Lentivector-mediated transfer of Bmi-1 and telomerase in muscle satellite cells yields a duchenne myoblast cell line with long-term genotypic and phenotypic stability.
Conditionally immortalized human cells are valuable substrates for basic biologic studies, as well as for the production of specific proteins and for the creation of bioartificial organs. We previously demonstrated that the lentivector-mediated transduction of immortalizing genes into human primary cells is an efficient method for obtaining such cell lines. Here, we used human muscle satellite cells as model targets to examine the impact of the transduced genes on the genotypic and phenotypic characteristics of the immortalized cells. The most commonly used immortalizing gene, the SV40 large T antigen (T-Ag), was extremely efficient at inducing the continuous growth of primary myoblasts, but the resulting cells rapidly accumulated major chromosomal aberrations and exhibited profound phenotypic changes. In contrast, the constitutive expression of telomerase and Bmi-1 in satellite cells from a control individual and from a patient suffering from Duchenne's muscular dystrophy yielded cell lines that remained diploid and conserved their growth factor dependence for proliferation. However, despite the absence of detectable cytogenetic abnormalities, clones derived from satellite cells of a control individual exhibited a differentiation block in vitro. In contrast, a Duchenne-derived cell line exhibited all the phenotypic characteristics of its primary parent, including an ability to differentiate fully into myotubes when placed in proper culture conditions. This cell line should constitute a useful reagent for a wide range of studies aimed at this disease.
Department of Genetics and Microbiology, University of Geneva, 1 rue Michel-Servet, 1211 Geneva 4, Switzerland.
Christophe
C
Cudré-Mauroux
Teresa
T
Occhiodoro
Stéphane
S
König
Patrick
P
Salmon
Laurent
L
Bernheim
Didier
D
Trono
D000293:Adolescent; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics*; D002454:Cell Differentiation; D002455:Cell Division; D002460:Cell Line; D017690:Cell Transplantation; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D007621:Karyotyping; D016086:Lentivirus / Q000235:genetics*; D008297:Male; D020388:Muscular Dystrophy, Duchenne / Q000473:pathology; D032446:Myoblasts / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D063150:Polycomb Repressive Complex 1; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000235:genetics* / Q000378:metabolism; D032496:Satellite Cells, Skeletal Muscle / Q000378:metabolism*; D019098:Telomerase / Q000235:genetics* / Q000378:metabolism
D000952:Antigens, Polyomavirus Transforming; C069077:BMI1 protein, human; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D012097:Repressor Proteins; D063150:Polycomb Repressive Complex 1; D019098:Telomerase