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Letters to the Editor
ered by multinucleated tartrate-resistant acid phos- phatase (TRAP) positive osteoclasts (OcS/MS %) (Figure 2D). Mirroring this histologic finding, gene expression of the osteoclast differentiation regulators RANKL and OPG evaluated by quantitative RT-PCR in basal BMSC from both sources was similar (Figure 2E). These data indicate an impaired osteogenic potential of AML-derived BMSC, suggesting that leukemia cells can interfere with the mat- uration of osteoblast precursors which in turn results in reduced bone formation in the absence of changes in osteoclastic activity. It cannot be excluded that the osteogenic differentiation blockade may be a result of epigenetic changes in AML-BMSC affecting the genes involved in osteogenic cell development, such as PTX2 and TBX15 transcription factors.7 Our finding is consis- tent with previous reports demonstrating perturbation of the osteogenic niche in AML mouse models and in AML and myelodysplastic patients.2,4,14 Specifically, studies have shown that AML leads to the accumulation of oster- ix-expressing osteoblast-primed cells in murine BM.15
We next assessed the ability of AML-BMSC to form a
BM cavity and a functional stromal niche using a model
consisting in in vivo implantation of cartilage pellets fol-
lowed by the progressive substitution of cartilage by mar-
row through a process we named “endochondral myelo-
genesis”.16 AML-BMSC and HD-BMSC were grown as
unmineralized pellets in chondrogenic differentiation
medium and then implanted subcutaneously into NSG
mice. After 8 weeks, implanted chondroid pellets were
replaced by a BM hematopoietic microenvironment com-
posed of human-derived skeletal tissues (bone, cartilage,
fat, and perivascular cells), as confirmed by staining with
human-specific LaminA/C, and mouse-derived
hematopoietic cells (Figure 3A-B). Furthermore, in AML-
ossicles we detected the presence of human CD146-pos-
itive stromal cells associated with the vessel wall (Figure
3B). Interestingly, AML-BMSC derived ossicles contained
a significantly increased fraction occupied by adipocytes,
when compared to HD-BMSC transplants (adipocyte
area/marrow area [Ad.Ar/Ma.Ar] %; AML-derived vs.
HD-derived implants: 1.92±0.42 vs. 0.73±0.22; P=0.037)
(Figure 3C). Our results agree with other reports showing
that BM-stromal progenitors from AML mice have an
3
Lastly, the amount of hematopoietic tissue and the myeloid/erythroid (MPO+/TER-119+) ratio in normal and patient-derived ossicles were similar (Figure 3D).
In conclusion, we have demonstrated using in vivo physiologic models that AML-BMSC function is signifi- cantly altered in mature bone formation and niche com- position. As demonstrated by these in vivo transplanta- tion assays, BM-stromal progenitors from pediatric AML patients, even when removed from their pathological environment, show an intrinsically abnormal differentia- tion pattern with altered osteogenesis and increased adi- pogenic potential, which is not easily detectable by canonical in vitro assays. This suggests an instructive role of leukemic cells on the BM microenvironment that can contribute to the generation of a supportive niche for leukemic cells themselves. As our study leveraged in vivo models that appropriately reproduce the human BM niche, our data may have an important clinical relevance. Understanding the unique characteristics of the AML osteogenic niche represents a critical step towards unrav- eling the mechanisms underlying osteogenic niche-medi- ated support of AML cells and leukemic progression. Our
data suggests the possibility to target stage-specific cells of the osteogenic lineage to normalize the hostile BM niche and suppress AML cell development and prolifera- tion with an ultimate goal of inducing deep remissions and controlling long-term disease.
Alice Pievani,1* Samantha Donsante,2* Chiara Tomasoni,1 Alessandro Corsi,2 Francesco Dazzi,3 Andrea Biondi,1,4 Mara Riminucci2 and Marta Serafini1
*AP and SD contributed equally as co-first authors
1Centro Ricerca M. Tettamanti, Department of Pediatrics, University of Milano-Bicocca, Monza, Italy; 2Department of Molecular Medicine, Sapienza University, Rome, Italy; 3Department of Hemato-Oncology, Rayne Institute, King’s College London, London, UK and 4Department of Pediatrics, Fondazione MBBM/San Gerardo Hospital, Monza, Italy
Correspondence: MARTA SERAFINI - serafinim72@gmail.com doi:10.3324/haematol.2020.247205
Disclosures: no conflicts of interest to disclose.
Contributions: AP collected and analyzed the data and wrote the manuscript; SD performed in vivo experiments, analyzed the data, and contributed to the manuscript writing; CT performed in vitro experiments; AC and MR interpreted the data and critically revised the manuscript, FD and AB critically revised the manuscript;
MS supervised the research and wrote the manuscript.
Acknowledgments: the authors would like to thank Fondazione Matilde Tettamanti, Comitato Maria Letizia Verga, Fondazione MBBM, Associazione SKO Arianna Amore Onlus for their generous support.
Funding: this work was partially supported by AIRC-5 per mille 2018-21147 to AB, and AIRC-2015-17248 to MS.
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increased adipogenic differentiation ability. Moreover, Lu et al. found that marrow of AML patients in remission had less adipocyte content than cases from non-remis- sion marrows as compared with diagnostic marrows.17
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