![Figure 1. Endogenous NrasG12D/+ induces increased proliferation and self-renewal in HSCs. (A-F) Control and NrasG12D/+ mice were treated with pI-pC and euthanized at various time points for analysis. We refer to the day of the first pI-pC injection as day 1. (A) Lin− CD41− CD48− c-Kit+ Sca-1+ CD150+ HSCs, (B) Lin− CD41− CD48− c-Kit+ Sca-1+ CD150− MPPs, and (C) LSK cells were quantified using flow cytometry. SP, spleen; H.L., hind limb, including tibia and femur; TBM, total bone marrow, ∼fourfold H.L. BM.18 (D) Cell cycle analysis of HSCs and WBM from control and NrasG12D/+ mice using Ki67/DAPI [4,6 diamidino-2-phenylindole] staining. (E) A 16-hour pulse of 5-ethynyl-2′-deoxyuridine (EdU) to quantify proliferating HSCs and WBM. (F) Twenty HSCs purified from control or NrasG12D/+ mice were transplanted with 2 × 105 congeneic BM cells into lethally irradiated mice. Donor-derived blood cells were regularly analyzed in the peripheral blood of recipients. Donor-derived HSCs were quantified in recipients 16 weeks after transplantation. (G) 2 × 106 BM cells isolated from primary recipients were transplanted into lethally irradiated secondary recipient mice. Donor-derived blood cells were regularly analyzed in the peripheral blood of secondary recipients. Data are presented as mean + standard deviation. *P < .05; **P < .01; ***P < .001.](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/121/26/10.1182_blood-2012-12-475863/4/5203f1.jpeg?Expires=1743231506&Signature=JPQ80biNe5UQRAQnUxGrcLj~ZfnAt1sOPlW0AAe8auEp93fNz4mLhGCWkyVYsEk1yV458Wcx5M41wuelvQ5-chT3vjSEOY978F19QQHFn1GmvbgIEOTGvp59weYeqWMbCE3WXDKKYMWLfJ9ol~IYwiZ00vIwMDZVWwa5~gzgfc~K4mvxcwB05vDebe9qJkQWhqq8~c3yG77q6wsVs8zlPfDExaf8Qf1lTUlb1PeIms2DSq0e~br71ZoCm7HV3bnrHJC8YP2sLfn19ptAt81-PUd8Ol5msp8ujzSgINglSSidfNg3j5Ee07vLWeAa0Opg8zmVOsbt~u7UOaLmp~zXXQ__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Endogenous NrasG12D/+induces increased proliferation and self-renewal in HSCs. (A-F) Control and NrasG12D/+ mice were treated with pI-pC and euthanized at various time points for analysis. We refer to the day of the first pI-pC injection as day 1. (A) Lin− CD41− CD48− c-Kit+ Sca-1+ CD150+ HSCs, (B) Lin− CD41− CD48− c-Kit+ Sca-1+ CD150− MPPs, and (C) LSK cells were quantified using flow cytometry. SP, spleen; H.L., hind limb, including tibia and femur; TBM, total bone marrow, ∼fourfold H.L. BM.18 (D) Cell cycle analysis of HSCs and WBM from control and NrasG12D/+ mice using Ki67/DAPI [4,6 diamidino-2-phenylindole] staining. (E) A 16-hour pulse of 5-ethynyl-2′-deoxyuridine (EdU) to quantify proliferating HSCs and WBM. (F) Twenty HSCs purified from control or NrasG12D/+ mice were transplanted with 2 × 105 congeneic BM cells into lethally irradiated mice. Donor-derived blood cells were regularly analyzed in the peripheral blood of recipients. Donor-derived HSCs were quantified in recipients 16 weeks after transplantation. (G) 2 × 106 BM cells isolated from primary recipients were transplanted into lethally irradiated secondary recipient mice. Donor-derived blood cells were regularly analyzed in the peripheral blood of secondary recipients. Data are presented as mean + standard deviation. *P < .05; **P < .01; ***P < .001.
