Accelerated telomere shortening in hematological lineages is limited to the first year following stem cell transplantation

Abstract

Using quantitative fluorescence in situ hybridization and flow cytometry, the telomere length of telomere repeat sequences after stem cell transplantation (SCT) were measured. The study included the telomeres of peripheral blood monocytes that should reflect the length of telomeres in stem cells and the telomeres of T lymphocytes that could shorten as a result of peripheral expansion. The loss of telomeres in monocytes and in memory T cells, although accelerated initially, became comparable to the loss of telomeres in healthy controls from the second year after transplantation. In addition, the telomere length in the naive T cells that were produced by the thymus was comparable to the telomere length in the naive T cells IntroductionTelomeres are specialized structures at the end of eukaryotic chromosomes that, in vertebrates, consist of hundreds to thousands of tandem repeats of the sequence TTAGGG. 1 Because telomere length decreases with the number of cell divisions in vitro and with aging in vivo, 2 it reflects the mitotic history of the cells. Furthermore, short telomeres might limit the cells' remaining replicative capacity. 3 This may be of special significance after stem cell transplantation (SCT) because the telomere length of leukocytes in the patient may be up to 2000 base pair (bp) shorter than in the donor. [4][5][6][7][8][9] Because the telomerase activity in stem cells does not prevent telomere shortening, 10 the shorter telomeres may be the consequence of the expansion of the transplanted stem cells needed to repopulate the host. In addition, the telomere loss previously observed in peripheral blood mononuclear cells 4,6,9 might be explained by the extensive proliferation of the T lymphocytes cotransfused with the marrow. In this report we show that both phenomena contribute significantly to the accelerated telomere shortening after SCT and that after an initial period of approximately one year, the rate of telomere shortening in patients becomes comparable to that in their donors. Study designThe average length of telomere repeats in individual cells was measured by flow fluorescence in situ hybridization (FISH), as previously described, 11 and expressed as molecules of equivalent soluble fluorochrome (MESF) units 12 after calibration with fluorescein isothiocyanate (FITC)-labeled fluorescent calibration beads (Quantum-24 Premixed; Flow Cytometry Standards Corporation, San Juan, Puerto Rico).Interexperimental variation was monitored by analyzing aliquots of a batch of frozen cells 13 and did not exceed 15%. Fluorescence-activated cell sorter (FACS) analysis of telomere length in monocytes and lymphocytes was performed with gates on single diploid cells with the characteristic light scatter properties. 13 To analyze naive T cells and cells with a memory phenotype, 2-5 ϫ 10 4 cells were fractionated into CD4 ϩ CD45RA ϩ CD45RO Ϫ and CD4 ϩ CD45RA Ϫ CD45RO ϩ by FACS analysis (FACStar Plus; Becton Dickinson, Basel, Switzerland) after staining with anti-CD4-APC (allophycocyanin) (Leu4; Becton Dickinso...

Measurable immune dysfunction and telomere attrition in long-term allogeneic transplant recipients

“…44 Several studies have shown that this accelerated telomere loss occurs within the first year post transplant with subsequent telomere loss continuing at a normal rate. 19,25,26 Most recently, it has been suggested that telomere homeostasis, rather than shortening, may occur during the first year after transplantation in patients who receive G-CSF-mobilized peripheral blood stem cells rather than bone marrow, 45 although this was not supported in the few patients in our study who received PBSCs. Given these findings, accelerated loss is most likely a result of the rapid cell division the hematopoietic system must undergo in the first year in order to reconstitute the bone marrow compartment after marrow ablation, and this loss may be attenuated by increasing the numbers of hematopoietic cells infused.…”

“…19 Furthermore, this accelerated loss is limited to the first year post transplant, with subsequent telomere shortening occurring at a normal rate, supporting the hypothesis that this loss is a result of extra cell division. 19,25,26 Despite increased hematopoietic demands early after transplantation, normal peripheral blood counts are maintained in long-term allogeneic recipients, even though their telomere lengths remain shorter than their respective donors. 27 The functional significance of prematurely shortened telomeres has yet to be determined and is controversial.…”

Concepts of human leukemic development

“…Unlike most normal human somatic cells, hematopoietic stem and progenitor cells exhibit some telomerase activity (Broccoli et al, 1995;Engelhardt et al, 1997). Nevertheless, telomeres in normal hematopoietic cells shorten with age (Rufer et al, 1999) and following transplantation (Rufer et al, 2001), suggesting that the telomerase activity in progenitor cells is not sufficient to completely prevent telomere shortening with proliferation. Given the greatly increased proliferative capacity of LSC, one of the essential elements of leukemogenesis is the acquisition of a mechanism to maintain telomere function and thereby provide limitless replicative potential (Hanahan and Weinberg, 2000).…”

X‐chromosome inactivation and telomere size in newborns resulting from intracytoplasmic sperm injection

“…In addition 19 cord blood samples from male ICSI cases were analyzed. Only newborn cord blood samples were used as controls since telomere length declines rapidly in the first year of life [Rufer et al, 2001]. An estimation of average telomere length was obtained using the Telo TAGGG Telomere Length Assay kit from Roche Diagnostics (Montreal, CA).…”