Hematopoiesis Remains Permissive to Bone Marrow Transplantation After Expansion of Progenitors and Resumption of Blood Cell Production

The immense regenerative power of hematopoietic tissue stems from the activation of the immature stem cells and the progenitor cells. After partial damage, hematopoiesis is reconstituted through a period of intense regeneration when blood cell production originates from erythro-myeloid progenitors in the virtual absence of stem cells. Since the damaged hematopoiesis can also be reconstituted from transplanted hematopoietic cells, we asked whether this also leads to the transient state when activated progenitors initially execute blood cell production. We first showed that the early reconstitution of hematopoiesis from transplanted cells gives rise to extended populations of developmentally advanced but altered progenitor cells, similar to those previously identified in the bone marrow regenerating from endogenous cells. We then identified the cells that give rise to these progenitors after transplantation as LSK CD48– cells. In the submyeloablative irradiated host mice, the transplanted LSK CD48– cells preferably colonized the spleen. Unlike the endogenous hematopoiesis reconstituting cells, the transplanted whole bone marrow cells and sorted LSK CD48– cells had greater potential to differentiate to B-lymphopoiesis. Separate transplantation of the CD150– and CD150+ subsets of LSK CD48– cells suggested that CD150– cells had a greater preference to B-lymphopoiesis than CD150+ cells. In the intensively regenerating hematopoiesis, the CD71/Sca-1 plot of immature murine hematopoietic cells revealed that the expanded populations of altered myeloid progenitors were highly variable in the different places of hematopoietic tissues. This high variability is likely caused by the heterogeneity of the hematopoiesis supporting stroma. Lastly, we demonstrate that during the period when active hematopoiesis resumes from transplanted cells, the hematopoietic tissues still remain highly permissive for further engraftment of transplanted cells, particularly the stem cells. Thus, these results provide a rationale for the transplantation of the hematopoietic stem cells in successive doses that could be used to boost the transplantation outcome.

Download Full-text

Primitive hematopoietic cells in murine bone marrow express the CD34 antigen

Blood ◽

10.1182/blood.v88.10.3774.bloodjournal88103774 ◽

1996 ◽

Vol 88 (10) ◽

pp. 3774-3784 ◽

Cited By ~ 65

Author(s):

F Morel ◽

SJ Szilvassy ◽

M Travis ◽

B Chen ◽

A Galy

Keyword(s):

Stem Cells ◽

Monoclonal Antibody ◽

Bone Marrow ◽

Progenitor Cells ◽

Significant Proportion ◽

Hematopoietic Cells ◽

Full Detail ◽

Hematopoietic Stem ◽

Cd34 Antigen

The CD34 antigen is expressed on most, if not all, human hematopoietic stem cells (HSCs) and hematopoietic progenitor cells, and its use for the enrichment of HSCs with repopulating potential is well established. However, despite homology between human and murine CD34, its expression on subsets of primitive murine hematopoietic cells has not been examined in full detail. To address this issue, we used a novel monoclonal antibody against murine CD34 (RAM34) to fractionate bone marrow (BM) cells that were then assayed in vitro and in vivo with respect to differing functional properties. A total of 4% to 17% of murine BM cells expressed CD34 at intermediate to high levels, representing a marked improvement over the resolution obtained with previously described polyclonal anti-CD34 antibodies. Sixty percent of CD34+ BM cells lacked lineage (Lin) markers expressed on mature lymphoid or myeloid cells. Eighty-five percent of Sca-1+Thy-1(10)Lin- /10 cells that are highly enriched in HSCs expressed intermediate, but not high, levels of CD34 antigen. The remainder of these phenotypically defined stem cells were CD34-. In vitro colony-forming cells, day-8 and -12 spleen colony-forming units (CFU-S), primitive progenitors able to differentiate into B lymphocytes in vitro or into T lymphocytes in SCID mice, and stem cells with radioprotective and competitive long-term repopulating activity were all markedly enriched in the CD34+ fraction after single-parameter cell sorting. In contrast, CD34-BM cells were depleted of such activities at the cell doses tested and were capable of only short-term B-cell production in vitro. The results indicate that a significant proportion of murine HSCs and multilineage progenitor cells express detectable levels of CD34, and that the RAM34 monoclonal antibody is a useful tool to subset primitive murine hematopoietic cells. These findings should facilitate more direct comparisons of the biology of CD34+ murine and human stem and progenitor cells.

Download Full-text

Disruption of CXCR4 Utilizing AMD3100 Bypasses Mobilization Deficiency Exhibited by CD26−/− Mice and Results in Efficient Mobilization of Myeloid Progenitors

Blood ◽

10.1182/blood.v112.11.3492.3492 ◽

2008 ◽

Vol 112 (11) ◽

pp. 3492-3492

Author(s):

Laura A. Paganessi ◽

Andrew L. Walker ◽

Stephanie A. Gregory ◽

Henry C. Fung ◽

Kent W. Christopherson

Keyword(s):

Bone Marrow ◽

Blood Cell ◽

Progenitor Cells ◽

Peripheral Blood ◽

Fold Increase ◽

Cxcr4 Antagonist ◽

Hematopoietic Stem ◽

Genetically Engineered Mice ◽

Colony Forming Units ◽

Myeloid Progenitors

Abstract The exopeptidase CD26 (also known as DPPIV/dipeptidylpeptidase IV) cleaves dipeptides from the N-terminus of proteins that contain the required X-Pro or X-Ala motif. We have previously reported that inhibition or loss of CD26 activity results in a deficiency in normal granulocyte-colony stimulating factor (G-CSF) induced mobilization, suggesting that CD26 is a necessary component of mobilization (Christopherson, et al Blood 2003 and Christopherson, et al Exp Hematol 2003). The chemokine CXCL12 (SDF-1, stromal cell derived factor-1) contains the appropriate recognition sequence for CD26 induced cleavage. This combined with the importance of CXCL12 in the trafficking of hematopoietic stem and progenitor cells (HSC/HPC) suggests CXCL12 as a likely functional target of CD26 during G-CSF induced mobilization. For this reason we therefore decided to investigate whether genetically engineered mice lacking CD26 (CD26−/−) could be mobilized utilizing the CXCR4 antagonist, AMD3100. To evaluate this, ten week old C57BL/6 and CD26−/− mice (also on a C57BL/6 background) received a single subcutaneous injection of AMD3100 (1mg/1kg). One hour following injection the mice were euthanized by CO2 inhalation. Peripheral blood was then obtained by heart stick with a 1.2 ml syringe containing EDTA as an anticoagulant. A complete blood count was taken for each peripheral blood sample. Following red blood cell lysis, cells were plated for myeloid colony formation in a standard 1% methylcellulose colony assay containing the appropriate cytokines. Following 7 days of incubation at 5% O2, 5% CO2 and 37°C plates were scored for colony-forming units-granulocyte macrophage (CFU-GM), burst-forming units-erythroid (BFU-E), and colony-forming units-granulocyte, erythroid, macrophage, and megakaryocytic (CFU-GEMM). Data is presented as the number of colonies per femur for the bone marrow and as the number of colonies per ml of whole blood for the peripheral blood. AMD3100 treatment resulted in an increase in white blood cell (WBC) counts from 5.05±0.48 × 106/ml in untreated mice to 10.21±0.88×106/ml in treated mice (p≤0.01). An increase in WBC counts was also observed during AMD3100 treatment in CD26−/− mice from 7.77±1.28×106/ml in untreated mice to 16.7 ±2.11 × 106/ml in treated mice (p<0.01). AMD3100 treatment resulted in an increase in circulating myeloid progenitors in the peripheral blood of C57BL/6 and CD26−/− mice as compared to untreated C57BL/6 and CD26−/− mice respectively (p≤0.01). Specifically, a 2.38, 3.75, 12.33 fold increase in CFU-GM, BFU-E, and CFU-GEMM were observed in the peripheral blood of C57BL/6 mice after treatment. A 2.63, 5.48, 14.29 fold increase in CFU-GM, BFU-E, and CFU-GEMM were observed in the peripheral blood of CD26−/− mice after treatment. Existing pre-clinical and clinical data suggest that the CXCR4 antagonist, AMD3100, rapidly mobilizes hematopoietic progenitor cells from the bone marrow into the periphery. The results presented here provide pre-clinical evidence that disruption of the interaction between the CXCR4 chemokine receptor and CXCL12, via sub-cutaneous injection of AMD3100, mobilizes significant numbers of myeloid progenitors in mice, even in the absence of CD26. These results support the notion that CD26 is downstream of G-SCF treatment. Additionally, these results support the potential use of AMD3100 to treat patients that may have an altered ability to respond to G-CSF treatment as a result of a reduction or loss in CD26 activity. Future studies are warranted to evaluate potential variations in CD26 levels or activity in the general population, in differing patient populations, and during different treatment regimens.

Download Full-text

Poly I:C Stimulates Sca-1 Expression in Murine Bone Marrow and Increases the Number of Phenotypic Hematopoietic Stem Cells.

Blood ◽

10.1182/blood.v114.22.2504.2504 ◽

2009 ◽

Vol 114 (22) ◽

pp. 2504-2504

Author(s):

Russell Garrett ◽

Gerd Bungartz ◽

Alevtina Domashenko ◽

Stephen G. Emerson

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Conflicts Of Interest ◽

Hematopoietic Cells ◽

Poly I:C ◽

Hematopoietic Stem ◽

Marrow Cells ◽

Myeloid Progenitors

Abstract Abstract 2504 Poster Board II-481 Polyinosinic:polycytidlyic acid (poly I:C) is a synthetic double-stranded RNA used to mimic viral infections in order to study immune responses and to activate gene deletion in lox-p systems employing a Cre gene responsive to an Mx-1 promoter. Recent observations made by us and others have suggested hematopoietic stem cells, responding to either poly I:C administration or interferon directly, enter cell cycle. Twenty-two hours following a single 100mg intraperitoneal injection of poly I:C into 10-12 week old male C57Bl/6 mice, the mice were injected with a single pulse of BrdU. Two hours later, bone marrow was harvested from legs and stained for Lineage, Sca-1, ckit, CD48, IL7R, and BrdU. In two independent experiments, each with n = 4, 41 and 33% of Lin- Sca-1+ cKit+ (LSK) IL-7R- CD48- cells from poly I:C-treated mice had incorporated BrdU, compared to 7 and 10% in cells from PBS-treated mice. These data support recently published reports. Total bone marrow cellularity was reduced to 45 and 57% in the two experiments, indicating either a rapid death and/or mobilization of marrow cells. Despite this dramatic loss of hematopoietic cells from the bone marrow of poly I:C treated mice, the number of IL-7R- CD48- LSK cells increased 145 and 308% in the two independent experiments. Importantly, the level of Sca-1 expression increased dramatically in the bone marrow of poly I:C-treated mice. Both the percent of Sca-1+ cells and the expression level of Sca-1 on a per cell basis increased after twenty-four hours of poly I:C, with some cells acquiring levels of Sca-1 that are missing from control bone marrow. These data were duplicated in vitro. When total marrow cells were cultured overnight in media containing either PBS or 25mg/mL poly I:C, percent of Sca-1+ cells increased from 23.6 to 43.7%. Within the Sca-1+ fraction of poly I:C-treated cultures, 16.7% had acquired very high levels of Sca-1, compared to only 1.75% in control cultures. Quantitative RT-PCR was employed to measure a greater than 2-fold increase in the amount of Sca-1 mRNA in poly I:C-treated cultures. Whereas the numbers of LSK cells increased in vivo, CD150+/− CD48- IL-7R- Lin- Sca-1- cKit+ myeloid progenitors almost completely disappeared following poly I:C treatment, dropping to 18.59% of control marrow, a reduction that is disproportionately large compared to the overall loss of hematopoietic cells in the marrow. These cells are normally proliferative, with 77.1 and 70.53% accumulating BrdU during the 2-hour pulse in PBS and poly I:C-treated mice, respectively. Interestingly, when Sca-1 is excluded from the analysis, the percent of Lin- IL7R- CD48- cKit+ cells incorporating BrdU decreases following poly I:C treatment, in keeping with interferon's published role as a cell cycle repressor. One possible interpretation of these data is that the increased proliferation of LSK cells noted by us and others is actually the result of Sca-1 acquisition by normally proliferating Sca-1- myeloid progenitors. This new hypothesis is currently being investigated. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

Mdm2 Haplo-Insufficiency in Hematopoietic and Mesenchymal Progenitor Cells Results in Cell Death

Blood ◽

10.1182/blood-2020-142662 ◽

2020 ◽

Vol 136 (Supplement 1) ◽

pp. 10-10

Author(s):

Rasoul Pourebrahim ◽

Rafael Heinz Montoya ◽

Zoe Alaniz ◽

Lauren B Ostermann ◽

Edward Ayoub ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Progenitor Cells ◽

Research Funding ◽

Hematopoietic Cells ◽

Mesenchymal Progenitor Cells ◽

Heterozygous Deletion ◽

Hematopoietic Stem ◽

Osteoprogenitor Cells

The murine double minute 2 (Mdm2) protein is an important negative regulator of the p53 tumor suppressor, required for normal embryonic development and homeostasis. In humans, a single nucleotide polymorphism in the MDM2 promoter is associated with increased risk of cancer suggesting the importance of MDM2 levels in tumorigenesis (Bond et al., 2004). Mice with Mdm2 haploinsufficiency were previously reported as phenotypically normal with increased p53-dependent response to ionizing radiation (IR) resulting in lethal bone marrow failure (Terzian et al., 2007). However, the mechanism of radiosensitivity in these mice is unknown. To better characterize the phenotype of Mdm2 haploinsufficient mice and explore the mechanism of IR sensitivity, we developed a lineage tracing system to genetically label and trace the fate of cells after heterozygous deletion of Mdm2 in hematopoietic as well as mesenchymal progenitor cells. We utilized mTmG allele as a traceable reporter in which green fluorescence (GFP) replaces red fluorescence (TdTomato) after Cre-mediated recombination. Using Vav-Cre or Mx1-Cre, we first targeted Mdm2 in hematopoietic cells and marked them by TdTomato (Mdm2-WT) and GFP (Mdm2+/-). Heterozygous deletion of Mdm2 in hematopoietic stem cells using Vav-Cre resulted in massive apoptosis of emerging hematopoietic progenitor cells in the aorta-gonad-mesonephros (AGM) region at E10.5. Marker segregation analysis by fluorescence microscopy and flow cytometry revealed a population of hematopoietic stem cells having both TdTomato and GFP markers that escaped from apoptosis and reconstituted the hematopoietic cells in the fetal liver. Deletion of p53 in these mice did not rescue the apoptotic phenotype of hematopoietic cells with Mdm2 haploinsufficiency suggesting that a non-p53 dependent function of Mdm2 is necessary for proper development of hematopoietic stem cells in early development. In adult mice, Mdm2 haploinsufficiency in hematopoietic cells resulted in significant reduction in bone marrow hematopoietic stem cells in the absence of IR induced cellular stress. In Mx1-Cre;mTmG;Mdm2fl/+ mice, induction of Cre activity by pIpC injection resulted in hematopoietic failure evident by pancytopenia in peripheral blood. To test whether the same apoptotic response to Mdm2 haploinsufficiency can occur in other lineages, we generated a traceable conditional model of Mdm2 haploinsufficiency in mesenchymal progenitor cells using Osx-Cre and Prx1-Cre. Mice with heterozygous deletion of Mdm2 (Osx-Cre;mTmG;Mdm2fl/+) showed apoptosis of emerging osteoprogenitor cells at E16.5. Analysis of bone at 4 weeks revealed significant apoptosis of emerging osteoprogenitor cells further supporting our findings in the hematopoietic lineage. Together, our data highlights the importance of Mdm2 levels in hematopoietic and mesenchymal stem cell hemostasis and identifies depletion of hematopoietic stem cells in the bone marrow as the mechanism of radiosensitivity in Mdm2 haploinsufficient mice. Disclosures Andreeff: Daiichi-Sankyo; Breast Cancer Research Foundation; CPRIT; NIH/NCI; Amgen; AstraZeneca: Research Funding; Daiichi-Sankyo; Jazz Pharmaceuticals; Celgene; Amgen; AstraZeneca; 6 Dimensions Capital: Consultancy; Centre for Drug Research & Development; Cancer UK; NCI-CTEP; German Research Council; Leukemia Lymphoma Foundation (LLS); NCI-RDCRN (Rare Disease Clin Network); CLL Founcdation; BioLineRx; SentiBio; Aptose Biosciences, Inc: Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding.

Download Full-text

Cdc42 Is Critical for Multi-Lineage Blood Cell Development from Hematopoietic Stem and Progenitor Cells.

Blood ◽

10.1182/blood.v108.11.633.633 ◽

2006 ◽

Vol 108 (11) ◽

pp. 633-633 ◽

Cited By ~ 1

Author(s):

Linda Yang ◽

Lei Wang ◽

Hartmut Geiger ◽

Jose A. Cancelas ◽

David A. Williams ◽

...

Keyword(s):

Bone Marrow ◽

Mouse Model ◽

Blood Cell ◽

Progenitor Cells ◽

Peripheral Blood ◽

Rho Gtpase ◽

Cell Development ◽

Hematopoietic Stem ◽

Lineage Differentiation ◽

Myeloid Progenitors

Abstract Cdc42 is a member of the Rho GTPase family regulating multiple functions in eukaryotic cells. However, its involvement in blood cell development is not well studied. In our recent work using an inducible, conditional gene targeted mouse model (Mx-Cre;Cdc42loxP/loxP), we have shown that Cdc42 is important for hematopoietic stem cell (HSC) homing, lodging and engraftment, as well as maintenance of long-term HSC quiescence in the hematopoietic microenvironment. To define the contribution of Cdc42 to multi-lineage differentiation of HSCs, we have examined this mouse model for lymphoid, erythroid and myeloid lineage development. Firstly, although Cdc42 gene disruption in HSCs does not alter the overall bone marrow (BM) cellularity, it affects the frequency of multipotent progenitor cells (Table). The number of common lymphoid progenitors (CLPs) remain unchanged, but the number of phenotypic common myeloid progenitors (CMP) decreases, the number of phenotypic granulocytic myeloid progenitors (GMP) increases, and the myeloerythroid progenitors (MEP) decreases, in the Cdc42−/− mice. Secondly, despite the normal CLP content in the BM, the cellularity of Cdc42−/− thymus and T-lymphoid populations in peripheral blood is markedly reduced (Table). The T-cell developmental defect is associated with a thymus-homing defect of the Cdc42−/− BM cells. Thirdly, the Cdc42−/− mice show severe anemia and splenomegaly that are associated with dramatically decreased BFU-E and CFU-E activities of the BM cells (Table). FACS analysis showed an accumulation of early (CD71hiTER119−/low) erythroid cells and a reduction of the more mature TER119hi populations of erythroid progenitors in the Cdc42−/− bone marrow (Table), suggesting that Cdc42-deficiency causes a block in erythroid differentiation. Finally, Cdc42-deficiency in the HSCs led to a myeloproliferative disorder (MPD) phenotype, characterized by significantly increased monocytes, granulocytes and myeloid progenitors in the peripheral blood (Table), massive deposit of Gr-1+/Mac-1+ myeloid cells in the BM, and infiltration by the myeloid cells into distal organs such as spleen, liver and lung. These effects are a consequence of increased cell cycle progression and cell expansion accompanying the markedly increased mobilization of the short-term HSCs and myeloid progenitors from the BM to the peripheral organs found in the Cdc42−/− mice. Taken together, our results show that in addition to an essential role in maintaining HSC quiescence in the bone marrow niche, the Rho GTPase Cdc42 is critically involved in regulating multi-lineage differentiation of HSCs, including lymphopoiesis, erythropoiesis and myelopoiesis, through distinct mechanisms. WT Cdc42−/− *p<0.05 **p<0.01 ***p<0.001 BM cellularity (X106) 113.0±15.2 116.8±26.4 CLP (X106) 0.0223±0.0139 0.0390±0.0277 CMP (X106) 0.228±0.093 0.077±0.029* GMP (X106) 1.47±0.33 2.70±1.07* MEP (X106) 0.16±0.058 0.051±0.031* Thymus weight (X10−3g/g body weight) 2.37±0.78 0.49±0.36*** CD4+CD8− (/ul PB) 614.3±340.7 282.2±179.1* CD4−CD8+ (/ul PB) 446.1±332.7 181.2±56.7* BFU-E (/105 BM cells) 36±8.9 14.4±3.7 CFU-E (/105 BM cells) 87.2±12.4 26.4±5.7* CD71+Ter119− cells in BM (Stage I) 1.3%±0.06% 3.4%±0.38%*** CD71−Ter119+ cells in BM (StageII-IV) 39.7%±10.8% 7.8%±1.3%** Gr1/Mac1+ cells in BM 47.8%±27.2% 83.3%±8.9%**

Download Full-text

Mechanism of Action of Diallyl Sulphide in Ameliorating the Hematopoietic Radiation Injury

Journal of Health and Allied Sciences NU ◽

10.1055/s-0041-1730094 ◽

2021 ◽

Author(s):

Omika Katoch ◽

Mrinalini Tiwari ◽

Namita Kalra ◽

Paban K. Agrawala

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Progenitor Cells ◽

Hematopoietic Stem ◽

Proliferation And Differentiation ◽

Stem And Progenitor Cells ◽

Radiation Induced ◽

Medicinal Properties ◽

Marrow Cellularity

AbstractDiallyl sulphide (DAS), the pungent component of garlic, is known to have several medicinal properties and has recently been shown to have radiomitigative properties. The present study was performed to better understand its mode of action in rendering radiomitigation. Evaluation of the colonogenic ability of hematopoietic progenitor cells (HPCs) on methocult media, proliferation and differentiation of hematopoietic stem cells (HSCs), and transplantation of stem cells were performed. The supporting tissue of HSCs was also evaluated by examining the histology of bone marrow and in vitro colony-forming unit–fibroblast (CFU-F) count. Alterations in the levels of IL-5, IL-6 and COX-2 were studied as a function of radiation or DAS treatment. It was observed that an increase in proliferation and differentiation of hematopoietic stem and progenitor cells occurred by postirradiation DAS administration. It also resulted in increased circulating and bone marrow homing of transplanted stem cells. Enhancement in bone marrow cellularity, CFU-F count, and cytokine IL-5 level were also evident. All those actions of DAS that could possibly add to its radiomitigative potential and can be attributed to its HDAC inhibitory properties, as was observed by the reversal radiation induced increase in histone acetylation.

Download Full-text

Identification of BCR/ABL-negative primitive hematopoietic progenitor cells within chronic myeloid leukemia marrow

Blood ◽

10.1182/blood.v81.3.801.bloodjournal813801 ◽

1993 ◽

Vol 81 (3) ◽

pp. 801-807 ◽

Cited By ~ 1

Author(s):

T Leemhuis ◽

D Leibowitz ◽

G Cox ◽

R Silver ◽

EF Srour ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Chronic Myeloid Leukemia ◽

Progenitor Cells ◽

Myeloid Leukemia ◽

Chronic Phase ◽

Polymerase Chain Reaction Analysis ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor ◽

Hematopoietic Stem

Chronic myeloid leukemia (CML) is a malignant disorder of the hematopoietic stem cell. It has been shown that normal stem cells coexist with malignant stem cells in the bone marrow of patients with chronic-phase CML. To characterize the primitive hematopoietic progenitor cells within CML marrow, CD34+DR- and CD34+DR+ cells were isolated using centrifugal elutriation, monoclonal antibody labeling, and flow cytometric cell sorting. Polymerase chain reaction analysis of RNA samples from these CD34+ subpopulations was used to detect the presence of the BCR/ABL translocation characteristic of CML. The CD34+DR+ subpopulation contained BCR/ABL(+) cells in 11 of 12 marrow samples studied, whereas the CD34+DR- subpopulation contained BCR/ABL(+) cells in 6 of 9 CML marrow specimens. These cell populations were assayed for hematopoietic progenitor cells, and individual hematopoietic colonies were analyzed by PCR for their BCR/ABL status. Results from six patients showed that nearly half of the myeloid colonies cloned from CD34+DR- cells were BCR/ABL(+), although the CD34+DR- subpopulation contained significantly fewer BCR/ABL(+) progenitor cells than either low-density bone marrow (LDBM) or the CD34+DR+ fraction. These CD34+ cells were also used to establish stromal cell-free long-term bone marrow cultures to assess the BCR/ABL status of hematopoietic stem cells within these CML marrow populations. After 28 days in culture, three of five cultures initiated with CD34+DR- cells produced BCR/ABL(-) cells. By contrast, only one of eight cultures initiated with CD34+DR+ cells were BCR/ABL(-) after 28 days. These results indicate that the CD34+DR- subpopulation of CML marrow still contains leukemic progenitor cells, although to a lesser extent than either LDBM or CD34+DR+ cells.

Download Full-text

Cell interactions in the bone marrow microenvironment affecting myeloid malignancies

Blood Advances ◽

10.1182/bloodadvances.2020002127 ◽

2020 ◽

Vol 4 (15) ◽

pp. 3795-3803 ◽

Cited By ~ 2

Author(s):

Konstantinos D. Kokkaliaris ◽

David T. Scadden

Keyword(s):

Bone Marrow ◽

Blood Cell ◽

Stromal Cells ◽

Therapeutic Intervention ◽

Oxygen Delivery ◽

Hematopoietic Cells ◽

Cell Interactions ◽

Hematopoietic Cell ◽

Cell Production ◽

Myeloid Malignancy

Abstract The bone marrow is a complex tissue in which heterogeneous populations of stromal cells interact with hematopoietic cells to dynamically respond to organismal needs in defense, hemostasis, and oxygen delivery. Physiologic challenges modify stromal/hematopoietic cell interactions to generate changes in blood cell production. When either stroma or hematopoietic cells are impaired, the system distorts. The distortions associated with myeloid malignancy are reviewed here and may provide opportunities for therapeutic intervention.

Download Full-text

In Vitro Murine Hematopoiesis Supported by Signaling from a Splenic Stromal Cell Line

Stem Cells International ◽

10.1155/2018/9896142 ◽

2018 ◽

Vol 2018 ◽

pp. 1-9 ◽

Cited By ~ 1

Author(s):

Hong Kiat Lim ◽

Pravin Periasamy ◽

Helen C. O’Neill

Keyword(s):

Bone Marrow ◽

Cell Line ◽

Stromal Cell ◽

Hematopoietic Cells ◽

Model Systems ◽

Hematopoietic Stem ◽

Cell Production ◽

Stromal Cell Line ◽

Reticular Cells

There are very few model systems which demonstrate hematopoiesis in vitro. Previously, we described unique splenic stromal cell lines which support the in vitro development of hematopoietic cells and particularly myeloid cells. Here, the 5G3 spleen stromal cell line has been investigated for capacity to support the differentiation of hematopoietic cells from progenitors in vitro. Initially, 5G3 was shown to express markers of mesenchymal but not endothelial or hematopoietic cells and to resemble perivascular reticular cells in the bone marrow through gene expression. In particular, 5G3 resembles CXCL12-abundant reticular cells or perivascular reticular cells, which are important niche elements for hematopoiesis in the bone marrow. To analyse the hematopoietic support function of 5G3, specific signaling pathway inhibitors were tested for the ability to regulate cell production in vitro in cocultures of stroma overlaid with bone marrow-derived hematopoietic stem/progenitor cells. These studies identified an important role for Wnt and Notch pathways as well as tyrosine kinase receptors like c-KIT and PDGFR. Cell production in stromal cocultures constitutes hematopoiesis, since signaling pathways provided by splenic stroma reflect those which support hematopoiesis in the bone marrow.

Download Full-text

Identification of BCR/ABL-negative primitive hematopoietic progenitor cells within chronic myeloid leukemia marrow

Blood ◽

10.1182/blood.v81.3.801.801 ◽

1993 ◽

Vol 81 (3) ◽

pp. 801-807 ◽

Cited By ~ 49

Author(s):

T Leemhuis ◽

D Leibowitz ◽

G Cox ◽

R Silver ◽

EF Srour ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Chronic Myeloid Leukemia ◽

Progenitor Cells ◽

Myeloid Leukemia ◽

Chronic Phase ◽

Polymerase Chain Reaction Analysis ◽

Hematopoietic Progenitor Cells ◽

Hematopoietic Progenitor ◽

Hematopoietic Stem

Abstract Chronic myeloid leukemia (CML) is a malignant disorder of the hematopoietic stem cell. It has been shown that normal stem cells coexist with malignant stem cells in the bone marrow of patients with chronic-phase CML. To characterize the primitive hematopoietic progenitor cells within CML marrow, CD34+DR- and CD34+DR+ cells were isolated using centrifugal elutriation, monoclonal antibody labeling, and flow cytometric cell sorting. Polymerase chain reaction analysis of RNA samples from these CD34+ subpopulations was used to detect the presence of the BCR/ABL translocation characteristic of CML. The CD34+DR+ subpopulation contained BCR/ABL(+) cells in 11 of 12 marrow samples studied, whereas the CD34+DR- subpopulation contained BCR/ABL(+) cells in 6 of 9 CML marrow specimens. These cell populations were assayed for hematopoietic progenitor cells, and individual hematopoietic colonies were analyzed by PCR for their BCR/ABL status. Results from six patients showed that nearly half of the myeloid colonies cloned from CD34+DR- cells were BCR/ABL(+), although the CD34+DR- subpopulation contained significantly fewer BCR/ABL(+) progenitor cells than either low-density bone marrow (LDBM) or the CD34+DR+ fraction. These CD34+ cells were also used to establish stromal cell-free long-term bone marrow cultures to assess the BCR/ABL status of hematopoietic stem cells within these CML marrow populations. After 28 days in culture, three of five cultures initiated with CD34+DR- cells produced BCR/ABL(-) cells. By contrast, only one of eight cultures initiated with CD34+DR+ cells were BCR/ABL(-) after 28 days. These results indicate that the CD34+DR- subpopulation of CML marrow still contains leukemic progenitor cells, although to a lesser extent than either LDBM or CD34+DR+ cells.

Download Full-text