Telomerized human bone marrow-derived cell clones maintain the phenotype of hematopoietic-supporting osteoblastic and myofibroblastic stromal cells after long-term culture.
OBJECTIVE
Gene transfer of the telomerase catalytic subunit (TERT) into primary human stromal cells prolonged their lifespan. However, primary human stromal cells are actually composed of adipocytes, myofibroblasts, osteoblasts, etc. Our objective was to investigate the phenotype and hematopoietic-support of the human telomerized stromal cell (HTS) in clonal level.


METHODS
We established HTS clones (HTS-1 to HTS-9) from a parental population of HTSs by limiting dilution. Hematopoietic-supporting activity of the HTS clones was examined by coculturing with CD34(+) cells.


RESULTS
HTS-1 to HTS-3 contained alkaline phosphatase (ALP)(+) cells, and HTS-4 to HTS-9 were composed of both ALP(+) and alpha-smooth muscle actin-positive cells. Although all HTS clones exhibited normal growth kinetics, one of the HTS clones exhibited a chromosomal abnormality. Moreover, the parental population of the HTS cells acquired an increased growth rate and anchorage independence after 4 years of culturing. Expression of hematopoietic growth factors, such as stem cell factor, angiopoietin-1, and hedgehog mRNA was detected in all HTS clones. The degree of hematopoietic progenitor support differed between the HTS clones, and the expansion level of CD34(+) cells was the highest in HTS-8.


CONCLUSIONS
Human telomerized stromal cell clones exhibited the phenotype of hematopoietic supporting osteoblastic and myofibroblastic cells after long-term culture. Clinical application of HTS cells should be limited because of their potential for neoplastic transformation after hTERT gene transfer. HTS cells may be useful for analyzing the molecular mechanism of hematopoietic support of human stromal cells.
Kobune
Masayoshi
M
Fourth Department of Internal Medicine, Sapporo Medical University School of Medicine, Japan. mkobune@sapmed.ac.jp
Kato
Junji
J
Chiba
Hiroki
H
Kawano
Yutaka
Y
Tanaka
Maki
M
Takimoto
Rishu
R
Hamada
Hirofumi
H
Niitsu
Yoshiro
Y
D001854:Bone Marrow Cells; D019070:Cell Lineage; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002999:Clone Cells; D018920:Coculture Techniques; D005347:Fibroblasts / Q000166:cytology*; D006410:Hematopoiesis; D016298:Hematopoietic Cell Growth Factors / Q000032:analysis / Q000235:genetics; D006412:Hematopoietic Stem Cells / Q000166:cytology; D006801:Humans; D016130:Immunophenotyping; D010006:Osteoblasts / Q000166:cytology*; D012333:RNA, Messenger / Q000032:analysis; D017154:Stromal Cells / Q000166:cytology* / Q000378:metabolism; D016615:Telomere / Q000235:genetics*; D013997:Time Factors; D014161:Transduction, Genetic
D016298:Hematopoietic Cell Growth Factors; D012333:RNA, Messenger