Multistep Nature of Microvascular Recruitment of Ex Vivo–expanded Embryonic Endothelial Progenitor Cells during Tumor Angiogenesis

Tissue neovascularization involves recruitment of circulating endothelial progenitor cells that originate in the bone marrow. Here, we show that a class of embryonic endothelial progenitor cells (Tie-2+, c-Kit+, Sca-1+, and Flk-1−/low), which were isolated at E7.5 of mouse development at the onset of vasculogenesis, retain their ability to contribute to tumor angiogenesis in the adult. Using intravital fluorescence videomicroscopy, we further defined the multistep process of embryonic endothelial progenitor cell (eEPC) homing and incorporation. Circulating eEPCs are specifically arrested in “hot spots” within the tumor microvasculature, extravasate into the interstitium, form multicellular clusters, and incorporate into functional vascular networks. Expression analysis and in vivo blocking experiments provide evidence that the initial cell arrest of eEPC homing is mediated by E- and P-selectin and P-selectin glycoprotein ligand 1. This paper provides the first in vivo insights into the mechanisms of endothelial progenitor cell recruitment and, thus, indicates novel ways to interfere with pathological neovascularization.

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Effects of androgens on endothelial progenitor cells in vitro and in vivo

Clinical Science ◽

10.1042/cs20090077 ◽

2009 ◽

Vol 117 (10) ◽

pp. 355-364 ◽

Cited By ~ 22

Author(s):

Gian Paolo Fadini ◽

Mattia Albiero ◽

Andrea Cignarella ◽

Chiara Bolego ◽

Christian Pinna ◽

...

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Endothelial Progenitor Cell ◽

Cell Biology ◽

Progenitor Cell ◽

Adult Males ◽

Endothelial Progenitor ◽

Healthy Human

The beneficial or detrimental effects of androgens on the cardiovascular system are debated. Endothelial progenitor cells are bone-marrow-derived cells involved in endothelial healing and angiogenesis, which promote cardiovascular health. Oestrogens are potent stimulators of endothelial progenitor cells, and previous findings have indicated that androgens may improve the biology of these cells as well. In the present study, we show that testosterone and its active metabolite dihydrotestosterone exert no effects on the expansion and function of late endothelial progenitors isolated from the peripheral blood of healthy human adult males, whereas they positively modulate early ‘monocytic’ endothelial progenitor cells. In parallel, we show that castration in rats is followed by a decrease in circulating endothelial progenitor cells, but that testosterone and dihydrotestosterone replacement fails to restore endothelial progenitor cells towards normal levels. This is associated with persistently low oestrogen levels after androgen replacement in castrated rats. In a sample of 62 healthy middle-aged men, we show that circulating endothelial progenitor cell levels are more directly associated with oestradiol, rather than with testosterone, concentrations. In conclusion, our results collectively demonstrate that androgens exert no direct effects on endothelial progenitor cell biology in vitro and in vivo.

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Does erythropoietin therapy affect circulating endothelial cells in hemodialysis patients?

Ukrainian Journal of Nephrology and Dialysis ◽

10.31450/ukrjnd.4(68).2020.05 ◽

2020 ◽

pp. 29-37

Author(s):

T. Bulduk ◽

A. U. Yalcin ◽

O. M. Akay ◽

S. G. Ozkurt ◽

H. U. Teke ◽

...

Keyword(s):

Endothelial Cells ◽

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Endothelial Progenitor Cell ◽

Peripheral Blood ◽

Progenitor Cell ◽

Hemodialysis Patients ◽

Circulating Endothelial Cells ◽

Control Group ◽

Endothelial Progenitor

Anemia is a common complication of chronic kidney disease (CKD). The most common cause of anemia in CKD is erythropoietin deficiency; and the most important cause of mortality in CKD patients is atherosclerotic vascular complications which are associated with endothelial damage. One of the methods evaluating vascular integrity is the cytometric measurement of circulating endothelial cells and endothelial progenitor cells in peripheral blood. The study aimed to investigate the effects of erythropoietin therapy on endothelial dysfunction by evaluating circulating endothelial cells and endothelial progenitor cells in peripheral blood using the technique of flow cytometry. Methods. A total of 55 hemodialysis patients were evaluated in three groups; those having erythropoietin therapy for at least last 3 months (n = 20) / not having erythropoietin for at least the last 3 months (n = 20) and the patients who started erythropoietin treatment during the study (n = 5). The control group consisted of 20 people. Blood values of the 3rd Group were investigated three times as baseline, 2nd week and 8th week CD34 +, CD105 + cells were evaluated as activated circulating endothelial cells; CD133 +, CD146 + cells were evaluated as activated endothelial progenitor cells. Results. There was no difference between the patients and healthy individuals in terms of circulating endothelial cells and endothelial progenitor cells. In the third group, no differences were observed in circulating endothelial cells / endothelial progenitor cell levels at baseline / 2nd and 8th weeks. There was no correlation between erythropoietin and circulating endothelial cells / endothelial progenitor cells. Conclusion. A correlation is not available between the therapeutic doses of erythropoietin used in hemodialysis patients and circulating endothelial cells / endothelial progenitor cell levels; supratherapeutic doses could change the results.

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Humanized large-scale expanded endothelial colony–forming cells function in vitro and in vivo

Blood ◽

10.1182/blood-2008-09-181362 ◽

2009 ◽

Vol 113 (26) ◽

pp. 6716-6725 ◽

Cited By ~ 138

Author(s):

Andreas Reinisch ◽

Nicole A. Hofmann ◽

Anna C. Obenauf ◽

Karl Kashofer ◽

Eva Rohde ◽

...

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Large Scale ◽

Proliferative Potential ◽

Vascular Networks ◽

Endothelial Progenitor ◽

Endothelial Colony Forming Cells ◽

Promising Tool

Abstract Endothelial progenitor cells are critically involved in essential biologic processes, such as vascular homeostasis, regeneration, and tumor angiogenesis. Endothelial colony–forming cells (ECFCs) are endothelial progenitor cells with robust proliferative potential. Their profound vessel-forming capacity makes them a promising tool for innovative experimental, diagnostic, and therapeutic strategies. Efficient and safe methods for their isolation and expansion are presently lacking. Based on the previously established efficacy of animal serum–free large-scale clinical-grade propagation of mesenchymal stromal cells, we hypothesized that endothelial lineage cells may also be propagated efficiently following a comparable strategy. Here we demonstrate that human ECFCs can be recovered directly from unmanipulated whole blood. A novel large-scale animal protein-free humanized expansion strategy preserves the progenitor hierarchy with sustained proliferation potential of more than 30 population doublings. By applying large-scale propagated ECFCs in various test systems, we observed vascular networks in vitro and perfused vessels in vivo. After large-scale expansion and cryopreservation phenotype, function, proliferation, and genomic stability were maintained. For the first time, proliferative, functional, and storable ECFCs propagated under humanized conditions can be explored in terms of their therapeutic applicability and risk profile.

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Role of β2-integrins for homing and neovascularization capacity of endothelial progenitor cells

Journal of Experimental Medicine ◽

10.1084/jem.20041402 ◽

2004 ◽

Vol 201 (1) ◽

pp. 63-72 ◽

Cited By ~ 230

Author(s):

Emmanouil Chavakis ◽

Alexandra Aicher ◽

Christopher Heeschen ◽

Ken-ichiro Sasaki ◽

Ralf Kaiser ◽

...

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Ex Vivo ◽

Limb Ischemia ◽

Endothelial Progenitor ◽

Cell Monolayers ◽

Β2 Integrins

The mechanisms of homing of endothelial progenitor cells (EPCs) to sites of ischemia are unclear. Here, we demonstrate that ex vivo–expanded EPCs as well as murine hematopoietic Sca-1+/Lin− progenitor cells express β2-integrins, which mediate the adhesion of EPCs to endothelial cell monolayers and their chemokine-induced transendothelial migration in vitro. In a murine model of hind limb ischemia, Sca-1+/Lin− hematopoietic progenitor cells from β2-integrin–deficient mice are less capable of homing to sites of ischemia and of improving neovascularization. Preactivation of the β2-integrins expressed on EPCs by activating antibodies augments the EPC-induced neovascularization in vivo. These results provide evidence for a novel function of β2-integrins in postnatal vasculogenesis.

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Redefining endothelial progenitor cells via clonal analysis and hematopoietic stem/progenitor cell principals

Blood ◽

10.1182/blood-2006-08-043471 ◽

2006 ◽

Vol 109 (5) ◽

pp. 1801-1809 ◽

Cited By ~ 1044

Author(s):

Mervin C. Yoder ◽

Laura E. Mead ◽

Daniel Prater ◽

Theresa R. Krier ◽

Karim N. Mroueh ◽

...

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Progenitor Cell ◽

Cell Activity ◽

Janus Kinase ◽

Proliferative Potential ◽

Endothelial Progenitor ◽

Hematopoietic Stem ◽

Vascular Regeneration

Abstract The limited vessel-forming capacity of infused endothelial progenitor cells (EPCs) into patients with cardiovascular dysfunction may be related to a misunderstanding of the biologic potential of the cells. EPCs are generally identified by cell surface antigen expression or counting in a commercially available kit that identifies “endothelial cell colony-forming units” (CFU-ECs). However, the origin, proliferative potential, and differentiation capacity of CFU-ECs is controversial. In contrast, other EPCs with blood vessel-forming ability, termed endothelial colony-forming cells (ECFCs), have been isolated from human peripheral blood. We compared the function of CFU-ECs and ECFCs and determined that CFU-ECs are derived from the hematopoietic system using progenitor assays, and analysis of donor cells from polycythemia vera patients harboring a Janus kinase 2 V617F mutation in hematopoietic stem cell clones. Further, CFU-ECs possess myeloid progenitor cell activity, differentiate into phagocytic macrophages, and fail to form perfused vessels in vivo. In contrast, ECFCs are clonally distinct from CFU-ECs, display robust proliferative potential, and form perfused vessels in vivo. Thus, these studies establish that CFU-ECs are not EPCs and the role of these cells in angiogenesis must be re-examined prior to further clinical trials, whereas ECFCs may serve as a potential therapy for vascular regeneration.

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Cilostazol Improves Proangiogenesis Functions in Human Early Endothelial Progenitor Cells through the Stromal Cell-Derived Factor System and Hybrid Therapy Provides a Synergistic EffectIn Vivo

BioMed Research International ◽

10.1155/2016/3639868 ◽

2016 ◽

Vol 2016 ◽

pp. 1-18 ◽

Cited By ~ 2

Author(s):

Shih-Ya Tseng ◽

Ting-Hsing Chao ◽

Yi-Heng Li ◽

Chung-Lung Cho

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Stromal Cell ◽

Ex Vivo ◽

Akt Signaling ◽

Endothelial Progenitor ◽

Hybrid Therapy ◽

Stromal Cell Derived Factor

This study investigated the effect of cilostazol on proangiogenesis functions in human early endothelial progenitor cells (EPCs)in vitroand the therapeutic implication of hybrid therapy with cilostazol and human early EPCsin vivo. Cilostazol significantly increased colony-forming units and enhanced differentiation of EPCs toward endothelial lineage. Treatments resulted in antiapoptotic effects and stimulated proliferation and migration andin vitrovascular tube formation through activation of stromal cell-derived factor-1 (SDF-1)/C-X-C chemokine receptor type 4 (CXCR4)/phosphatidylinositol-3 kinase (PI3K)/Akt signaling pathway. Blood flow recovery and capillary density in murine ischemic hindlimbs were significantly improved in cilostazol-treated, human early EPCs-treated, and cotreatment groups. The effects were attenuated with SDF-1αinhibition. Plasma SDF-1αlevels were significantly higher in 3 active treatment groups after surgery, with greatest effects observed in hybrid therapy. The angiogenic effects of transplanted EPCs pretreated with cilostazolex vivowere superior to untreated EPCs usingin vivoMatrigel assay. Implanted EPCs were incorporated into the capillary, with pretreatment or cotreatment with cilostazol resulting in enhanced effects. Taken together, cilostazol promotes a large number of proangiogenic functions in human early EPCs through activation of SDF-1/CXCR4/PI3K/Akt signaling, and hybrid therapy provides a synergistic effectin vivo. Cotreatment may be beneficial in ischemic disease.

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Endothelial progenitor cell transplantation alleviated ischemic brain injury via inhibiting C3/C3aR pathway in mice

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x19892777 ◽

2019 ◽

Vol 40 (12) ◽

pp. 2374-2386

Author(s):

Yuanyuan Ma ◽

Lu Jiang ◽

Liping Wang ◽

Yongfang Li ◽

Yanqun Liu ◽

...

Keyword(s):

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Cell Transplantation ◽

Endothelial Progenitor Cell ◽

Progenitor Cell ◽

Brain Infarct ◽

Endothelial Progenitor ◽

Ischemic Brain ◽

Neurological Outcomes ◽

The Brain

Endothelial progenitor cell transplantation is a potential therapeutic approach in brain ischemia. However, whether the therapeutic effect of endothelial progenitor cells is via affecting complement activation is unknown. We established a mouse focal ischemia model ( n = 111) and transplanted endothelial progenitor cells into the peri-infarct region immediately after brain ischemia. Neurological outcomes and brain infarct/atrophy volume were examined after ischemia. Expression of C3, C3aR and pro-inflammatory factors were further examined to explore the role of endothelial progenitor cells in ischemic brain. We found that endothelial progenitor cells improved neurological outcomes and reduced brain infarct/atrophy volume after 1 to 14 days of ischemia compared to the control ( p < 0.05). C3 and C3aR expression in the brain was up-regulated at 1 day up to 14 days ( p < 0.05). Endothelial progenitor cells reduced astrocyte-derived C3 ( p < 0.05) and C3aR expression ( p < 0.05) after ischemia. Endothelial progenitor cells also reduced inflammatory response after ischemia ( p < 0.05). Endothelial progenitor cell transplantation reduced astrocyte-derived C3 expression in the brain after ischemic stroke, together with decreased C3aR and inflammatory response contributing to neurological function recovery. Our results indicate that modulating complement C3/C3aR pathway is a novel therapeutic target for the ischemic stroke.

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In vivo vasculogenic potential of human blood-derived endothelial progenitor cells

Blood ◽

10.1182/blood-2006-12-062471 ◽

2007 ◽

Vol 109 (11) ◽

pp. 4761-4768 ◽

Cited By ~ 349

Author(s):

Juan M. Melero-Martin ◽

Zia A. Khan ◽

Arnaud Picard ◽

Xiao Wu ◽

Sailaja Paruchuri ◽

...

Keyword(s):

Endothelial Cells ◽

Progenitor Cells ◽

Endothelial Progenitor Cells ◽

Umbilical Vein ◽

Human Umbilical Cord Blood ◽

Human Umbilical Cord ◽

Vascular Networks ◽

Endothelial Progenitor ◽

Immunodeficient Mice

Abstract Vascularization of tissues is a major challenge of tissue engineering (TE). We hypothesize that blood-derived endothelial progenitor cells (EPCs) have the required proliferative and vasculogenic activity to create vascular networks in vivo. To test this, EPCs isolated from human umbilical cord blood or from adult peripheral blood, and human saphenous vein smooth muscle cells (HSVSMCs) as a source of perivascular cells, were combined in Matrigel and implanted subcutaneously into immunodeficient mice. Evaluation of implants at one week revealed an extensive network of human-specific lumenal structures containing erythrocytes, indicating formation of functional anastomoses with the host vasculature. Quantitative analyses showed the microvessel density was significantly superior to that generated by human dermal microvascular endothelial cells (HDMECs) but similar to that generated by human umbilical vein endothelial cells (HUVECs). We also found that as EPCs were expanded in culture, their morphology, growth kinetics, and proliferative responses toward angiogenic factors progressively resembled those of HDMECs, indicating a process of in vitro maturation. This maturation correlated with a decrease in the degree of vascularization in vivo, which could be compensated for by increasing the number of EPCs seeded into the implants. Our findings strongly support the use of human EPCs to form vascular networks in engineered organs and tissues.

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