Role of SCF-Expressing Bone Marrow Populations in Hematopoietic and Leukemic Stem Cell Regulation

Stem cell factor (SCF), the ligand for the c-Kit receptor, stimulates growth of early hematopoietic stem/progenitors cells (HSCs) directly and/or by stimulating other cytokines. Using transgenic and driver/reporter mice, studies have shown that, in physiological conditions the Scf gene is expressed in bone marrow microenvironmental (BMM) cells, in particular endothelial and perivascular stroma. Deletion of exon 1 from the Scf gene in endothelial cells (using Tek-cre), perivascular cells (using Lepr-cre) and adipocytes (using adipoq-cre) significantly reduced the frequency and function of hematopoietic stem cells (HSCs) in young adult mice (Zhou et al. Nature cell biology. 2017; Ding et al. Nature. 2013). These studies have been useful in identifying the different Scf-expressing niche compartments from the BMM that are important for HSC maintenance. However, leukemic stem cell (LSC ) regulation by specific BMM cells in CML is poorly understood. In our study, the different Scf-expressing niche populations were identified using the Scf-GFP knock-in model (Scf^gfp/+). CD45^-TER119^-CD31^-CD140⁺ (PαS) and CD45^-TER119^-CD31^-CD51⁺CD105⁺6C3⁺ (6C3) populations, representing mesenchymal progenitors and HSC-supporting stromal cells, respectively, showed the highest levels of Scf expression in normal mice, with endothelial cells also expressing SCF but at a lower level. We confirmed the role of Scf in normal hematopoiesis by showing that Scf^f/f - Ubc-cre mice generated by crossing Scf^f/f mice (LoxP sites flanking exon 1) with Ubc-cre mice (Cre mediated excision in widespread tissues following Tamoxifen administration) showed decreased stem/progenitor counts. To further evaluate the contribution of Scf-expressing populations towards regulation of normal HSC and LSCs, the Scf^f/f mice (LoxP sites flanking exon 1) were crossed with Bglap-cre (osteoblasts), Osx-cre (osteoprogenitors), Prx1-cre (mesenchymal progenitors) and Tek-cre (endothelial cells) mice. Deletion of Scf in the target populations was confirmed. We observed markedly reduced long-term HSC (LT-HSC; Lin^-Sca-1⁺Kit⁺CD48^-CD150⁺) numbers in the BM of 4-week old Scf^f/f-Prx1-cre mice, and to a lesser extent in Scf^f/f - Bglap-cre mice, and reduced short-term HSC (ST-HSC; Lin^-Sca-1⁺Kit⁺CD48^-CD150^-). We observed markedly reduced ST-HSC numbers, and LT-HSC to a lesser extent, in the BM of 4-week old Scf^f/f - Osx-cre mice, supporting a role for these Scf -expressing populations in normal HSC maintenance in developing animals. In contrast to previous reports, we did not observe changes in stem/progenitor numbers in Tek-cre animals. To further evaluate effects of Scf-expressing niche populations on regulation of normal HSC and CML LSC, BM cells from wild-type (1x10⁶ CD45.1 cells) and SCL-tTA-BCR-ABL mice (1x10⁶ CD45.1/2) were transplanted into irradiated (8Gy) Scf^f/f Cre- or Cre+ knockout animals and followed for hematopoietic reconstitution. Scf^f/f-Prx1-cre animals showed decreased numbers of normal LT-HSC and ST-HSC in the BM compared to WT mice, but did not show changes in CML LSC numbers, although BM and peripheral blood (PB) myeloid counts were reduced. Scf^f/f - Osx-cre mice showed decreased numbers of ST-HSC, but no changes in changes in normal HSC or CML LSC were seen. Scf^f/f-Bglap-cre mice did not show significant reduction in normal and CML LT-HSC in BM. Collectively, our results show that Scf -expressing mesenchymal stem cells play an important role in normal LT-HSC maintenance, and osteoprogenitors in normal ST-HSC maintenance. In contrast, CML LSCs are not dependent on SCF expression on the different mesenchymal populations studied here. Therefore, normal and leukemic stem cells appear to be differentially regulated by SCF-expressing BMM populations. Further studies to better define the Scf -expressing populations that support CML LSCs, and the effects of SCF-expressing BMM populations on LSC response to tyrosine kinase inhibitors are underway.