Figure 2. GPI-80 expression defines HSPC with self-renewal and engraftment ability.

A. In vitro self-renewal/expansion assay of GPI-80⁺ and GPI-80⁻ HSPC co-cultured on OP9M2 stroma and analyzed at 2 and 4 weeks for the presence of undifferentiated HSPC is shown. Growth of total cells and expansion of CD34⁺CD38^lo/−CD90⁺ HSPC on OP9M2 are shown (n=3 different donor tissues, which are representative examples of 10 independent experiments that showed the same result). Flow cytometry plots of hematopoietic populations after 2 and 4 weeks of co-culture on OP9M2 stroma are shown. Error bars represent mean ± SD. B. Cell cycle analysis of FACS sorted FL HSPC cultured on OP9M2 overnight and pulsed with BRDU is shown (n=3 different donor tissues analyzed in 2 independent experiments). Representative flow cytometry plots assessing BRDU incorporation in GPI-80⁺ and GPI-80⁻ cells are shown. Error bars represent mean ± SEM. C. Scheme of transplant assay for analysis of engraftment ability of GPI-80⁺ and GPI-80⁻ HSPC. Each mouse was transplanted with sorted HSPC subset originating from 50,000 CD34⁺ cells. Flow cytometry analysis of human hematopoietic engraftment in peripheral blood, spleen and BM of mice transplanted with GPI-80⁺ and GPI-80⁻HSPC from human FL is shown (p≤0.01)(n=3 donor tissues assessed in 3 independent experiments). D. Scheme of transplant assay for CFSE labeled HSPC. Flow cytometry analysis of murine bone marrow 16 hours post transplant is shown (n=3 donor tissues assessed in 2 independent experiments). See also Figure S2 and Table S2.