Exosomes Derived from AML/MDS Cells Is Involved in Stromal Dysfunction and Bone Marrow Failure

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4627-4627
Author(s):  
Hiroto Horiguchi ◽  
Masayoshi Kobune ◽  
Shohei Kikuchi ◽  
Wataru Jomen ◽  
Kazuyuki Murase ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Mrna Expression ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Leukemic Cells ◽  
Boyden Chamber ◽  
Hematopoietic Stem ◽  
Coculture System ◽  
Qpcr Array

Abstract The failure of normal hematopoiesis in myelodysplastic syndrome (MDS)/acute myeloid leukemia (AML) could be induced by a variety of mechanism such as the alteration of property of hematopoietic stem cells and stem cell niche. However, it has not yet been clarified precise mechanism how MDS stem/progenitor cells could replace normal hematopoietic stem/progenitor cells especially regarding involvement of mesenchymal stromal cells (MSCs). To gain insight into the mechanism of stromal dysfunction, comparative analyses of transcriptomes were conducted between normal and MDS/AML-derived MSCs. Further, we attempted to identify certain effectors originated from MDS/AML cells could alter the function of bone marrow (BM) MSCs. The MSCs derived from healthy volunteer (HV)-derived (normal) and MDS/AML-derived stromal cells were established and analyzed mRNA expression by quantitive PCR (qPCR) array. Additionally, the supporting activity of MSCs for BM CD34+ progenitor/stem cells was examined using serum free coculture system. The interaction between MDS/AML cells and MSCs were evaluated by using Boyden Chamber and the changes of mRNA expression were analyzed. The results of qPCR array revealed that the expression of hematopoietic factors was drastically altered in MDS/AML-derived MSCs as compared with normal MSCs. Among these factors, the expression of SCF and JAG1 mRNA were significantly and consistently reduced in all MDS/AML patients examined. Functional assay of these MSCs demonstrated that the number of colony-forming units (CFU) mixed cells (MIXs) and cobblestone area-forming cells (CAFCs) derived from CD34+ cells was significantly reduced after coculture with MDS/AML-derived MSCs as compared with normal MSCs. Even non-contact culture using Boyden Chamber between leukemic cells and MSCs induced the reduction of SCF and JAG1 mRNA, indicating that certain inducers could be soluble factors. Interestingly, this effect of transcriptomes alteration was negated by nSMase2 inhibitor (GW4869). Exosome transfer assay using Boyden Chamber revealed that GFP and PKH26 in leukemic cells transmit onto MSCs in non-contact coculture system and this transfer of exosome was significantly inhibited by GW4869 or nSMase siRNA. The multiple type of microRNA in exosome derived from MDS/AML cells was transferred into MSCs, suggesting that exosome could contribute to the alteration of mRNA expression in stromal cells. Collectively, these results indicated that exosome derived from MDS/AML cells could be involved in the reduction of SCF/JAG mRNA and the stromal supporting activity of normal hematopoietic stem/progenitor cells. Disclosures No relevant conflicts of interest to declare.

scholarly journals Acute exercise mobilizes hematopoietic stem and progenitor cells and alters the mesenchymal stromal cell secretome

2016 ◽  
Vol 120 (6) ◽  
pp. 624-632 ◽  
Author(s):  
Russell Emmons ◽  
Grace M. Niemiro ◽  
Olatomide Owolabi ◽  
Michael De Lisio
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Acute Exercise ◽  
Marrow Cell ◽  
Exercise Bout ◽  
Hematopoietic Stem ◽  
Stem And Progenitor Cells

Transplantation of hematopoietic stem and progenitor cells (HSPC), collected from peripheral blood, is the primary treatment for many hematological malignancies; however, variable collection efficacy with current protocols merits further examination into factors responsible for HSPC mobilization. HSPCs primarily reside within the bone marrow and are regulated by mesenchymal stromal cells (MSC). Exercise potently and transiently mobilizes HSPCs from the bone marrow into peripheral circulation. Thus the purpose of the present study was to evaluate potential factors in the bone marrow responsible for HSPC mobilization, investigate potential sites of HSPC homing, and assess changes in bone marrow cell populations following exercise. An acute exercise bout increased circulating HSPCs at 15 min (88%, P < 0.001) that returned to baseline at 60 min. Gene expression for HSPC homing factors (CXCL12, vascular endothelial growth factor-a, and angiopoietin-1) were increased at 15 min in skeletal muscle and HSPC content was increased in the spleen 48 h postexercise (45%, P < 0.01). Acute exercise did not alter HSPCs or MSCs quantity in the bone marrow; however, proliferation of HSPCs (40%, P < 0.001), multipotent progenitors (40%, P < 0.001), short-term hematopoietic stem cells (61%, P < 0.001), long-term hematopoietic stem cells (55%, P = 0.002), and MSCs (20%, P = 0.01) increased postexercise. Acute exercise increased the content of the mobilization agent granulocyte-colony stimulating factor, as well as stem cell factor, interleukin-3, and thrombopoeitin in conditioned media collected from bone marrow stromal cells 15 min postexercise. These findings suggest that the MSC secretome is responsible for HSPC mobilization and proliferation; concurrently, HSPCs are homing to extramedullary sites following exercise.

Reversible Expansion of Hematopoietic Stem Cells (HSC) and CML-Like Disease in Transgenic Mice Expressing BCR-ABL under the Control of the SCL 3′ Enhancer.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 715-715
Author(s):  
Steffen Koschmieder ◽  
Berthold Goettgens ◽  
Pu Zhang ◽  
Tajhal Dayaram ◽  
Kristin Geary ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Transgenic Mice ◽  
Hematopoietic Stem Cells ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Molecular Mechanisms ◽  
Leukemic Cells ◽  
Hematopoietic Stem ◽  
Double Transgenic Mice

Abstract Chronic myeloid leukemia (CML) is a malignant disorder originating from the transformation of hematopoietic stem cells (HSC) by the BCR-ABL oncogene. Using the tet-off system, we have generated double-transgenic mice in which BCR-ABL is expressed under the control of the murine SCL 3′ enhancer, which targets expression to the vast majority of HSC and progenitors. After induction of BCR-ABL, all mice developed progressive chronic neutrophilia and leukocytosis (20–40 K/ul), and the animals died or were sacrificed in moribund condition within 58+/−28 days. Upon necropsy, bone marrow granulocytic hyperplasia, splenomegaly as well as organ infiltration by leukemic cells (liver, kidney, lung, small intestine, skin) were found. In addition, 31% of the mice subsequently developed ALL or lymphomas. BCR-ABL mRNA and protein expression were demonstrated in the affected organs. Expression of the transactivating transgene tTA was high in HSC, CMP, and CLP, but low in GMP and MEP, as assessed by real-time PCR, suggesting that the SCL 3′ enhancer indeed directed BCR-ABL expression to the most primitive hematopoietic cells within the bone marrow. The percentage of HSC in the bone marrow was expanded 7- and 26-fold in double-transgenic as compared to single-transgenic or wild-type control mice within 12 and 21 days, respectively, after BCR-ABL induction. GMP were increased 2- and 3-fold while the number of CMP was decreased 2-fold after 12 days but was increased 1.5-fold after 21 days. MEP were decreased 3-fold at both time points. In keeping with these results, the percentage of Ter-119 positive erythroid cells was decreased while the percentage of Gr-1 positive granulocytic cells was increased in the bone marrow. To assess reversibility of the phenotype, we readministered tetracycline to abrogate BCR-ABL expression. Double-transgenic mice showed rapid clinical improvement, reversion of neutrophilia and leukocytosis, normalization of Gr-1/Mac-1 positive cells in the peripheral blood and spleen, and reversion of splenomegaly. In addition, in mice that had developed lymphoblastic disease, readministration of tetracycline led to disappearance of lymphomas and of B220/BP-1 positive lymphoblastic cells in the peripheral blood. Furthermore, expansion of the HSC compartment in the bone marrow was also reversible, and the percentage of HSC decreased to levels observed in control mice. Repeated induction of BCR-ABL expression by removal of tetracycline led to reappearance of the myeloid and lymphoid phenotype. Again, the disease was reversible, and none of the animals relapsed while on tetracycline, suggesting that the phenotype remained completely dependent on the expression of the oncogene. In conclusion, we present a model of BCR-ABL mediated CML-like disease with expansion of phenotypic hematopoietic stem cells and myeloid progenitor cells in the bone marrow. The target cell population in this model closely resembles the origin of transformation in patients with CML, allowing for in vivo monitoring of early molecular mechanisms of BCR-ABL transformation. We are currently studying the function of the expanded HSC and progenitor cells in transplantation experiments.

scholarly journals 5-Fluorouracil Treatment Leads to Activation of Stem Cell Niche By Reconstructing Mesenchymal Stromal Cells and Exert a Distinct Microenvironmental Impact on Normal and Leukemic Cells

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1197-1197
Author(s):  
Seon-Yeong Jeong ◽  
Jin-A Kim ◽  
Il-Hoan Oh
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Stromal Cells ◽  
Mesenchymal Cells ◽  
Leukemic Cells ◽  
Bone Marrow Niche ◽  
Hematopoietic Stem ◽  
Cellular Changes ◽  
Rapid Emergence

Abstract Reactivation of endogenous hematopoietic stem cells (HSCs) are initiated by stimulation of bone marrow niche triggered by various injury signals. Here, we show that treatment with 5-fluorouracil (5-FU) leads to reconstruction of bone marrow (BM) microenvironment to establish an activated niche stimulating hematopoietic stem cells (HSCs). First, we show that pre-treatment with 5-FU leads to engraftment of donor cells in non-irradiated recipient mice without affecting the homing efficiency of HSCs into BM. The HSC activation effects were reproduced in-vitro by co-culturing hematopoietic cells with CD45-Ter119- stromal cells derived from 5-FU treated BM, but not by co-culture with CD45+ cells or stromal cells obtained from enzymatic digestion of bone from the same mice. Examination of BM mesenchymal cells after 5-FU treatment revealed a rapid emergence of high-proliferating mesenchymal progenitors exhibiting large size colony (CFU-F) and higher self-renewal of colonogenic cells 3-5 days after 5-FU treatment, which was concomitantly associated with regeneration of CD34+Lin-Sca-1+c-kit+ (LSK) cells in the same BM. The cellular changes in mesenchymal stroma was associated with rapid emergence of characteristic mesenchymal cell populations (PDGFR-a+/Leptin receptor+/SSEA-3+: PLS) with 650-folds increase of the PLS cells in BM in 3 days after 5-FU treatment. However, the increase of these PLS mesenchymal cells were not associated with increase in mitotic activity of mesenchymal cells (<5% BrdU+ cells), indicating phenotypic conversion of subpopulation in BM. Moreover, cellular changes in mesenchymal niche were associated with rapid increase of mesenchymal cells expressing cross-talk molecules such as CXCL-12 (20-folds), Jagged-1 (13-folds) and DLL-1 (15-folds). Furthermore, in-vivo administration of chemicals blocking CXCL-12 and notch signaling during the recovery from the 5-FU treatment led to the significant loss of LSK-SLAM cells in the regenerated BM. Interestingly, the BM niche activated by 5-FU exerted a distinct effect on normal and leukemic cells in a manner that it provide higher support on the primitive state of normal HSCs than for MN-1 induced leukemia cells. Thus, leukemic mice engrafted with MN-1 cells exhibited a decrease in primitive leukemic cell (Lin-c-kit+) and higher survival by 5-FU treatment than those treated by radiation. Taken together, our study reveals the cellular reconstruction of mesenchymal niche in BM during stimulus-induced niche activation and provides an insight on the selective niche targeting as a novel therapeutic strategies for hematological diseases. Disclosures No relevant conflicts of interest to declare.

Id1 Provides a Proper Hematopoietic Progenitor Niche Function

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2427-2427
Author(s):  
Hyung Chan Suh ◽  
Ming Ji ◽  
John Gooya ◽  
Michael Lee ◽  
Kimberly Klarmann ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Mrna Expression ◽  
Progenitor Cells ◽  
Stromal Cells ◽  
Hematopoietic Progenitor ◽  
Vegf Mrna ◽  
Hematopoietic Stem ◽  
Gm Csf

Abstract Development of hematopoietic stem cells (HSC) and their progeny is maintained by the interaction with cells in the microenvironment. In addition to hematopoietic cells, Id1 is expressed in stromal cells known to support hematopoiesis, and is involved in cell proliferation, differentiation and senescence. Therefore, to investigate the role of Id1 in hematopoiesis, we examined hematologic phenotypes of Id1−/− mice. In this study, we found increased neutrophils and macrophages, and decreased B cells and platelets in peripheral blood, and decreased BM cellularity. While the percentages of hematopoietic stem cells (HSC) in Id1−/− mice were increased relative to the Id1+/+ mice, their total numbers and function appeared normal. For example, Id1 was not required for self-renewal or repopulation of HSC. In contrast, we found that there were increased numbers of hematopoietic progenitor cells (HPC) in S phase of cell cycle in Id1−/− mice BM, suggesting that the loss of Id1 within HPC promotes proliferation. However, purified Id1−/− HPC had the same proliferation potential as Id1+/+ HPC when cultured in vitro. In transplantation experiments, we proved that BM microenvironment in Id1−/− mice is defective by showing that the Id1+/+ HSC showed impaired hematopoietic development in Id1−/− mice, while the Id1−/− HSC had normal repopulation potential in an Id1+/+ microenvironment. In agreement with these findings, Id1−/− BM stromal cell cultures supported enhanced proliferation of hematopoietic progenitors. Furthermore, quantitative PCR showed that SCF, M-CSF, OPN, SDF-1 and TGF-α mRNA expression was decreased in Id1−/− stromal cells relative to Id1+/+ stromal cells, while G-CSF, GM-CSF, and VEGF mRNA expression was significantly increased. Id1−/− BM showed decreased number of mesenchymal stem/progenitor cells. Thus, Id1 does not play a role in maintaining HSC, but is involved in regulating hematopoietic progenitor niche. Funded by NCI contract No. N01-CO-12400.

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

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.

The role of children's bone marrow mesenchymal stromal cells in the ex vivo expansion of autologous and allogeneic hematopoietic stem cells

2015 ◽  
Vol 39 (10) ◽  
pp. 1099-1110 ◽  
Author(s):  
Iordanis Pelagiadis ◽  
Eftichia Stiakaki ◽  
Christianna Choulaki ◽  
Maria Kalmanti ◽  
Helen Dimitriou
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Hematopoietic Stem Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ex Vivo ◽  
Ex Vivo Expansion ◽  
Hematopoietic Stem

scholarly journals Primitive hematopoietic cells in murine bone marrow express the CD34 antigen

Blood ◽  
1996 ◽  
Vol 88 (10) ◽  
pp. 3774-3784 ◽  
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.

scholarly journals Major Histocompatibility Complex Restriction Between Hematopoietic Stem Cells and Stromal Cells In Vivo

Blood ◽  
1997 ◽  
Vol 89 (1) ◽  
pp. 49-54 ◽  
Author(s):  
Futoshi Hashimoto ◽  
Kikuya Sugiura ◽  
Kyoichi Inoue ◽  
Susumu Ikehara
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Major Histocompatibility Complex ◽  
Hematopoietic Stem Cells ◽  
Stromal Cells ◽  
Graft Failure ◽  
Major Histocompatibility ◽  
Hematopoietic Stem ◽  
Histocompatibility Complex ◽  
Colony Forming Units

Graft failure is a mortal complication in allogeneic bone marrow transplantation (BMT); T cells and natural killer cells are responsible for graft rejection. However, we have recently demonstrated that the recruitment of donor-derived stromal cells prevents graft failure in allogeneic BMT. This finding prompted us to examine whether a major histocompatibility complex (MHC) restriction exists between hematopoietic stem cells (HSCs) and stromal cells. We transplanted bone marrow cells (BMCs) and bones obtained from various mouse strains and analyzed the cells that accumulated in the engrafted bones. Statistically significant cell accumulation was found in the engrafted bone, which had the same H-2 phenotype as that of the BMCs, whereas only few cells were detected in the engrafted bones of the third-party H-2 phenotypes during the 4 to 6 weeks after BMT. Moreover, the BMCs obtained from the MHC-compatible bone showed significant numbers of both colony-forming units in culture (CFU-C) and spleen colony-forming units (CFU-S). These findings strongly suggest that an MHC restriction exists between HSCs and stromal cells.

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

Blood ◽  
1993 ◽  
Vol 81 (3) ◽  
pp. 801-807 ◽  
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.

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