Growth Of Endothelial Progenitor Cells Research Articles

Primary aldosteronism (PA) and endothelial progenitor cell (EPC) bioavailability

Patients with primary aldosteronism (PA) experience more cardiovascular events than patients with essential hypertension matched for risk factor profile. Endothelial progenitor cells (EPC) represent a bone marrow-derived cell population implicated in vascular healing whose number correlates to the cardiovascular risk factor profile. Aldosterone has been reported to decrease EPC proliferation in rats. We assessed (i) the growth characteristics of EPC from six PA patients and six matched normotensive controls; (ii) the growth characteristics of EPC treated with increasing doses of aldosterone. Senescence and cell-cycle analysis of EPC from PA patients and normotensive controls and of aldosterone-treated EPC from healthy volunteers. No difference was found in the senescence rate between EPC from PA patients (72.4% senescent cells) and controls (70.7%, P > 0.05). No difference was also found in the cell-cycle distribution determined by FACS (controls: 75.2% cells in G0/G1 phase; PA: 73.5%, P > 0.05). Incubation of EPC with aldosterone did not modify their senescence rate (controls: 72.4% senescent cells; aldosterone 10 nmol/l: 70.9%; aldosterone 100 nmol/l 71.6%, P > 0.05 for all comparisons) and cell-cycle distribution (controls: 73.3% cells in G0/G1 phase; aldosterone 10 nmol/l: 74.9%; aldosterone 100 nmol/l: 75.4%, P > 0.05 for all comparisons). No expression of the mineralocorticoid receptor (MR) transcript was found in EPC by RT-PCR analysis. High aldosterone levels, both in PA patients and in vitro, exert no direct or indirect effect on EPC growth characteristics.

Effects of erythropoietin after an acute myocardial infarction: Rationale and study design of a prospective, randomized, clinical trial (HEBE III)

Preclinical studies have consistently shown that erythropoietin (EPO), administered after an acute myocardial infarction (AMI), reduces infarct size and improves left ventricular function. Furthermore, EPO promotes endothelial progenitor cell growth, which increases angiogenesis. A recent pilot study in patients with AMI suggested that a single bolus of EPO was safe and well tolerated. The HEBE III is a multicenter, prospective, randomized, open-label trial with blinded evaluation of the primary end point. The primary objective is to study the effect on left ventricular ejection fraction (LVEF) of a single bolus of EPO, administered directly after a primary percutaneous coronary intervention (PCI) for a first AMI. A total of 466 patients with thrombolysis in myocardial infarction 0/1 flow before the PCI procedure and 2/3 flow after a successful PCI are randomly assigned to either receive standard medical care or a single bolus with 60,000 IU of EPO on top of standard medical care within 3 hours of the PCI procedure. Primary end point of the study is LVEF after 6 weeks, assessed by planar radionuclide ventriculography. If an improvement of LVEF with a single bolus of EPO is demonstrated, this simple approach might further improve clinical outcome of patients with AMI.

Insulin Stimulates the Clonogenic Potential of Angiogenic Endothelial Progenitor Cells by IGF-1 Receptor-Dependent Signaling

Endothelial progenitor cells (EPCs) have been shown to be involved in vascular regeneration and angiogenesis in experimental diabetes. Because insulin therapy mobilizes circulating progenitor cells, we studied the effects of insulin on outgrowth of EPCs from peripheral blood mononuclear cells of healthy volunteers and patients with type 2 diabetes. Insulin increased the formation of EPC colony-forming units in a dose-dependent manner, half-maximal at 1.5 nM and peaking at 15 nM. Inhibiting the insulin receptor with neutralizing antibodies or antisense oligonucleotides had no effect on EPC outgrowth.(1) In contrast, targeting the human insulin-like growth factor 1 (IGF-1) receptor with neutralizing antibodies significantly suppressed insulin-induced outgrowth of EPCs from both healthy controls and patients with type 2 diabetes. This IGF-1 receptor-mediated insulin effect on EPC growth was at least in part dependent on MAP kinases(2) and was abrogated when extracellular signal-regulated kinase 1/2 (Erk1/2) and protein kinase 38 (p38) activity was inhibited. To study the functional relevance of the observed insulin effects, we studied EPC-induced tube formation of bovine endothelial cells in vitro. Insulin-stimulated EPCs incorporated into the endothelial tubes and markedly enhanced tube formation. In conclusion, this is the first study showing an insulin-mediated activation of the IGF-1 receptor leading to an increased clonogenic and angiogenic potential of EPCs in vitro.

Rafs constitute a nodal point in the regulation of embryonic endothelial progenitor cell growth and differentiation

Mouse embryonic endothelial progenitor cells (eEPCs) acquire a mature phenotype after treatment with cyclic adenosine monophosphate (cAMP), suggesting an involvement of Raf serine/threonine kinases in the differentiation process. To test this idea, we investigated the role of B-Raf and C-Raf in proliferation and differentiation of eEPCs by expressing fusion proteins consisting of the kinase domains from Raf molecules and the hormone binding site of the estrogen receptor (ER), or its variant, the tamoxifen receptor. Our findings show that both B- and C-Raf kinase domains, when lacking adjacent regulatory parts, are equally effective in inducing eEPC differentiation. In contrast, the C-Raf kinase domain is a more potent stimulator of eEPC proliferation than B-Raf. In a complimentary approach, we used siRNA silencing to knockdown endogenously expressed B-Raf and C-Raf in eEPCs. In this experimental setting, we found that eEPCs lacking B-Raf failed to differentiate, whereas loss-of C-Raf function primarily slowed cell growth without impairing cAMP-induced differentiation. These findings were further corroborated in B-Raf null eEPCs, isolated from the corresponding knockout embryos, which failed to differentiate in vitro. Thus, gain- and loss-of-function experiments point to distinct roles of B-Raf and C-Raf in regulating growth and differentiation of endothelial progenitor cells, which may harbour therapeutic implications.

Association of circulating endothelial progenitor cell growth in patients with Type 2 diabetes with type of glucose-lowering treatment

Diabetic MedicineVolume 24, Issue 8 p. 926-927 Association of circulating endothelial progenitor cell growth in patients with Type 2 diabetes with type of glucose-lowering treatment H. Sourij, H. Sourij Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this authorD. Strunk, D. Strunk Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this authorE. Rohde MD, E. Rohde MD Department of Blood Group Serology and Transfusion Medicine, Medical University of Graz, AustriaSearch for more papers by this authorT. C. Wascher, T. C. Wascher Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this author H. Sourij, H. Sourij Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this authorD. Strunk, D. Strunk Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this authorE. Rohde MD, E. Rohde MD Department of Blood Group Serology and Transfusion Medicine, Medical University of Graz, AustriaSearch for more papers by this authorT. C. Wascher, T. C. Wascher Department of Internal Medicine, Metabolism and Vascular Biology Unit, Division of Hematology andSearch for more papers by this author First published: 19 July 2007 https://doi.org/10.1111/j.1464-5491.2007.02214.x DOI: 10.1111/j.1464-5491.2007.02214.x Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat No abstract is available for this article. Volume24, Issue8August 2007Pages 926-927 RelatedInformation

Purification and growth of endothelial progenitor cells from murine bone marrow mononuclear cells

This study reports the culture and purification of murine bone marrow endothelial progenitor cells (EPCs) using endothelial cell-conditioned medium (EC-CM). Endothelial-like cells appeared at day 5 in culture of bone marrow mononuclear cells in the presence of EC-CM in the culture system, and these cells incorporated acetylated low-density lipoproteins (Ac-LDL) and reacted with endothelial-specific Ulex Europaeus Lectin. Continued incubation of these cells at low density with EC-CM for longer than 10 days resulted in the formation of endothelial cell colonies which gave rise to colonies of endothelial progeny and can be passed for many generations in the EC-CM culture system. Cells derived from these colonies expressed endothelial cell markers such as vWF and CD31, incorporated Dil-Ac-LDL, stained positive for Ulex Europaeus Lectin, formed capillary-like structures on Matrigel, and demonstrated a high proliferative capacity in culture. These bone marrow-derived adherent cells were identified as EPCs. The purification and the formation of EPC colonies by using EC-CM were associated with the cytokines secreted in the EC-CM. VEGF, bFGF, and GM-CSF in the EC-CM stimulated the proliferation and growth of EPCs, whereas AcSDKP (tetrapeptide NAc-Ser-Asp-Lys-Pro) in EC-CM suppressed the growth of mesenchymal stem cells (MSC) and fibroblasts. This approach is efficient for isolation/purification and outgrowth of bone marrow EPCs in vitro, a very important cell source in angiogenic therapies and regenerative medicine.

Benfotiamine Counteracts Glucose Toxicity Effects on Endothelial Progenitor Cell Differentiation via Akt/FoxO Signaling

Dysfunction of mature endothelial cells is thought to play a major role in both micro- and macrovascular complications of diabetes. However, recent advances in biology of endothelial progenitor cells (EPCs) have highlighted their involvement in diabetes complications. To determine the effect of glucotoxicity on EPCs, human EPCs have been isolated from peripheral blood mononuclear cells of healthy donors and cultured in the presence or absence of high glucose (33 mmol/l) or high glucose plus benfotiamine to scavenge glucotoxicity. Morphological analysis revealed that high glucose significantly affected the number of endothelial cell colony forming units, uptake and binding of acLDL and Lectin-1, and the ability to differentiate into CD31- and vascular endothelial growth factor receptor 2-positive cells. Functional analysis outlined a reduced EPC involvement in de novo tube formation, when cocultured with mature endothelial cells (human umbilical vein endothelial cells) on matrigel. To explain the observed phenotypes, we have investigated the signal transduction pathways known to be involved in EPC growth and differentiation. Our results indicate that hyperglycemia impairs EPC differentiation and that the process can be restored by benfotiamine administration, via the modulation of Akt/FoxO1 activity.

Modulation of Endothelial Progenitor Cells by Polyunsaturated Omega‐3 Fatty Acids

Introduction Omega-3 polyunsaturated fatty acids (Omega-3 PUFA) are emerging as dietary and pharmaceutical agents with multiple health benefits in both cancer and cardiovascular disease. We hypothesized that the effects of docosahexaenoic acid (DHA), an Omega-3 PUFA, may in part be mediated by its effects on endothelial progenitor cells (EPCs), which are key components of angiogenesis in both cardiovascular disease and cancer. Methods EPCs were isolated as previously published from umbilical cord blood and cultured on collagen in endothelial growth medium. We exposed them for 24 hours to varying doses of DHA. Cell growth was assessed by cell counts and also by flow cytometric analysis of DNA content. Results Cell counts demonstrated that the mean EPC number showed a trend towards an increase in low DHA doses of 1 micromolar, from a baseline mean of 169000 to 201000 cells per well. Interestingly, the cell number decreased at 20 micromolar DHA to lower than baseline levels of 128000 cells per well. DNA staining suggested a similar dose-dependent effect, but also showed inter-individual variation in the DHA effects. Conclusions Our results suggest that DHA may have a biphasic effect on EPC growth. This may explain why omega-3 PUFA can be beneficial in cardiovascular disease, where EPC growth would be required and also have benefits in cancer, where EPC growth and angiogenesis need to be inhibited.

Intrarenal Injection of Bone Marrow-Derived Angiogenic Cells Reduces Endothelial Injury and Mesangial Cell Activation in Experimental Glomerulonephritis

Loss of glomerular endothelial cells has been suggested to contribute to the progression of glomerular injury. Although therapeutic angiogenesis induced by administration of bone marrow-derived endothelial progenitor cells has been observed in disease models of endothelial injury, the effects on renal disease have not been clarified. Whether administration of culture-modified bone marrow mononuclear cells would mitigate the glomerular endothelial injury in anti-Thy1.1 nephritis was investigated. After cultivation under conditions that promote endothelial progenitor cell growth, bone marrow mononuclear cells were labeled with CM-DiI, a fluorescence marker, and injected into the left renal artery of Lewis rats with anti-Thy1.1 glomerulonephritis. The decrease in glomerular endothelial cells was significantly attenuated in the left kidney, as compared with the right, in nephritic rats that received the cell infusion. Glomerular injury score, the area positive for mesangial alpha-smooth muscle actin, and infiltration of macrophages were significantly decreased in the left kidney. CM-DiI-positive cells were distributed in glomeruli of the left kidney but not in those of the right kidney. Among CM-DiI-labeled cells incorporated into glomeruli, 16.5 +/- 1.2% of cells were stained with an endothelial marker, rat endothelial cell antigen-1. Culture-modified mononuclear cells secreted 281.2 +/- 85.0 pg of vascular endothelial growth factor per 10(5) cells per day. In conclusion, intra-arterial administration of culture-modified bone marrow mononuclear cells reduced endothelial injury and mesangial activation in anti-Thy1.1 glomerulonephritis. Incorporation into the glomerular endothelial lining and production of angiogenic factor(s) are likely to contribute to the protective effects of culture-modified mononuclear cells against glomerular injury.

Stem Cells Combined With Gene Transfer for Therapeutic Vasculogenesis

Discoveries in one scientific field sometimes have the unexpected result of igniting a major step forward or new direction in another field. We are in the early phases of one of these sea changes. Advances in stem cell therapies are progressing at a rapid clip and are providing the field of gene therapy with an expanded platform of technologies for the introduction of recombinant genes into human tissues. Likewise, a decade worth of experience in gene therapy provides stem cell researchers with a jump-start. Indeed, it is likely and probable that these 2 fields will eventually merge due to significant scientific and technical overlap. See p 732 The major contribution of the study by Iwaguro et al1 in this issue of Circulation is the demonstration that gene transfer of vectors encoding VEGF into endothelial progenitor cells (EPCs) ex vivo and injection of gene modified EPCs into ischemic muscle in vivo increases neovascularization in an animal model of hindlimb ischemia. Peripheral blood mononuclear cells from human volunteers were cultured in a special cocktail that facilitates growth of EPCs—plating on fibronectin and grown in the presence of multiple growth factors, such as endothelial cell basal medium, fetal bovine serum, human VEGF-A, human FGF-2, human EGF, IGF-1, and ascorbic acid—as previously characterized by this group.2 After 7 days in culture, EPCs were transduced with adenoviral vectors encoding a murine VEGF164 gene or a control reporter gene. Proliferative indices and adhesion properties were measured ex vivo in order to characterize the phenotype of the cells. The authors then conducted pilot studies in vivo to determine the dose of gene-modified cells required to produce a similar magnitude of neovascularization achieved with nontransduced EPCs. Interestingly, they found that 30 times fewer VEGF-transduced EPCs (1.5×104 cells) produced equivalent neovascularization compared with 4.5×105 …