Mesenchymal Stromal Cells, Instigator or Suppressor for the Development of MLL-AF9 Induced Acute Myeloid Leukemia?

It has been reported that bone marrow (BM) hematopoietic niche is critical for the maintenance of normal hematopoiesis and the development of hematopoietic disorders. However, little is known about how the different BM niche components contribute to the initiation and progression of acute myeloid leukemia (AML). To explore the role of BM stromal cells in the development of AML, we here characterized the BM stromal cells in AML mouse model induced by transplantation of mouse BM cellstransduced with MLL-AF9 oncogene. We subdivided the BM stromal cells (CD45^-TER119^-) into mesenchymal stem cells (MSCs, CD31^-CD44^-CD51⁺SCA1⁺), mesenchymal progenitor cells (MPCs, CD31^-CD44^-CD51⁺SCA1^-), and endothelial cells (CD31⁺) in the mice after development of AML by multicolor fluorescent activated cell sorting (FACS) (Qian et al., Mol Cell Biol, 2013). We found significantly increased frequencies of BM MSCs (p < 0.0001), MPCs (p < 0.0001), and endothelial cells (p < 0.0001) in the AML mice, indicating an AML-induced alteration of BM stromal cell composition. The increased frequency of MSCs in AML mouse BM was confirmed by the increased number of colony foming unit-fibroblasts in the AML mice (p < 0.0001). Furthermore, in vitro multi-lineage differentiation assay revealed increased adipogenic and osteogenic differentiation capacities of the BM MSCs in the AML mice. Consistent with the increased osteogenic differentiation capacity of AML MSCs, Runx2 mRNA expression was increased in freshly sorted AML MSCs compared to that in the control mice. In addition, quantitative real time PCR on freshly sorted BM MPCs showed a dramatic downregulation of the genes important in hematopoietic stem cell regulation, including angiopoietin like 1 (Angptl1), C-X-C motif chemokine (Cxcl12), and kit ligand (Kitl) in the AML mice. The altered gene expression might contribute to the AML progression.

To specifically determine the contribution of MSCs to AML development, we analyzed the MSC population expressing Early B cell factor 2 (Ebf2) (Qian et al., Mol Cell Biol, 2013) by establishing the AML in Ebf2-Egfp transgenic recipient mice. We detected an increased frequency of Ebf2⁺ cells in the recipient BM after development of AML (p = 0.0018). In addition, by using the triple transgenic Tg(Ebf2-Egfp x Ebf2-Cre^ER x Rosa26-^loxpStop^loxp-Tomato) mouse model, where Ebf2⁺ MSCs and their progenies can be traced by tomato fluorescent protein after tamoxifen treatment, we found that the MSCs and MPCs generated from the Ebf2⁺ MSCs were also increased in the AML mouse BM (p < 0.05).

To further investigate the functional contribution of Ebf2⁺ MSCs to the AML progression, we took advantage of the transgenic Tg(Ebf2-Cre^ER x Rosa26-^loxpStop^loxp-DTA) mouse model allowing specific deletion of the Ebf2-expressing MSCs. Strikingly, we observed an accelerated initiation of AML, illustrated by the shorter survival of the MSC depleted recipients (Figure 1), compared to the non-depleted recipients, suggesting that Ebf2⁺ MSCs could suppress AML progression.

Taken together, our data show that BM stromal cell composition and function are altered in the mice with AML and the BM MSCs may act to suppress the progression of the MLL-AF9 induced AML. These findings provide new evidence for the niche contribution to the development of AML.

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