Characterization of Angiogenic Potential of Late Endothelial Progenitor Cells on Human Cord Blood and Bone Marrow

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5404-5404
Author(s):  
Eun-Sun Yoo ◽  
Jee-Young Ahn ◽  
KiHwan Kwon ◽  
Soo-Ah Oh ◽  
Moon-Young Choi ◽  
...  
Keyword(s):  
Cord Blood ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Adhesion Molecule ◽  
Mononuclear Cells ◽  
Marker Genes ◽  
Endothelial Progenitor ◽  
Angiogenic Potential ◽  
Cell Based Therapy

Abstract Background: The identification of circulating endothelial progenitor cells (EPCs) has revolutionized approaches to cell-based therapy for injured and ischemic tissues. Recently, we have demonstrated that there are 2 distinct types of EPCs from UCB having different biologic properties for angiogenic capabilities in vitro and in vivo. In present study, the aim is to directly compare umbilical cord blood (UCB)- and BM-derived late EPC surface phenotypes and in vitro functional capacity. Methods: Mononuclear cells from UCB and BM cultured using EGM-2 medium with VEGF, IGF-1 and FGF for 21 days. Late outgrowing endothelail cells(late OECs) which were in peak growth at third weeks of culture were analyzed for expression of various surface markers by flow cytometry/RT-PCR/IF, tube formation in Matrigel plates, proliferation assay, endothelial colony assay and the role of SDF-1/VEGF on migration. Results: The adherent cells after culture of 7 days exhibited a fibroblast like shape in BM and a cobblestone shaped cells in UCB. Although two sources of OECs were comparable in expression of endothelial and various adhesion molecule markers, BM-derived OECs contained higher proportion of cells expressing smooth muscle cell markers(SMMHC), several adhesion molecule(CD49d, CD62L and VCAM-1), whereas the expression of CXCR-4, PECAM-1 and CD62E and expression of mRNA on endothelial marker genes were higher in UCB-derived OECs. UCB-OECS stained positive for uptake of acetylated low-density lipoprotein and had more migratory ability in the presence of SDF-1 and VEGF compared with BM-OECs. Both sources OECs effectively formed capillary tubes in Matrigel plates. Conclusion: We directly compared OECs derived from UCB and BM and two source of OECs differ in aspect of several adhesion molecule and angiogenic potential in vitro. These difference of UCB render it potentially advantageous for human therapeutic OECs applications for potential applications for a “cell therapy” in the situations on vascular injuries when compared with patients-derived BM.

scholarly journals Interleukin-1β Augments Angiogenic Responses of Murine Endothelial Progenitor Cells in Vitro

2009 ◽  
Vol 29 (5) ◽  
pp. 933-943 ◽  
Author(s):  
Anna Rosell ◽  
Ken Arai ◽  
Josephine Lok ◽  
Tongrong He ◽  
Shuzhen Guo ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Mononuclear Cells ◽  
Therapeutic Angiogenesis ◽  
Interleukin 1Β ◽  
Endothelial Progenitor ◽  
Angiogenic Potential ◽  
Diphenyl Tetrazolium Bromide ◽  
In Vitro Angiogenesis

Endothelial progenitor cells (EPCs) may provide novel opportunities for therapeutic angiogenesis after ischemic diseases. However, it is unclear how the angiogenic potential of EPCs might be affected by an inflammatory environment. We examine how the potent cytokine interleukin-1β (IL-1β) affects angiovasculogenic responses in EPCs in culture. Mononuclear cells isolated from mouse spleen were plated on fibronectin-coated wells and grown in EGM-2 MV media. Endothelial progenitor cells were phenotyped using multiple markers (UEA-Lectin, ac-LDL, CD133, CD34, vWillebrand Factor, Flk-1) and to identify the IL-1 Receptor-I. We quantified cell and colony counts and performed MTT (3-(4,5-dimethylthiazol-2-yl)2,5-diphenyl-tetrazolium bromide) and Matrigel assays, in vitro, under control and IL-1β (10 ng/mL) conditions. Endothelial progenitor cells exposed to IL-1β increased in the number of cells and colonies compared with untreated cells, without any effect on cell metabolic integrity. Furthermore, IL-1β treatment augmented EPC angiogenic function, significantly increasing the number of vessel-like structures in the Matrigel assay. An early phosphorylation of ERK1/2 occurred after IL-1β stimulation, and this pathway was inhibited if IL-1 Receptor-I was blocked. Our results suggest that IL-1β is a potent stimulator of in vitro angiogenesis through ERK signaling in mouse EPCs. Further studies are warranted to assess how interactions between proinflammatory environments and EPC responses may be leveraged to enhance therapeutic angiogenesis.

Abstract 1743: Stem/Progenitor Markers Sca-1 versus C-kit and CD31 Determine Angiogenic Potential of Mouse Bone Marrow-derived Endothelial Progenitor Cells

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Toshikazu D Tanaka ◽  
Masaaki Ii ◽  
Haruki Sekiguchi ◽  
Kentaro Jujo ◽  
Sol Misener ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Mononuclear Cells ◽  
Tube Formation ◽  
Mouse Bone Marrow ◽  
Endothelial Progenitor ◽  
Angiogenic Potential ◽  
Ischemic Tissue

Background: Endothelial progenitor cells (EPCs) have been shown to have angiogenic potential contributing to neovascularization. However, the definition of EPC, including surface marker expression of EPCs promoting vasculo-/angiogenesis in ischemic tissue, remains uncertain. We hypothesized that stem/progenitor (c-kit vs. sca-1) and endothelial cell (EC) markers (CD31) may identify cells with enhanced EPC potential. Methods and Results: Mononuclear cells (MNCs) were isolated from mouse bones, and Lin+ cells were depleted by magnetic cell sorting. Lin- cells were further sorted with the following markers (% of total MNCs) by FACS: c-kit+ (1.87%), sca-1+ (0.6%), c-kit+ /CD31+ (1.1%) and sca-1+ /CD31+ (0.28%). Non-sorted MNCs were used as a control. To examine EC phenotype in culture, cells were labeled with DiI and co-cultured with mature ECs (human microvascular endothelial cells: HMVECs). The percent incorporation of DiI labeled cells into HMVEC tube structures 12 hours after co-culture and BS1-lectin positivity/acLDL uptake were: sca-1+ /CD31+ cells (87 ± 2%) > c-kit+ /CD31+ (79 ± 8%) > sca-1+ (62 ± 8%) > c-kit+ (59 ± 5%) > MNC (50 ± 3% ) . Next, we examined homing capacity of these cells to ischemic myocardium using a mouse myocardial infarction (MI) model. DiI-labeled cells (5x10 4 , IV) were injected to splenectomized mice 3 days after MI, and the hearts were excised 24 hours after the cell injection for histological analysis. Interestingly, the number of recruited/retained DiI-labeled-cells in the MI hearts exactly replicated the findings of the in vitro tube formation assay (cells/HPF): sca-1+ /CD31+ (108 ± 26) > c-kit+ /CD31+ (77 ± 16) > sca-1+ (71 ± 14) > c-kit+ (67 ± 1) > MNCs (48 ± 6) , suggesting that sca-1+ /CD31+ cells might have great functional activities as endothelial precursors. Conclusions: Both stem/progenitor marker Sca-1 and EC marker CD31 expressing EPCs exhibited high potential angiogenic capacity with EC phenotypic features compared with c-kit expressing cells. Our data suggest that Sca-1+ /CD31+ cells may represent EPC-rich cell population, and Sca-1/CD31 could be useful markers to enrich for cells with EPC potential. Ongoing studies will determine the in vivo characteristics of these cells for ischemic tissue repair.

Abstract 3955: Chemokines Fused to a Fractalkine Backbone and a GPI-Anchor Attract Embryonic Endothelial Progenitor Cells to the Site of Ischemia

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Chiraz El-Aouni ◽  
Franziska Globisch ◽  
Achim Pfosser ◽  
Georg Stachel ◽  
Rabea Hinkel ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Adhesion Molecule ◽  
Capillary Density ◽  
Gpi Anchor ◽  
Endothelial Progenitor ◽  
Collateral Growth ◽  
Artificial Adhesion

Recruitment of endothelial progenitor cells to the sites of ischemia is a prerequisite for efficient therapeutic neovascularization via vasculogenesis. Chemokines play a major role in the homing of EPCs at the ischemic vasculature, a mechanism fading in chronic ischemia. To overcome this limitation, we constructed an artificial adhesion molecule consisting of a GPI-anchor, a fractalkine-backbone and an SDF-1 head (SDF-1-fra-GPI), which was applied for enhanced recruitment of embryonic EPCs (eEPCs: CXCR4++, Tie2++, Thrombomodulin++, CD34-, MHCI-, vWF inducible, eNOS inducible) in vitro and in vivo . Methods: In a flow chamber adhesion assay, Control plasmids (pcDNA or GPI-SDF-1 cDNA) were compared to the SDF-1-fra-GPI construct for eEPC recruitment 24h after liposomal transfection of rat endothelial cells. In vivo, in rabbits (n=5 per group) at day 7 (d7) after femoral artery excision, 1 mg of the SDF-1-fra-GPI or eGFP cDNA was transfected into the ischemic limb. At d9, ischemic hindlimbs were retroinfused with 5x10 6 eEPCs. Angiography was performed for collateral quantification and frame count score at d9 and d37 (% of d9), capillary density was assessed via PECAM-1-staining (capillaries/muscle fiber = c/mf). Results: In vitro, eEPC adhesion (16±12 cells/field) was increased to a higher extent by SDF-1-fra-GPI (79±13) than SDF1-GPI (54±8) or control vector (37±8). In vivo , eEPC adhesion in the ischemic hindlimb after SDF-1-fra-GPI transfection compared to mock transfection (30±3 vs. 9±1 cells/field). Whereas capillary density was unaffected (1.66±0.30 SDF-1-Fra-GPI vs. 1.56±0.29 eEPCs), collateral growth (152±10% SDF-1-fra-GPI vs. 124±13%) as well as perfusion score (198±17% SDF-1-fra-GPI vs.160±6% eEPCs) further increased after SDF-1-fra-GPI transfection (controls: 1.24±0.12 c/mf, collaterals 105±8%, perfusion score 112±11%). We conclude that recruitment of EPCs expressing CXCR4 (the SDF-1 receptor) may benefit from pre-treatment of the recipient vasculature with SDF-1-Fra-GPI, an artificial adhesion molecule. This approach might be valuable for enhancing EPC recruitment in the scenario of therapeutic neovascular-ization of chronic ischemic syndromes.

Circulating Endothelial Progenitor Cells Are Increased in Patients with Myelofibrosis and Do Not Harbor V617F JAK-2 or W515L MPL Mutations

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1535-1535 ◽  
Author(s):  
Elisa Bonetti ◽  
Vittorio Rosti ◽  
Laura Villani ◽  
Rita Campanelli ◽  
Gaetano Bergamaschi ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Healthy Subjects ◽  
Mononuclear Cells ◽  
Median Frequency ◽  
Endothelial Progenitor ◽  
Circulating Endothelial Progenitor Cells ◽  
Mutational Status

Abstract Bone marrow and spleen neoangiogenesis is a relevant feature of patients with myelofibrosis (MF). We have previously reported that patients with MF have an increased percentage of circulating endothelial progenitor cells (EPC) assessed as CD34+CD133+VEGFR2+ cells compared with patients with other Ph-negative myeloproliferative disorders (polycythemia vera, PV, and essential thrombocytemia, ET) and healthy subjects. However, neither the functional activity of these putative EPC nor their belonging to the malignant clone have been yet fully characterized. In order to address these issues we have grown in vitro EPC-derived colonies from the peripheral blood (PB) of 36 patients with MF, 9 patients with PV or ET and 10 healthy subjects. Seventeen MF patients harbored a V617F JAK-2 mutation (8 heterozygous and 9 homozygous) whereas 2 patients showed a W515L MPL mutation (both heterozygous). Eight out of 9 PV/ET patients had a V617F JAK-2 mutation (5 heterozygous and 3 homozygous). Mononuclear cells were cultured in collagen coated 6 well plates in the presence of EBM-2MV medium according to Ingram et al (Blood104:2752; 2004). The endothelial origin of the colonies was ascertained by assessment of the expression of CD105, CD146, CD144, CD31, vWf, VEGFR-2, CD14 and CD45 antigens. V617F JAK-2 and W515L MPL mutations were assessed by PCR, followed by enzymatic digestion, of endothelial cells after tripsinization of the EPC-derived colonies. The median frequency (number of colonies per 107 mononuclear cells plated) of EPC-derived colonies was statistically higher in MF patients (0.25, range 0–8.1) compared to healthy subjects (0.05, 0–0.3; P=0.037), but not different form that of PV/ET patients (0, 0–4.4; P=NS). Immunophenotyping confirmed that the cells expressed the endothelial antigens CD105, CD146, CD144, CD31, vWf, and VEGFR-2 but not the hematopoietic specific antigens CD45 and CD14. The capacity of colony-derived endothelial cells of MF patients to form capillary-like structures in the Matrigel assay was not different from that of healthy subjects. No correlation was found between the number of colonies and the mutational status of either JAK-2 or MPL. In 11 MF patients harboring either a JAK-2 (n=9) or a MPL (n=2) mutation, colony growth was observed and PCR was performed on EPC-derived colonies. In 0/9 and 0/2 cases neither JAK-2 nor MPL mutations were found, respectively. In addition, no V617F JAK-2 mutation was found in the EPC-derived colonies of 8 PV/ET patients who carried the mutation in their granulocytes. Taken together, our data show that patients with MF have an increased frequency of EPC in their PB compared to healthy subjects and that these mobilized EPC are not clonally-related to the JAK-2 or MPL mutated clone. Whether or not circulating EPC derive from an earlier progenitor cell compared to the one in which the JAK-2/MPL mutations arise remains to be determined.

P3475Dabigatran inhibits the activation of endothelial progenitor cells induced by thrombin

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
S Papadaki ◽  
S Sidiropoulou ◽  
I Moschonas ◽  
A Tselepis
Keyword(s):  
Endothelial Cells ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Adhesion Molecule ◽  
Mononuclear Cells ◽  
Umbilical Vein ◽  
Cell Types ◽  
The European Union ◽  
Membrane Expression ◽  
Endothelial Progenitor

Abstract Introduction Thrombin is a coagulation serine protease, which also activates various cell types, including endothelial cells. Late-outgrowth endothelial cells (OECs) are a type of endothelial progenitor cells that contribute to endothelial regeneration and angiogenesis. Dabigatran is a direct oral anticoagulant that inhibits thrombin's action and is widely used in everyday clinical practice. Purpose We investigated the effect of dabigatran on thrombin-induced activation of OECs, using the membrane expression of adhesion molecule ICAM-1 as activation marker. Control experiments were conducted in mature human umbilical vein endothelial cells (HUVECs). Methods CD34+ cells were isolated from cord blood mononuclear cells, using human CD34 Microbead Kit and appropriately cultured for OEC formation. HUVECs were purchased from Lonza. Confluent OECs and HUVECs were incubated with 10 or 20 μM dabigatran for 10 min, before activation with 8 U/mL thrombin, for 24 h. The effect of dabigatran on ICAM-1 expression (anti-CD54-PE) was evaluated in both cell types as % gated CD31+/CD54+ cells, using flow cytometry. Results Thrombin induced ICAM-1 membrane expression by 33±15% on OECs and by 123±22% on HUVECs, compared with respective untreated cells (p<0.05 for both comparisons, from 6 different experiments). Dabigatran at 10 μM significantly inhibited ICAM-1 expression by 36±7% on OECs and 57±8% on HUVECs, as well as at 20 μM by 51±8% on OECs and 79±7% on HUVECs (p<0.05 for all comparisons, from 4 different experiments). Conclusions We show for the first time that dabigatran at concentrations relevant to those existing in vivo after oral administration, inhibits the thrombin-induced membrane expression of the adhesion molecule ICAM-1 on OECs, a phenomenon that is also observed on mature endothelial cells. The significance of this effect regarding the pathophysiological role of OECs and mature endothelial cells at the clinical level remains to be established. Acknowledgement/Funding Operational Program “Human Resources Development, Education and Lifelong Learning”. Co-financed by the European Union and Greek national funds

scholarly journals MRI Tracking of FePro Labeled Fresh and Cryopreserved Long Term In Vitro Expanded Human Cord Blood AC133+ Endothelial Progenitor Cells in Rat Glioma

PLoS ONE ◽  
2012 ◽  
Vol 7 (5) ◽  
pp. e37577 ◽  
Author(s):  
Branislava Janic ◽  
Kourosh Jafari-Khouzani ◽  
Abbas Babajani-Feremi ◽  
A. S. M. Iskander ◽  
Nadimpalli Ravi S. Varma ◽  
...  
Keyword(s):  
Cord Blood ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Endothelial Progenitor ◽  
Human Cord Blood ◽  
Rat Glioma

scholarly journals Reduced Ischemic Brain Injury by Partial Rejuvenation of Bone Marrow Cells in Aged Rats

2010 ◽  
Vol 31 (3) ◽  
pp. 855-867 ◽  
Author(s):  
Akihiko Taguchi ◽  
Pengxiang Zhu ◽  
Fang Cao ◽  
Akie Kikuchi-Taura ◽  
Yukiko Kasahara ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Mononuclear Cells ◽  
Bone Marrow Cells ◽  
Aged Rats ◽  
Endothelial Progenitor ◽  
Ischemic Brain ◽  
Marrow Cells

Circulating bone marrow-derived immature cells, including endothelial progenitor cells, have been implicated in homeostasis of the microvasculature. Decreased levels of circulating endothelial progenitor cells, associated with aging and/or cardiovascular risk factors, correlate with poor clinical outcomes in a range of cardiovascular diseases. Herein, we transplanted bone marrow cells from young stroke-prone spontaneously hypertensive rats (SHR-SP) into aged SHR-SP, the latter not exposed to radiation or chemotherapy. Analysis of recipient peripheral blood 28 days after transplantation revealed that 5% of circulating blood cells were of donor origin. Cerebral infarction was induced on day 30 posttransplantation. Animals transplanted with bone marrow from young SHR-SP displayed an increase in density of the microvasculature in the periinfarction zone, reduced ischemic brain damage and improved neurologic function. In vitro analysis revealed enhanced activation of endothelial nitric oxide synthase and reduced activation p38 microtubule-associated protein (MAP) kinase, the latter associated with endothelial apoptosis, in cultures exposed to bone marrow-derived mononuclear cells from young animals versus cells from aged counterparts. Our findings indicate that partial rejuvenation of bone marrow from aged rats with cells from young animals enhances the response to ischemic injury, potentially at the level of endothelial/vascular activation, providing insight into a novel approach ameliorate chronic vascular diseases.

Nrf2 deficiency diminishes angiogenic potential of endothelial progenitor cells in vitro but improves neovascularization under ischemic conditions in vivo

2012 ◽  
Vol 56 (5-6) ◽  
pp. 317
Author(s):  
Urszula Florczyk ◽  
Agnieszka Jazwa ◽  
Monika Maleszewska ◽  
Szymon Czauderna ◽  
Anna Grochot-Przeczek ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Endothelial Progenitor Cells ◽  
Endothelial Progenitor ◽  
Angiogenic Potential
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