Figure 4
Figure 4. Hematopoietic stem cells are not impaired in KRNxG7 mice. (A) Immunophenotypic analyses of HSC containing populations. BM from 3-, 6-, and 7-week-old KRN, B6xG7, and KRNxG7 mice were subjected to FACS analyses for cKit, Sca1, and lineage or CD150, CD48, and CD41 markers. The frequency ± SD of KSL or SLAM is shown on the y-axis (n ≥ 4). (B) Cell-cycle analysis of BM subpopulations. Six- to 8-week-old mice were injected with a single dose of BrdU proportionate to body mass for 2 to 3 hours before death. Different BM fractions were analyzed for BrdU incorporation by flow cytometry. Shown is the mean ± SD of BrdU+ cells for each population for 2 independent experiments (n = 4 or 5 total mice). (C) Quiescent fraction analysis of BM KSL cells. BM cells from KRNxG7 and B6xG7 mice (n = 4 or 5) of different ages were lineage depleted, surface stained for c-Kit and Sca-1, and subjected to intracellular staining for Ki-67 and Hoechst (supplemental Figure 5). Quiescent cells do not express Ki-67 (Ki-67−) and stain low for Hoechst because of their 2N DNA content (vs 4N DNA content of S/G2/M phase cells). Ki-67−Hoechstlow (quiescent cell) fraction of the stem cell–enriched Lin−c-Kit+Sca1+ (KSL) population and the non–stem cell–enriched Lin−cKit+Sca1− population are shown. As expected, KSL cells are more quiescent than Lin−cKit+Sca1− progenitors. However, there is no appreciable difference in quiescent fraction in KRNxG7 KSL cells compared with B6xG7 KSL cells. (D) B6xG7 (CD45.2xCD45.2) or KRNxG7 (CD45.2xCD45.2) BM was transplanted into lethally irradiated B6xG7 (CD45.1xCD45.2) recipients. PB was analyzed for donor contribution (CD45.2) every 6 weeks after BM transplantation for 6 months. The percentage ± SD of CD45.2+ chimerism is shown on the y-axis (n = 4/genotype). (E) B6xG7 (CD45.2+) or KRNxG7 (CD45.2+) BM cells (2 × 105) were mixed with B6xG7 (CD45.1+; CD45.2+) competitor BM cells (2 × 105) and injected intravenously into lethally irradiated (1000 cGy) B6xG7 (CD45.1xCD45.2) recipient mice. PB CD45.2+ cells of were analyzed 3 months and 5 months after transplantation. Data represent the average percentages of PB chimerism ± SEM. *P < .05. **P < .01.

Hematopoietic stem cells are not impaired in KRNxG7 mice. (A) Immunophenotypic analyses of HSC containing populations. BM from 3-, 6-, and 7-week-old KRN, B6xG7, and KRNxG7 mice were subjected to FACS analyses for cKit, Sca1, and lineage or CD150, CD48, and CD41 markers. The frequency ± SD of KSL or SLAM is shown on the y-axis (n ≥ 4). (B) Cell-cycle analysis of BM subpopulations. Six- to 8-week-old mice were injected with a single dose of BrdU proportionate to body mass for 2 to 3 hours before death. Different BM fractions were analyzed for BrdU incorporation by flow cytometry. Shown is the mean ± SD of BrdU+ cells for each population for 2 independent experiments (n = 4 or 5 total mice). (C) Quiescent fraction analysis of BM KSL cells. BM cells from KRNxG7 and B6xG7 mice (n = 4 or 5) of different ages were lineage depleted, surface stained for c-Kit and Sca-1, and subjected to intracellular staining for Ki-67 and Hoechst (supplemental Figure 5). Quiescent cells do not express Ki-67 (Ki-67−) and stain low for Hoechst because of their 2N DNA content (vs 4N DNA content of S/G2/M phase cells). Ki-67−Hoechstlow (quiescent cell) fraction of the stem cell–enriched Lin−c-Kit+Sca1+ (KSL) population and the non–stem cell–enriched Lin−cKit+Sca1− population are shown. As expected, KSL cells are more quiescent than Lin−cKit+Sca1− progenitors. However, there is no appreciable difference in quiescent fraction in KRNxG7 KSL cells compared with B6xG7 KSL cells. (D) B6xG7 (CD45.2xCD45.2) or KRNxG7 (CD45.2xCD45.2) BM was transplanted into lethally irradiated B6xG7 (CD45.1xCD45.2) recipients. PB was analyzed for donor contribution (CD45.2) every 6 weeks after BM transplantation for 6 months. The percentage ± SD of CD45.2+ chimerism is shown on the y-axis (n = 4/genotype). (E) B6xG7 (CD45.2+) or KRNxG7 (CD45.2+) BM cells (2 × 105) were mixed with B6xG7 (CD45.1+; CD45.2+) competitor BM cells (2 × 105) and injected intravenously into lethally irradiated (1000 cGy) B6xG7 (CD45.1xCD45.2) recipient mice. PB CD45.2+ cells of were analyzed 3 months and 5 months after transplantation. Data represent the average percentages of PB chimerism ± SEM. *P < .05. **P < .01.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal