrome P-450 Enzyme System
Urea
Albumins
Intercellular Signaling Peptides and Proteins
Cell Cycle
Transcription Factors
Up-Regulation

ASJC Scopus subject areas

  • Gastroenterology

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Telomerase reconstitution immortalizes human fetal hepatocytes without disrupting their differentiation potential. / Wege, Henning; Le, Hai T.; Chui, Michael S.; Liu, Li; Wu, Jian; Giri, Ranjit; Malhi, Harmeet; Sappal, Baljit S.; Kumaran, Vinay; Gupta, Sanjeev; Zern, Mark A.

In: Gastroenterology, Vol. 124, No. 2, 01.02.2003, p. 432-444.

Research output: Contribution to journalArticle

Wege, H, Le, HT, Chui, MS, Liu, L, Wu, J, Giri, R, Malhi, H, Sappal, BS, Kumaran, V, Gupta, S & Zern, MA 2003, 'Telomerase reconstitution immortalizes human fetal hepatocytes without disrupting their differentiation potential', Gastroenterology, vol. 124, no. 2, pp. 432-444. https://doi.org/10.1053/gast.2003.50064
Wege, Henning ; Le, Hai T. ; Chui, Michael S. ; Liu, Li ; Wu, Jian ; Giri, Ranjit ; Malhi, Harmeet ; Sappal, Baljit S. ; Kumaran, Vinay ; Gupta, Sanjeev ; Zern, Mark A. / Telomerase reconstitution immortalizes human fetal hepatocytes without disrupting their differentiation potential. In: Gastroenterology. 2003 ; Vol. 124, No. 2. pp. 432-444.
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abstract = "Background & Aims: The availability of in vitro expandable human hepatocytes would greatly advance liver-directed cell therapies. Therefore, we examined whether human fetal hepatocytes are amenable to telomerase-mediated immortalization without inducing a transformed phenotype and disrupting their differentiation potential. Telomerase is a ribonucleoprotein that plays a pivotal role in maintaining telomere length and chromosome stability. Human somatic cells, including hepatocytes, exhibit no telomerase activity. Consequently, their telomeres progressively shorten with each cell cycle until critically short telomeres trigger replicative senescence. Methods: The catalytic subunit, telomerase reverse transcriptase, was expressed in human fetal hepatocytes. Transduced cells were characterized for telomerase activity, telomere length, proliferative capacity, hepatocellular functions, oncogenicity, and their in vivo maturation potential. Results: The expression of human telomerase reverse transcriptase restored telomerase activity in human fetal hepatocytes. Telomerase-reconstituted cells were capable of preserving elongated telomeres, propagated in culture beyond replicative senescence for more than 300 cell doublings (to date), and maintained their liver-specific nature, as analyzed by a panel of hepatic growth factors, growth factor receptors, and transcription factors as well as albumin, glucose-6-phosphatase, glycogen synthesis, cytochrome P450 (CYP) expression profiles, and urea production. Moreover, the immortalized cells exhibited no oncogenicity, and no upregulation of c-Myc was detected. The cells engrafted and survived in the liver of immunodeficient mice with hepatocellular gene expression. Conclusions: Reconstitution of telomerase activity induces indefinite replication in human fetal hepatocytes and offers unique opportunities for examining basic biologic mechanisms and for considering development of stable cell lines for liver-directed therapies.",
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AU - Wege, Henning

AU - Le, Hai T.

AU - Chui, Michael S.

AU - Liu, Li

AU - Wu, Jian

AU - Giri, Ranjit

AU - Malhi, Harmeet

AU - Sappal, Baljit S.

AU - Kumaran, Vinay

AU - Gupta, Sanjeev

AU - Zern, Mark A.

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N2 - Background & Aims: The availability of in vitro expandable human hepatocytes would greatly advance liver-directed cell therapies. Therefore, we examined whether human fetal hepatocytes are amenable to telomerase-mediated immortalization without inducing a transformed phenotype and disrupting their differentiation potential. Telomerase is a ribonucleoprotein that plays a pivotal role in maintaining telomere length and chromosome stability. Human somatic cells, including hepatocytes, exhibit no telomerase activity. Consequently, their telomeres progressively shorten with each cell cycle until critically short telomeres trigger replicative senescence. Methods: The catalytic subunit, telomerase reverse transcriptase, was expressed in human fetal hepatocytes. Transduced cells were characterized for telomerase activity, telomere length, proliferative capacity, hepatocellular functions, oncogenicity, and their in vivo maturation potential. Results: The expression of human telomerase reverse transcriptase restored telomerase activity in human fetal hepatocytes. Telomerase-reconstituted cells were capable of preserving elongated telomeres, propagated in culture beyond replicative senescence for more than 300 cell doublings (to date), and maintained their liver-specific nature, as analyzed by a panel of hepatic growth factors, growth factor receptors, and transcription factors as well as albumin, glucose-6-phosphatase, glycogen synthesis, cytochrome P450 (CYP) expression profiles, and urea production. Moreover, the immortalized cells exhibited no oncogenicity, and no upregulation of c-Myc was detected. The cells engrafted and survived in the liver of immunodeficient mice with hepatocellular gene expression. Conclusions: Reconstitution of telomerase activity induces indefinite replication in human fetal hepatocytes and offers unique opportunities for examining basic biologic mechanisms and for considering development of stable cell lines for liver-directed therapies.

AB - Background & Aims: The availability of in vitro expandable human hepatocytes would greatly advance liver-directed cell therapies. Therefore, we examined whether human fetal hepatocytes are amenable to telomerase-mediated immortalization without inducing a transformed phenotype and disrupting their differentiation potential. Telomerase is a ribonucleoprotein that plays a pivotal role in maintaining telomere length and chromosome stability. Human somatic cells, including hepatocytes, exhibit no telomerase activity. Consequently, their telomeres progressively shorten with each cell cycle until critically short telomeres trigger replicative senescence. Methods: The catalytic subunit, telomerase reverse transcriptase, was expressed in human fetal hepatocytes. Transduced cells were characterized for telomerase activity, telomere length, proliferative capacity, hepatocellular functions, oncogenicity, and their in vivo maturation potential. Results: The expression of human telomerase reverse transcriptase restored telomerase activity in human fetal hepatocytes. Telomerase-reconstituted cells were capable of preserving elongated telomeres, propagated in culture beyond replicative senescence for more than 300 cell doublings (to date), and maintained their liver-specific nature, as analyzed by a panel of hepatic growth factors, growth factor receptors, and transcription factors as well as albumin, glucose-6-phosphatase, glycogen synthesis, cytochrome P450 (CYP) expression profiles, and urea production. Moreover, the immortalized cells exhibited no oncogenicity, and no upregulation of c-Myc was detected. The cells engrafted and survived in the liver of immunodeficient mice with hepatocellular gene expression. Conclusions: Reconstitution of telomerase activity induces indefinite replication in human fetal hepatocytes and offers unique opportunities for examining basic biologic mechanisms and for considering development of stable cell lines for liver-directed therapies.

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