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Cdc42 and aging of human HSC
HSC to undergo first division was considerably longer than cells treated with 5 mM or 10 mM CASIN albeit not statistically significant (P=0.06, Figure 4H) suggesting the inhibition of Cdc42 activity may facilitate or support early division onset.
Aged hematopoietic stem cells show an elevated level of chimerism but balanced differentiation in NBSGW xenotransplants
Murine xenotransplant models that are used to test the potential of human HSC usually require irradiation of the recipient animal for establishing human engraftment.17 Irradiation of the recipients though might contribute to variable secondary effects not linked to the potential of the transplanted HSC. In order to circumvent the necessi- ty for irradiation of the recipients in xenotransplants, we utilized NBSGW animals as recipients33 to assess the func- tion of young, aged and CASIN-treated aged human HSC in vivo.
To this end, 500 HSC were injected via the tail vein into mice and BM chimerism was analyzed at week 8 and 12 post xenotransplantation. Mice were considered to be suc- cessfully engrafted when the level of human hematopoiet- ic cells (huCD45+) detected was higher than 0.1% of total CD45+ cells.34 In general, transplants with human BM-
derived HSC result in a much lower level of chimerism in comparison to transplants with CB-derived HSC (data not shown). Animals transplanted with aged human HSC showed a significantly higher level of engraftment (mean 3.7%) compared to recipients that received young HSC (mean 0.6%) and the aged + CASIN groups (mean 1.6%, 2.1%) (Figure 5A). Chimerism driven by aged HSC further increased at 12 weeks post-transplantation (mean 5%) compared to chimerism driven by young HSC (Figure 5B). This result was surprising, as the small number of previous studies in which aged human cells were transplanted into irradiated recipients demonstrated a lower16 or at least sim- ilar chimerism15 stemming from aged compared to young human HSC. In order to test whether irradiation of recipi- ents might influence the reconstitution potential of aged HSC, we also transplanted young and aged human HSC into irradiated (1.6 Gy) NBSGW mice. Recipients of aged HSC displayed similar levels of chimerism compared to recipients of young HSC, and only marginal differences in their differentiation profile (Online Supplementary Figure S6A to C). Irradiation of recipients might thus influence the reconstitution potential of aged human HSC in xenotrans- plantation experiments. In summary, our results demon- strate that aged HSC show an overall elevated contribution to BM upon transplantation into NBSGW animals.
AB
CD
Figure 5. Analysis of xenotransplantation experiment under non-irradiated settings. Frequency of human cells in the bone marrow (BM) of NBSGW mice at (A) 8 weeks and (B) 12 weeks post-transplant. (C) Emergence of human myeloid cells and (D) B cells post-transplant from young (blue), aged (green) and CASIN-treated aged (orange, brown) -recipients. Red line represents averages of non-transplanted controls. Bars represent the mean "±" standard deviation (SD). n = 6 different donors per cohort, 46 mice at 8 weeks and 42 mice at 12 weeks. Donor age: young =24-35 years (yr), median =27 yr; aged =60-81 yr, median =62 yr.
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