Treatment of Bone Marrow Mononuclear Cells with Erythropoietin and Platelet-Derived Growth Factor-bb Promotes Cellular Growth and Expression of Mesenchymal and Endothelial Cell Surface Markers In Vitro

2006 ◽  
Vol 12 (6) ◽  
pp. S33
Author(s):  
Jordan J. Lancaster ◽  
Nicholle M. Johnson ◽  
Mohamed A. Gaballa ◽  
Hoang M. Thai ◽  
Steven Goldman
Keyword(s):  
Bone Marrow ◽  
Endothelial Cell ◽  
Mononuclear Cells ◽  
Cellular Growth ◽  
Platelet Derived Growth Factor ◽  
Surface Markers ◽  
Bone Marrow Mononuclear Cells ◽  
Cell Surface Markers ◽  
Endothelial Cell Surface

scholarly journals Combining Effects of the SMO Inhibitor and Jak1 Inhibitor in Terminal Differentiation of MDS-Derived Induced Potent Stem Cells (iPSC)

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3150-3150
Author(s):  
Tetsuzo Tauchi ◽  
Seiichi Okabe ◽  
Seiichiro Katagiri ◽  
Yuko Tanaka ◽  
Kazuma Ohyashiki
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Research Funding ◽  
Mononuclear Cells ◽  
Nuclear Translocation ◽  
Terminal Differentiation ◽  
Scid Mice ◽  
Cytokine Signaling ◽  
Bone Marrow Mononuclear Cells

Abstract Background: Myelodysplastic syndromes (MDS) are clonal hematopoietic disorders characterized by no efficient hematopoiesis and frequent progression to acute myeloid leukemia (AML). Even in low risk MDS, clonal hematopoiesis already dominates at diagnosis, and clones found in secondary AML originate from the MDS stage of disease, highlighting the need to specifically target the MDS-initiating clone. PF-0449913 is a potent and selective hedgehog pathway inhibitor that act by binding Smoothened (SMO) and blocking signal transduction. PF-04449913 demonstrated preliminary antitumor activity in a phase I trial, when given as monotherapy in patients with several hematopoietic malignancy. Jak1 tyrosine kinase plays an important role in cytokine signaling. Jak1 functions to phosphorylate STAT3 transcription factor, which triggers their dimerization and nuclear translocation. In the present study, we investigated the combining effects of PF-04449913 and Jak1 inhibitor, PF-6667291 in terminal differentiation of MDS-derived induced potent stem cells (iPSC). Methods: We generated iPSCs from bone marrow mononuclear cells of two MDS patients (RAEB1 and RAEB2 by WHO classification) with chromosome 5 deletion and complex karyotypic abnormalities, respectively. Karyotyping analysis revealed that MDS-derived iPSCs have identical abnormalities to primary MDS cells. We also generated iPSCs from bone marrow mononuclear cells of normal volunteer as control. To investigate the effects of PF-04449913 on self-renewal and the relevance as a therapeutic target in MDS initiating cells, we examined the activity of PF-04449913 against MDS-derived iPSCs transferred NOD/SCID mice in vivo. NOD/SCID mice were injected subcutaneously with MDS-derived iPSCs or normal iPSCs then treated with PF-04449913 (100 mg/kg; p.o.) from day 10 for 28 days. We also used MDS-L, a myelodysplastic cell line established from MDS patient with del (5q) and complex karyotypic abnormalities for in vitro studies. In vitro re-differentiation of MDS-iPSCs was performed with differentiation media (30 ng/ml VEGF, 30 ng/ml BMP-4, 40 ng/ml SCF, 50 ng/ml Activin) for 4 days. At day 14, a single cell suspension expressing CD34+CD38- was achieved with hematopoietic cytokines (300 ng/ml Flt-3 ligand, 10 ng/ml IL-3, 10 ng/ml IL-6, 50 ng/ml G-CSF, 25 ng/ml BMP-4). Results: Both MDS-derived iPSCs transferred NOD/SCID mice and normal iPSCs transferred NOD/SCID mice demonstrated the engraftment of CD34+CD38- positive cells by flow cytometry. However, the treatment with PF-04449913 reduced the population of CD34+CD38- positive cells in MDS-derived iPSCs transferred NOD/SCID mice. We isolated human CD45+ cells from the spleen of mice from each treatment group and injected equivalent numbers of CD45+ cells into secondary recipients. Following 50 days, all mice treated with vehicle engrafted with CD34+CD38- positive cells. In contrast, CD34+CD38- positive cells engraftment was not detected in recipient mice (n=3) from PF-04449913-treated donors. These results demonstrate the persistent effects of PF-0449913 on long term self-renewing MDS-initiating cells. Next we performed in vitro re-differentiation of MDS-iPSCs, which express CD34+CD38- population. CD34+CD38- cells from MDS-derived iPSCs were cultured with 2 μM of PF-04449913 and 1 μM of PF-6667291 in STEMdiff APEL medium for 14 days for CFC activities. Treatments with PF-04449913 and PF-6667291 significantly reduced the colony formations of mature erythroid, granulocyte-macrophage, and mixed of these hematopoietic cells. To identify the mechanisms that limit the terminal differentiation of MDS-derived iPSC by PF-04449913 and PF-6667291, MDS-L cells were cultured with PF-04449913 and PF-6667291 for 72 hrs. The treatments with PF-04449913 and PF-6667291 induced the expressions of p21Cip1, cleaved PARP and reduced the expression of BMI-1, c-Myc, Nanog, and phospho-Stat3. Conclusion: Our preclinical results indicate that the combination with PF-04449913 and PF-6667291 have potential as an important option for controlling the terminal differentiation of MDS-initiating cells. It is expected that the combination with PF-04449913 and PF-6667291 may become extremely useful therapeutic interventions in a number of hematological neoplasms, including MDS. Disclosures Tauchi: Pfizer Inc.: Research Funding. Ohyashiki:Bristol-Myers Squibb: Research Funding; Novartis International AG,: Honoraria, Research Funding.

Isolation of Human Bone Marrow Mesenchymal Stem Cells by a Novel Monoclonal Antibody, ZUC3.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2562-2562
Author(s):  
Xiaoyu Lai ◽  
He Huang ◽  
Li Huang ◽  
Fenfang Zeng
Keyword(s):  
Stem Cells ◽  
Monoclonal Antibody ◽  
Bone Marrow ◽  
Flow Cytometry ◽  
Cell Sorting ◽  
Mononuclear Cells ◽  
Human Mscs ◽  
Bone Marrow Mononuclear Cells ◽  
Multiple Differentiation

Abstract Objective: Due to absence of a single definitive marker of mesenchymal stem cells (MSCs) and low incidence in human bone marrow, the primary culture of MSCs, conventionally isolated with its characteristic of adherent, were considered to be heterogeneous containing of several subpopulations, which had currently limited our understanding of their biology and therapeutic applications. In our previous study, a novel murine monoclonal antibody (McAb) ZUC3 was produced by hybridoma technology, which was specifically reactive with human MSCs, while showed negative cross-reactivity when screened against a variety of human tissues. Now, ZUC3 antigen positive MSCs population would be further identified by magnetic-activated cell sorting (MACS). Methods: Bone marrow were taken from the iliac crest of normal healthy adult volunteers, and mononuclear cells were separated by density gradient centrifugation, then separated into positively- and negatively-labelled fractions with McAb ZUC3 by immunomagnetic activated cell sorting. The purity of positive cells was analyzed by flow cytometry, then ZUC3 antigen positive and negative cells were plated respectively in human MSCs medium consisting of 10% FBS, LG-DMEM. Characteristics of ZUC3 antigen positive cells phenotype was analyzed by flow cytometry, and proliferation and multiple differentiation potential of the cells was observed in vitro. Results: Flow cytometric analysis showed that ZUC3 antigen expression by cultured MSCs and mononuclear cells derived from bone marrow were 91.31±2.92%, 0.96±0.28% respectively, and western blotting showed the molecular mass of antigen was about 33KD. The purity of the recovered fractions for ZUC3 by MACS was 76.82±6.32%. The positive cells have adhered to culture flask in vitro, and the quantity of adhered cells that had fibroblast-like morphology increased and proliferated during primary expansion period, while the negative cells were observed as round shape cells without any proliferation. It was demonstrated that ZUC3 antigen positive cells continued growth with spindle-shape, extending beyond 30 population doublings in long-term culture. Analyzed by flow cytometry, the culture-expanded positive cells were uniformly positive for CD29, CD44, CD105, CD106, and lack typical hematopoietic antigens such as CD14, CD34, CD45, HLA-DR, which demonstrated that ZUC3 postive cells sorted from bone marrow mononuclear cells by McAb were MSCs. With proper medium, the ZUC3 antigen positive cells could be successfully induced to differentiate into adipocytes, osteoblasts, and neuro-like cells which were positive of neuron markers such as nestin, NSE and NF-M. Conclusion: ZUC3 McAb was a specific surface marker against human MSCs for cell sorting. The ZUC3 antigen positive cells separated from bone marrow mononuclear cells had potential capacity of high proliferation and multiple differentiation.

scholarly journals Human rib bone marrow mononuclear cells spontaneously synthesize and secrete IgE in vitro

2008 ◽  
Vol 83 (1) ◽  
pp. 163-168 ◽  
Author(s):  
R. P. MACDERMOTT ◽  
G. A. S. JENDRlSAK ◽  
G. S. NASH ◽  
S. SCHREIBER ◽  
M. J. BERTOVICH ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Bone Marrow Mononuclear Cells ◽  
Human Rib Bone

Bone marrow mononuclear cells of MDS patients are characterized by in vitro proliferation and increased apoptosis independently of stromal interactions

2007 ◽  
Vol 31 (12) ◽  
pp. 1659-1667 ◽  
Author(s):  
Lambert F.R. Span ◽  
Eugenie Rutten ◽  
Anita Gemmink ◽  
Jan B.M. Boezeman ◽  
Reinier A.P. Raymakers ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Bone Marrow Mononuclear Cells ◽  
In Vitro Proliferation

The Induction of Dendritic Cells with IFN-alpha and GM-CSF from Bone Marrow Mononuclear Cells from Patients with Chronic Myeloid Leukemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4873-4873
Author(s):  
Shuier Zhen ◽  
Jie Jin ◽  
Xiangmin Tong
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
T Cell ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Chronic Phase ◽  
Myelogenous Leukemia ◽  
Bone Marrow Mononuclear Cells ◽  
Gm Csf

Abstract Chronic myelogenous leukemia (CML) is a malignant myeloproliferative disease arising from the clonal expansion of a stem cell with the typical Philadelphia (Ph) chromosome cytogenetic abnormality. IFN-a has been proven to be effective for patients in the chronic phase of myelogenous leukemia (CML), yet the mechanisms of the antitumor action of these cytokines are still a matter of debate. Dendritc cells (DCs) are potent antigen-presenting cells that prime effective T-cell response aginst tumour antigens. Recent studies have shown that IFN-a can exert a variety of effects on dendritic cells (DCs), which may play an important role in the induction of an antitumor immunity. Human DCs can be generated in vitro from peripheral blood(PB) monocytes or from CD34+ haematopoietic precursor cells in culture medium containing human granulocyte macrophage-colony stimulating factor (GM-CSF), IL-4 and some other cytokines. Previous studies have shown a new effective protocol for the generation of human DCs from unseparated BM aspirate cells with excellent functional capacity of antigen uptake and of stimulating naive and memory T cell responses superior to that of DCs from peripheral blood(PB) monocytes. We, therefore, explored whether treatment with IFN-a may influence the CML bone marrow mononuclear cells(BMMNCs) derived DCs in vitro. Treatment BMMNCs of 12 patients with CML in chronic phase with IFN-a+rhGM-CSF(IFN-a-DC) generated DCs with more mature phenotype properties expressing higher of CD80,CD86,HLA-DR,CD83 compared to the CML- BMMNCs treated with rhGM-CSF+IL-4(IL-4-DC). And in parallel with phenotypes, IFN-a-DC also showed more effective than IL-4-DC in eliciting an allogeneic mixed lymphocyte reaction by MTT assay. FISH confirmed the DCs of both groups were leukemic origin. These findings demonstrate that IFN-a promotes the differentiation/maturation of DCs derived from BMMNCs of patients with CML in vitro, these studies also broaden the clinical scope of IFN-a as a promising agent in the immunotherapy of CML.

scholarly journals Bone Marrow Mononuclear Cells Activate Angiogenesis via Gap Junction–Mediated Cell-Cell Interaction

Stroke ◽  
2020 ◽  
Vol 51 (4) ◽  
pp. 1279-1289 ◽  
Author(s):  
Akie Kikuchi-Taura ◽  
Yuka Okinaka ◽  
Yukiko Takeuchi ◽  
Yuko Ogawa ◽  
Mitsuyo Maeda ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Gap Junction ◽  
Mononuclear Cells ◽  
Cell Interaction ◽  
Bone Marrow Mononuclear Cells ◽  
Hematopoietic Stem ◽  
Direct Cell ◽  
Cell Cell

Background and Purpose— Bone marrow mononuclear cells (BM-MNCs) are a rich source of hematopoietic stem cells and have been widely used in experimental therapies for patients with ischemic diseases. Activation of angiogenesis is believed to be one of major BM-MNC mode of actions, but the essential mechanism by which BM-MNCs activate angiogenesis have hitherto been elusive. The objective of this study is to reveal the mechanism how BM-MNCs activate angiogenesis. Methods— We have evaluated the effect of direct cell-cell interaction between BM-MNC and endothelial cell on uptake of VEGF (vascular endothelial growth factor) into endothelial cells in vitro. Cerebral ischemia model was used to evaluate the effects of direct cell-cell interaction with transplanted BM-MNC on endothelial cell at ischemic tissue. Results— The uptake of VEGF into endothelial cells was increased by BM-MNC, while being inhibited by blockading the gap junction. Low-molecular-weight substance was transferred from BM-MNC into endothelial cells via gap junctions in vivo, followed by increased expression of hypoxia-inducible factor-1α and suppression of autophagy in endothelial cells. The concentration of glucose in BM-MNC cytoplasm was significantly higher than in endothelial cells, and transfer of glucose homologue from BM-MNC to endothelial cells was observed. Conclusions— Our findings demonstrated cell-cell interaction via gap junction is the prominent pathway for activation of angiogenesis at endothelial cells after ischemia and provided novel paradigm that energy source supply by stem cell to injured cell is one of the therapeutic mechanisms of cell-based therapy. Visual Overview— An online visual overview is available for this article.

Sign in / Sign up

Export Citation Format

Share Document