176 Bone marrow mononuclear cells producing IL-1 and IL-2 inhibited by HBV in vitro and in vivo

1994 ◽  
Vol 23 ◽  
pp. 129
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Bone Marrow Mononuclear Cells

Targeting the Hedgehog Signaling Pathway By PF-04449913 Limits the Self-Renewal of MDS-Derived Induced Potent Stem Cells (iPSC): Molecular Mechanisms

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 791-791
Author(s):  
Tetsuzo Tauchi ◽  
Seiichi Okabe ◽  
Seiichiro Katagiri ◽  
Yuko Tanaka ◽  
Kaoru Tohyama ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Research Funding ◽  
Molecular Mechanisms ◽  
Mononuclear Cells ◽  
Scid Mice ◽  
The Self ◽  
Bone Marrow Mononuclear Cells ◽  
Self Renewal

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. In xenograft models of human coloirectal and pancreatic cancer, treatment with PF-04449913 in combination with other anticancer agents reduced the tumor growth. Furthermore, PF-04449913 demonstrated preliminary antitumor activity in a phase I trial, when given as monotherapy in patients with several hematopoietic malignancy. In the present study, we investigated the molecular mechanisms by which PF-04449913 regulate the self-renewal of MDS-derived iPS cells (iPSCs) in vivo. Methods: We generated iPSCs from bone marrow mononuclear cells of two MDS patients (RAEB1 and RAEB2 by WHO clssification) with chromosome 5 deletion and complex karyotypic abnormalities, respectivly. 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 sucutaneously 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 establised from MDS patient with del(5q) and complex karyotypic abnormalities for in vitro studies. 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. We further examined the effects of Nanog pathway modulation on in vitro clonogenic growth. CD34+CD38- cells from MDS-derived iPSCs transferred NOD/SCID mice and MDS-L cells were treated with 2 mM of PF-04449913 for 72 hrs, washed free of drugs, and plated in quadruplicate in methylcellulose. At 14 days, colonies were counted as initial plating. The representative plate was then washed and cells were re-suspended and re-plated. After an additional 14 days, colonies were counted as secondary re-plating. Clonogenic recovery of untreated cells was normalized to 100% and plating results from all treatment groups were expressed as % control. PF-04449913 had only minimum effects on colony formation after initial plating over control cells. However, upon serial re-plating, secondary colony formations were significantly inhibited by PF-04449913 (p<0.001). To identify the mechanisms that limit the self-renewal of MDS-initiating ells by PF-04449913, NOD/SCID mice engrafted with CD34+CD38- fractions from MDS-derived iPSCs were treated with PF-04449913 (100 mg/kg; p.o.) for 14 days. PF-04449913 induced the expressions of p21Cip1, cleaved PARP and reduced the expression of BMI-1, c-Myc, Nanog, and Bcl-XL. Conclusion: Our preclinical results indicate that PF-04449913 have potential as an important option for controlling the drug-resistant MDS-initiating cells. It is expected that the PF-04449913 may become extremely useful therapeutic interventions in a number of hematological neoplasms, including MDS, where the persistence of cancer stem cells. Disclosures Ohyashiki: Sumitomo Dainippon: Membership on an entity's Board of Directors or advisory committees; Chugai Pharna KK: Research Funding; Bristol Meyer Squib KK: Research Funding; Jansen Pharma KK: Honoraria, Research Funding, Speakers Bureau; Celegen KK: Consultancy, Honoraria, Research Funding, Speakers Bureau; Novartis Pharma KK: Honoraria, Research Funding, Speakers Bureau; Kyowa Kirin KK: Honoraria; MSD KK: Honoraria; Nippo Shinyaku KK: Speakers Bureau; Toyama Kagaku KK: Speakers Bureau; Shinbaio Pharma KK: Honoraria; Asteras: Research Funding; Alexion Pharma KK: Research Funding; Teijin Pharma KK: Research Funding; Asahikasei: Research Funding; Taiho Yakuhin KK: Research Funding.

scholarly journals Sustained, retransplantable, multilineage engraftment of highly purified adult human bone marrow stem cells in vivo

Blood ◽  
1996 ◽  
Vol 88 (11) ◽  
pp. 4102-4109 ◽  
Author(s):  
CI Civin ◽  
G Almeida-Porada ◽  
MJ Lee ◽  
J Olweus ◽  
LW Terstappen ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Cell Population ◽  
Mononuclear Cells ◽  
Cell Subset ◽  
Fetal Sheep ◽  
Adult Human

Abstract Data from many laboratory and clinical investigations indicate that CD34+ cells comprise approximately 1% of human bone marrow (BM) mononuclear cells, including the progenitor cells of all the lymphohematopoietic lineages and lymphohematopoietic stem cells (stem cells). Because stem cells are an important but rare cell type in the CD34+ cell population, investigators have subdivided the CD34+ cell population to further enrich stem cells. The CD34+/CD38-cell subset comprises less than 10% of human CD34+ adult BM cells (equivalent to < 0.1% of marrow mononuclear cells), lacks lineage (lin) antigens, contains cells with in vitro replating capacity, and is predicted to be highly enriched for stem cells. The present investigation tested whether the CD34+/CD38-subset of adult human marrow generates human hematopoiesis after transfer to preimmune fetal sheep. CD34+/ CD38- cells purified from marrow using immunomagnetic microspheres or fluorescence-activated cell sorting generated easily detectable, long- term, multilineage human hematopoiesis in the human-fetal sheep in vivo model. In contrast, transfer of CD34+/CD38+ cells to preimmune fetal sheep generated only short-term human hematopoiesis, possibly suggesting that the CD34+/CD38+ cell population contains relatively early multipotent hematopoletic progenitor cells, but not stem cells. This work extends the prior in vitro evidence that the earliest cells in fetal and adult human marrow lack CD38 expression. In summary, the CD34+/ CD38-cell population has a high capacity for long-term multilineage hematopoietic engraftment, suggesting the presence of stem cells in this minor adult human marrow cell subset.

Donor Origin of Multipotent Adult Progenitor Cells in Radiation Chimeras.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1395-1395
Author(s):  
Morayma Reyes ◽  
Jeffrey S. Chamberlain
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Progenitor Cells ◽  
Y Chromosome ◽  
Stromal Cells ◽  
Mononuclear Cells ◽  
Multipotent Adult Progenitor Cells

Abstract Multipotent Adult Progenitor Cells (MAPC) are bone marrow derived stem cells that can be extensively expanded in vitro and can differentiate in vivo and in vitro into cells of all three germinal layers: ectoderm, mesoderm, endoderm. The origin of MAPC within bone marrow (BM) is unknown. MAPC are believed to be derived from the BM stroma compartment as they are isolated within the adherent cell component. Numerous studies of bone marrow chimeras in human and mouse point to a host origin of bone marrow stromal cells, including mesenchymal stem cells. We report here that following syngeneic bone marrow transplants into lethally irradiated C57Bl/6 mice, MAPC are of donor origin. When MAPC were isolated from BM chimeras (n=12, 4–12 weeks post-syngeneic BM transplant from a transgenic mouse ubiquitously expressing GFP), a mixture of large and small GFP-positive and GFP-negative cells were seen early in culture. While the large cells stained positive for stroma cell markers (smooth muscle actin), mesenchymal stem cell makers (CD73, CD105, CD44) or macrophages (CD45, CD14), the small cells were negative for all these markers and after 30 cell doublings, these cells displayed the classical phenotype of MAPC (CD45−,CD105−, CD44−, CD73−, FLK-1+(vascular endothelial growth factor receptor 2, VEGFR2), Sca-1+,CD13+). In a second experiment, BM obtained one month post BM transplant (n=3) was harvested and mononuclear cells were sorted as GFP-positive and GFP-negative cells and were cultured in MAPC expansion medium. MAPC grew from the GFP-positive fraction. These GFP positive cells displayed the typical MAPC-like immunophenotypes, displayed a normal diploid karyotype and were expanded for more than 50 cell doublings and differentiated into endothelial cells, hepatocytes and neurons. To rule out the possibility that MAPC are the product of cell fusion between a host and a donor cell either in vivo or in our in vitro culture conditions, we performed sex mismatched transplants of female GFP donor BM cells into a male host. BM from 5 chimeras were harvested 4 weeks after transplant and MAPC cultures were established. MAPC colonies were then sorted as GFP-positive and GFP- negative and analyzed for the presence of Y-chromosome by FISH analysis. As expected all GFP-negative (host cells) contained the Y-chromosome whereas all GFP-positive cells (donor cells) were negative for the Y-chromosome by FISH. This proves that MAPC are not derived from an in vitro or in vivo fusion event. In a third study, BM mononuclear cells from mice that had been previously BM-transplanted with syngeneic GFP-positive donors (n=3) were transplanted into a second set of syngeneic recipients (n=9). Two months after the second transplant, BM was harvested and mononuclear cells were cultured in MAPC medium. The secondary recipients also contained GFP-positive MAPC. This is the first demonstration that BM transplantation leads to the transfer of cells that upon isolation in vitro generate MAPCs and, whatever the identity of this cell may be, is eliminated by irradiation. We believe this is an important observation as MAPC hold great clinical potential for stem cell and/or gene therapy and, thus, BM transplant may serve as a way to deliver and reconstitute the MAPC population. In addition, this study provides insight into the nature of MAPC. The capacity to be transplantable within unfractionated BM transplant renders a functional and physiological distinction between MAPC and BM stromal cells. This study validates the use of unfractionated BM transplants to study the nature and possible in vivo role of MAPC in the BM.

p38MAPK Inhibitor LSN2322600 Modulates the Bone Marrow Microenvironment and Inhibits Osteoclastogenesis in Multiple Myeloma.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5042-5042
Author(s):  
Kenji Ishitsuka ◽  
Teru Hideshima ◽  
Paola Neri ◽  
Sonia Vallet ◽  
Norihiko Shiraishi ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Stromal Cells ◽  
Bone Marrow Stromal Cells ◽  
Mononuclear Cells ◽  
Severe Combined Immunodeficiency ◽  
Marrow Stromal Cells ◽  
Skeletal Complications

Abstract The interaction between multiple myeloma (MM) cells and the bone marrow (BM) microenvironment plays a crucial role not only in proliferation and survival of MM cells, but also in osteoclastogenesis. In this study, we examined diverse potential of novel p38MAPK inhibitor LSN2322600 (LSN) for MM therapy in vitro and in vivo. The cytotoxic activity of LSN against MM cell lines was modest; however, LSN significantly enhances the cytotoxicity of Bortezomib by down-regulating Bortezomib-induced heat shock protein (HSP) 27 phosphorylation. We next examined the effects of LSN on cytokine secretion in MM cells, bone marrow stromal cells and osteoclast precursor cells. LSN inhibited IL-6 secretion from long-term cultured-bone marrow stromal cells (LT-BMSCs) and bone marrow mononuclear cells (BMMNCs) from MM patients in remission. LSN also inhibited MIP-1 α secretion by fresh tumor cells, BMMNCs and CD14 positive cells. Since these cytokines mediate osteoclastogenesis, we further examined whether LSN could inhibit osteoclastogenesis. Importantly, LSN inhibited in vitro osteoclastogenesis induced by macrophage-colony stimulating factor (M-CSF) and soluble receptor activator of nuclear factor- κ B ligand (sRANKL), as well as osteoclastogenesis in the severe combined immunodeficiency (SCID)-Hu mouse model of human MM. These results suggest that LSN represents a promising novel targeted strategy to reduce skeletal complications as well as to sensitize or overcome resistance to Bortezomib.

SGX70393 Inhibits Bcr-AblT315I In Vitro and In Vivo and Completely Suppresses Resistance When Combined with Nilotinib or Dasatinib.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 535-535 ◽  
Author(s):  
Thomas O’Hare ◽  
Christopher A. Eide ◽  
Jeffrey W. Tyner ◽  
Amie S. Corbin ◽  
Matthew J. Wong ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mononuclear Cells ◽  
Kinase Inhibitor ◽  
Single Agent ◽  
Resistance Profile ◽  
Murine Bone Marrow ◽  
Abl Kinase ◽  
A Cell

Abstract Overview: Bcr-AblT315I is detected in the majority of CML patients who relapse after dasatinib- or nilotinib-based second-line Bcr-Abl kinase inhibitor therapy. SGX70393, an azapyridine-based Abl kinase inhibitor, is effective against Bcr-Abl and Bcr-AblT315I at low nanomolar concentrations in vitro and in cell lines. Here, we comprehensively profiled SGX70393 against native and mutant Bcr-Abl in vitro and in vivo. We also used a cell-based mutagenesis screen to evaluate the resistance profile of SGX70393 alone and in combination with imatinib, nilotinib, or dasatinib. Methods: We assessed colony formation in the presence of SGX70393 by murine bone marrow infected with retroviruses for expression of Bcr-Abl, Bcr-AblT315I, or a variety of other mutants. Toxicity was tested in clonogenic assays of normal bone marrow. SGX70393 effects on cellular tyrosine phosphorylation were measured by immunoblot and FACS in primary Bcr-AblT315I cells isolated from patients with CML or Ph+ B-ALL. In vivo activity was evaluated in a xenograft model using Ba/F3 cells expressing Bcr-AblT315I. Lastly, the resistance profile of SGX70393 was evaluated alone and in dual combinations with imatinib, nilotinib, or dasatinib in a cell-based mutagenesis assay. Results: Colony formation by murine bone marrow cells expressing Bcr-AblT315I (IC50: 180 nM) was reduced by SGX70393 in a dose dependent manner, while no toxicity was observed in colony forming assays of normal human or murine mononuclear cells at concentrations up to 2 μM. Ex vivo exposure of human Bcr-AblT315I mononuclear cells to SGX70393 decreased CrkL phosphorylation, while imatinib, nilotinib, or dasatinib had no effect. SGX70393 inhibited Bcr-AblT315I-driven tumor growth in mice and this was correlated with reduced levels of pCrkL in tumor tissue, while imatinib was ineffective. A cell-based mutagenesis screen revealed a profile of resistant clones confined to four p-loop residues and position 317. SGX70393 in combination with imatinib contracted the spectrum of resistant mutations relative to either single agent, though outgrowth could not be completely suppressed. Combining SGX70393 with low concentrations of nilotinib or dasatinib narrowed the resistance profile still further (residues 248 and 255 for nilotinib; 317 for dasatinib) and, with clinically achievable doses of either second drug, completely abrogated emergence of resistant subclones. Conclusions: SGX70393, a potent inhibitor of Bcr-AblT315I, exhibits a resistance profile centered around the p-loop and residue 317 of Bcr-Abl. Remarkably, in combination with nilotinib or dasatinib, outgrowth of resistant clones is completely suppressed. Single-agent therapy with an effective T315I inhibitor may provide a viable option for patients who relapse with Bcr-AblT315I. However, as a broader spectrum of mutations accounts for imatinib resistance, patients with acquired dasatinib or nilotinib resistance may continue to harbor residual mutant clones other than T315I. Thus, the full clinical potential of SGX70393 may be realized in combinations with a second Abl kinase inhibitor. Our findings provide the first demonstration that Abl kinase inhibitor combinations that include a T315I-targeted component such as SGX70393 have the potential to pre-empt Bcr-Abl-dependent resistance.

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.

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