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Blood, 15 June 2004, Vol. 103, No. 12, pp. 4440-4448.
Prepublished online as a Blood First Edition Paper on January 15, 2004; DOI 10.1182/blood-2003-09-3079.
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HEMATOPOIESIS
Elevated telomerase activity and minimal telomere loss in cord blood long-term cultures with extensive stem cell replication
Loretta Gammaitoni,
Katja C. Weisel,
Monica Gunetti,
Kai-Da Wu,
Stefania Bruno,
Silvana Pinelli,
Antonio Bonati,
Massimo Aglietta,
Malcolm A. S. Moore, and
Wanda Piacibello
From the Laboratory of Clinical Oncology, IRCC—Institute for Cancer Research and Treatment, Torino, Italy; James Ewing Laboratory of Developmental Hematopoiesis, Memorial Sloan-Kettering Cancer Center, New York, NY; and Department of Clinical Sciences, Section of Hemato-Oncology, University of Parma Medical School, Parma, Italy.
Telomerase activity, telomere length, stem/progenitor cell production, and function of CD34+ cells from cord blood (CB), bone marrow, and mobilized peripheral blood were evaluated in long-term cultures. CB cells were cultured either on OP-9 stromal cells transduced with an adenovector expressing thrombopoietin (TPO) or stimulated by a cytokine cocktail in the absence of stroma, with, in one method, CD34+ cells reisolated at monthly intervals for passage. Continuous expansion of stem cells as measured by in vitro cobblestone area and secondary colony-forming assays was noted for 18 to 20 weeks and by severe combined immunodeficiency (SCID)-repopulating cells (SRCs), capable of repopulating and serially passage in nonobese diabetic/SCID mice, for 16 weeks. Despite this extensive proliferation, telomere length initially increased and only at late stages of culture was evidence of telomere shortening noted. This telomere stabilization correlated with maintenance of high levels of telomerase activity in the CD34+ cell population for prolonged periods of culture. Cytokine-stimulated cultures of adult CD34+ cells showed CD34+ and SRC expansion (6-fold) for only 3 to 4 weeks with telomere shortening and low levels of telomerase. There is clearly a clinical value for a system that provides extensive stem cell expansion without concomitant telomere erosion. (Blood. 2004;103:4440-4448)
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