Retrovirally mediated telomerase immortalization of neural progenitor cells.
Traditional methods of generating immortalized lines of both somatic cells and their progenitors have relied on the use of oncogenes. However, the resulting lines are typically anaplastic in vitro and tumorigenic in vivo, and hence of limited utility. The overexpression of telomerase, as mediated by the induced overexpression of human telomerase reverse transcriptase (hTERT), has permitted the generation of stable, non-oncogenic lines of a variety of cell types. This strategy for immortalization has found special utility in the central nervous system, as few stable lines are available for the study of either human neural progenitor cells, or of neurons or glia of restricted phenotype. We describe the use of retroviral hTERT overexpression for generating lines of immortalized human neural progenitor cells, whose neuronal progeny are phenotypically restricted, post-mitotic and functionally competent. Although we focus here on telomerase immortalization of spinal neural progenitors, this is a broadly applicable protocol for using hTERT to immortalize human fetal neural progenitors of any pre-selected phenotype and for characterizing the cell lines thereby generated.
Roy
Neeta S
NS
Department of Neurology, Weill Medical College of Cornell University, New York, New York 10021, USA. ner2004@med.cornell.edu
Chandler-Militello
Devin
D
Lu
Gang
G
Wang
Su
S
Goldman
Steven A
SA
D018929:Cell Culture Techniques; D003470:Culture Media; D053686:Fetal Stem Cells / Q000166:cytology*; D006801:Humans; D007621:Karyotyping; D009474:Neurons / Q000166:cytology*; D011691:Puromycin; D012190:Retroviridae / Q000235:genetics*; D013116:Spinal Cord / Q000166:cytology / Q000196:embryology; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism; D016615:Telomere / Q000502:physiology; D014162:Transfection
D003470:Culture Media; D011691:Puromycin; C509186:TERT protein, human; D019098:Telomerase