Evidence that the number of hematopoietic stem cells per animal is conserved in mammals

Abstract

Humans and larger mammals require more blood cells per lifetime than mice because of their larger size and longer life expectancy. To investigate this evolutionary adaptation, we calculated the total number of nucleated marrow cells (NMCs) per cat, observing the distribution of 59 Fe to marrow, then multiplied this value (1.9 ؎ 0.9 ؋ 10 10 [mean ؎ SD]) times the frequency of feline hematopoietic stem cells (HSCs) (6 HSCs/10 7 NMCs) to derive the total number of HSCs per cat (11 400 ؎ 5400). Surprisingly, when the total number of HSCs per mouse was calculated with a similar experimental and computational approach, the value was equivalent. These data imply that the output of differentiated cells per feline HSC must vastly exceed that of murine HSCs. IntroductionHematopoietic stem cells (HSCs) are the functional units of blood cell production. Through self-renewal (replication) and differentiation, HSCs maintain hematopoiesis throughout an animal's lifetime. HSC number and kinetics have been studied extensively in mice and are genetically regulated. Less is known about HSC behavior in larger mammals or humans, in whom hematopoietic demand (ie, number of mature cells required) is significantly higher. To understand the adaptation to increased size and longevity, we have previously investigated HSC behavior in cats (which make the same number of red cells in 8 days as humans in 1 day or mice in a 2-year lifetime) 1 and have demonstrated that the frequencies of HSCs differ by 100-fold in mouse and cat marrow. [2][3][4][5][6][7][8] In this study, we determined the total number of HSCs. Surprisingly, it appears that the total number of HSCs in a cat is similar to, and within-estimation variability overlaps with, the total number of HSCs in a mouse. Study designTo compute the total number of HSCs in cats, HSC frequency (6 HSCs/10 7 nucleated marrow cells [NMCs] in adolescent cats, as determined by limiting-dilution competitive transplantation 6 ) was multiplied by the total number of NMCs. Because the number of NMCs per cat was unknown, the value was derived by observing the distribution of iron Fe 59 ( 59 Fe)-transferrin to the erythroid marrow of 4-to 9-month-old cats, using the methods of previous murine 7 and human 8 studies. In this way, a direct comparison of the results between species was feasible.In a preliminary study, we confirmed that 59 Fe had a t 1 ⁄2 in the circulation of 39 minutes. 9 We demonstrated that infused 59 Fe was undetectable in blood by 16 hours (which is 25 t 1 ⁄2 s), and we assumed complete redistribution to marrow at this time. Heparinized plasma (1.5-2.5 mL) was incubated with 3 Ci (0.111 MBq) 59 Fe as ferric chloride at 37°C for 30 minutes to allow transferrin saturation 9 and then was infused into cats intravenously. Studies were performed in 8 adolescent cats (4-to 10-monthold cats were used in studies to determine the HSC frequency 6 ). Marrow and blood were collected at baseline, 2 minutes after infusion, and at 16 to 23 hours. The number of NMCs and radioactivity (Auto-Gamma Coun...

In vivo transduction of primitive mobilized hematopoietic stem cells after intravenous injection of integrating adenovirus vectors

“…Total BM MNCs of animals injected with both vectors showed GFP in 2.5%, 1.1%, 0.4%, and 0.9% of cells at 4,8,12, and 20 weeks after transduction. The loss of transduced cells over time was most likely due to exit or turnover of short-term reconstituting HSPCs and differentiated cells.…”

“…First, the ability of GFP 1 lineage-depleted BM cells harvested at week 4,8,12, and 20 after HD-Ad injection to form multilineage progenitor colonies (or CFUs) was analyzed ( Figure 3A). The percentage of GFP 1 CFUs increased during the first 12 weeks in treated animals, suggesting enrichment of transduced colony-forming cells and the loss of transduced less primitive cells ( Figure 3B).…”

“…To evaluate stable long-term transgene expression after HSPC transduction, groups of mice were injected with both vectors, or HDAd-GFP alone and euthanized 4,8,12, and 20 weeks after transduction ( Figure 2D). Total BM MNCs of animals injected with both vectors showed GFP in 2.5%, 1.1%, 0.4%, and 0.9% of cells at 4,8,12, and 20 weeks after transduction.…”

“…3 A problem for in vivo HSPC transduction is the low number of HSPCs in the BM. Only about 1 in 10 8 nucleated marrow cells are HSPCs 4 ; the vast majority of cells in the BM are blood cell progenitor cells with different levels of lineage commitment. A long-term therapeutic effect of gene therapy requires that gene transfer vectors target these rare HSPCs.…”

Blood cell progenitors: Insights into the properties of stem cells

“…The frequency of human and murine hematopoietic stem cells has been estimated to be 1/10 4 -10 5 bone marrow cells (Harrison et al, 1989;Sutherland et al, 1989;Harrison, 1993;Szilvassy and Hoffman, 1995;Bhatia et al, 1998;Abkowitz et al, 2002). The rarity of this population presented considerable impediments to isolation until recently.…”