Reconstitution of telomerase activity combined with HPV-E7 expression allow human preadipocytes to preserve their differentiation capacity after immortalization

doi:doi:10.1038/sj.cdd.4401273

Cell Death and Differentiation (2003) 10, 1025–1031. doi:10.1038/sj.cdd.4401273

Reconstitution of telomerase activity combined with HPV-E7 expression allow human preadipocytes to preserve their differentiation capacity after immortalization

C Darimont¹, I Zbinden¹, O Avanti¹, P Leone-Vautravers¹, V Giusti², P Burckhardt², A M A Pfeifer¹ and K Macé

¹Nestlé Research Center, PO Box 44, Vers-Chez-Les-Blanc, 1000 Lausanne 26, Switzerland
²Département de Médecine, CHUV, Rue du Bugnon 46, 1011 Lausanne, Switzerland

Correspondence: C Darimont, Tel.: +41 21 785 84 02; Fax: +41 21 785 85 44; E-mail: christian.darimont@rdls.nestle.com

Received 25 November 2002; Revised 11 April 2003; Accepted 17 April 2003.

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Abstract

Overexpression of SV40 T-antigen (SV40 T-Ag) has been widely used to overcome replicative senescence of human primary cells and to promote cell immortalization. However, in the case of certain cell types, such as preadipocytes, the differentiation process of immortalized cells is blocked by SV40 T-Ag expression. In this study, human telomerase reverse transcriptase (hTERT) and papillomavirus E7 oncoprotein (HPV-E7) genes were coexpressed in human preadipocytes to test whether this combination could maintain cell differentiation capacity after immortalization. We demonstrated that the HPV-E7/hTERT expressing preadipocytes displayed an indefinite life span. Interestingly, immortalized cells were diploid and presented no chromosomic alterations. These immortalized cells were able to accumulate and hydrolyze intracellular triglycerides and to express adipocyte markers. These data demonstrate, for the first time, that coexpression of hTERT and HPV-E7 in human preadipocytes allows cells not only to display an indefinite life span but also to retain their capacity to differentiate.

Keywords:

oncogenes, adipogenesis, proliferation

Abbreviations:

aP2, adipocyte fatty acid binding protein; BSA, bovine serum albumin; C/EBP, CCAAT/enhancer binding protein alpha; Dex, dexamethasone; FBS, fetal bovine serum; GPDH, glycerol-3-phosphate dehydrogenase; hTERT, human telomerase reverse transcriptase; HPV-E7, human papillomavirus E7 oncoprotein; IBMX, isobutyl methyl xanthine; LPL, lipoprotein lipase; OA, oleic acid; PPAR, peroxisome proliferator-activated receptor gamma; RPLp0, ribosomal protein large P0; SV40 T-Ag, SV40 T-antigen