STEM CELLS IN HEMATOLOGY Mitochondrial DNA spectra of single human CD34 cells, T cells, B cells, and granulocytes

Previously, we described the age-dependent accumulation of mitochon-drial DNA (mtDNA) mutations, leading to a high degree of mtDNA heterogeneity among normal marrow and blood CD34 clones and in granulocytes. We estab-lished a method for sequence analysis of single cells. We show marked, distinct mtDNA heterogeneity from correspond-ing aggregate sequences in isolated cells of 5 healthy adult donors—37.9 % 3.6% heterogeneity in circulating CD34 cells, 36.4 % 14.1 % in T cells, 36.0 % 10.7% in B cells, and 47.7 % 7.4 % in granulo-cytes. Most heterogeneity was caused by poly-C tract variability; however, base substitutions were also prevalent, as fol-lows: 14.7 % 5.7 % in CD34 cells, 15.2 % 9.0 % in T cells, 15.4 % 6.7 % in B cells, and 32.3 % 2.4 % in granulo-cytes. Many poly-C tract length differ-ences and specific point mutations seen in these same donors but assayed 2 years earlier were still present in the new CD34 samples. Additionally, specific poly-C tract differences and point mutations were frequently shared among cells of the lym-phoid and myeloid lineages. Secular sta-bility and lineage sharing of mtDNA se-quence variability suggest that mutations arise in the lymphohematopoietic stem cell compartment and that these changes may be used as a natural genetic marker to estimate the number of active stem cells. (Blood. 2005;106:3271-3284) © 2005 by The American Society of Hematolog