Abstract
Engraftment of clonal hematopoietic precursor cells from patients with myelodysplastic syndrome (MDS) in immunodeficient mice has been difficult to achieve by intravenous (i.v.) injection. We used i.v. coadministration of the human marrow stroma cell line HS27a with CD34+ MDS cells in Nod.cg-Prkdcscid Il2rgtm1wjll (NSG) mice to provide signals that would facilitate engraftment. Hematopoietic cells from 24 MDS patients were transplanted. Cells from all patients were engrafted, and engraftment was documented in 44 of 46 evaluable mice (95%). Immunohistochemistry revealed human HS27a stroma colocalizing with human hematopoietic cells in mouse spleens. Human CD34+ precursors harvested from marrow and spleen of primary murine recipients, when combined with HS27a cells, were also engrafted successfully in secondary NSG recipients, showing persistence of the original clonal characteristics. This observation supports the concept that clonal markers were present in long-term repopulating cells. We suggest that HS27a stroma cells ‘traveled' in direct contact with hematopoietic precursors and enabled their propagation. An essential signal for engraftment appears to be CD146, which is prominently expressed on HS27a cells. This xenotransplantation model will allow to further dissect signals that control engraftment of MDS cells and should be amenable to in vivo treatment studies.
Highlights
Propagation of clonal CD34 þ cells derived from the marrow of patients with myelodysplastic syndrome (MDS), in contrast to cells from patients with acute myeloid leukemia (AML),[1,2] has proven difficult in vitro and has met with limited success in xenogeneic transplant models in vivo.[3,4,5,6] Extensive research has shown the central importance of the marrow microenvironment in supporting hematopoiesis, and vascular and stromal stem cell niches have been described.[7,8] Importantly, contact with components of the microenvironment supports normal hematopoiesis, and protects malignant cells, for example, in patients with AML, against the effects of cytotoxic therapy.[9,10] Additional data suggest that marrow stroma cells in patients with MDS show abnormal DNA methylation[11] and are functionally altered.[12,13] Raaijimarkers et al.[8] showed in a murine model that deletion of Dicer 1 in osteoblast progenitors resulted in the development of dysplastic hematopoiesis
MATERIALS AND METHODS Patients myelodysplastic syndrome (MDS) cells were obtained from marrow aspirates or from peripheral blood (PB) of patients referred to the Fred Hutchinson Cancer Research Center (FHCRC) for consultation or therapy
Intravenous coadministration of HS27a, but not HS5 stroma cells, supports engraftment of clonal human CD34 þ MDS cells in Nod.cg-Prkdcscid Il2rgtm1wjll (NSG) mice Previous studies had shown that the combined coinjection of two stroma cell lines, HS5 and HS27a, together with MDS marrow cells into the marrow cavity of immunodeficient mice, was associated with an inconsistent graft-enhancing effect.[3]
Summary
Propagation of clonal CD34 þ cells derived from the marrow of patients with myelodysplastic syndrome (MDS), in contrast to cells from patients with acute myeloid leukemia (AML),[1,2] has proven difficult in vitro and has met with limited success in xenogeneic transplant models in vivo.[3,4,5,6] Extensive research has shown the central importance of the marrow microenvironment in supporting hematopoiesis, and vascular and stromal stem cell niches have been described.[7,8] Importantly, contact with components of the microenvironment supports normal hematopoiesis, and protects malignant cells, for example, in patients with AML, against the effects of cytotoxic therapy.[9,10] Additional data suggest that marrow stroma cells in patients with MDS show abnormal DNA methylation[11] and are functionally altered.[12,13] Raaijimarkers et al.[8] showed in a murine model that deletion of Dicer 1 in osteoblast progenitors resulted in the development of dysplastic hematopoiesis. Cells were harvested from the bone marrows and spleens of two primary recipient NSG mice (transplanted with cells from the same patient and killed at 12–13 weeks), human CD45 þ cells (Supplementary Figure S2) were pooled and 7–10 Â 106 human CD45 þ cells, containing 4–5% CD34 þ cells, were injected together with HS27a cells (as used in primary recipients) into two secondary recipients.
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