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Concept: Endothelial progenitor cell

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Type 1 diabetes is associated with increased cardiovascular disease (CVD). Decreased endothelial progenitor cells (EPCs) number plays a pivotal role in reduced endothelial repair and development of CVD. We aimed to determine if cardioprotective effect of metformin is mediated by increasing circulating endothelial progenitor cells (cEPCs), pro-angiogenic cells (PACs) and decreasing circulating endothelial cells (cECs) count whilst maintaining unchanged glycemic control.

Concepts: Atherosclerosis, Diabetes mellitus type 2, Diabetes mellitus, Diabetes mellitus type 1, Blood vessel, Diabetes, Endothelial progenitor cell, Insulin resistance

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BACKGROUND: Far infra-red (IFR) therapy was shown to exert beneficial effects in cardiovascular system, but effects of IFR on endothelial progenitor cell (EPC) and EPC-related vasculogenesis remain unclear. We hypothesized that IFR radiation can restore blood flow recovery in ischemic hindlimb in diabetic mice by enhancement of EPCs functions and homing process.Materials and methodsStarting at 4 weeks after the onset of diabetes, unilateral hindlimb ischemia was induced in streptozotocine (STZ)-induced diabetic mice, which were divided into control and IFR therapy groups (n = 6 per group). The latter mice were placed in an IFR dry sauna at 34[DEGREE SIGN]C for 30 min once per day for 5 weeks. RESULTS: Doppler perfusion imaging demonstrated that the ischemic limb/normal side blood perfusion ratio in the thermal therapy group was significantly increased beyond that in controls, and significantly greater capillary density was seen in the IFR therapy group. Flow cytometry analysis showed impaired EPCs (Sca-1+/Flk-1+) mobilization after ischemia surgery in diabetic mice with or without IFR therapy (n = 6 per group). However, as compared to those in the control group, bone marrow-derived EPCs differentiated into endothelial cells defined as GFP+/CD31+ double-positive cells were significantly increased in ischemic tissue around the vessels in diabetic mice that received IFR radiation. In in-vitro studies, cultured EPCs treated with IFR radiation markedly augmented high glucose-impaired EPC functions, inhibited high glucose-induced EPC senescence and reduced H2O2 production. Nude mice received human EPCs treated with IFR in high glucose medium showed a significant improvement in blood flow recovery in ischemic limb compared to those without IFR therapy. IFR therapy promoted blood flow recovery and new vessel formation in STZ-induced diabetic mice. CONCLUSIONS: Administration of IFR therapy promoted collateral flow recovery and new vessel formation in STZ-induced diabetic mice, and these beneficial effects may derive from enhancement of EPC functions and homing process.

Concepts: Blood, Angiogenesis, Heart, Blood vessel, Capillary, Endothelial progenitor cell, Flow cytometry, Cardiovascular system

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Smoking is a major risk factor for atherosclerosis. In this study, we evaluated the effects of benzo[a]pyrene (BaP, a prominent component of tobacco smoke) on the function and pro-inflammatory response of human endothelial progenitor cells (EPCs). EPCs were isolated from umbilical cord blood and treated with different concentrations (10, 20 and 50 µmol/l) of BaP. The proliferation, migration, adhesion and angiogenesis of BaP-treated EPCs were evaluated using the cell counting kit-8 (CCK-8), Transwell assay, adhesion assay and in vitro tube formation assay, respectively. The activation of nuclear factor-κB (NF-κB) was evaluated by measuring the mRNA expression of NF-κB p65 and p50 by real-time RT-PCR and NF-κB translocation assay. Reactive oxygen species (ROS) production was determined by the reduction of fluorescent 2',7'-dichlorofluorescein diacetate (DCFH-DA). The results demonstrated that BaP treatment significantly inhibited the proliferation, migration, adhesion and angiogenesis of EPCs in vitro. In addition, BaP induced the release of interleukin (IL)-1β and tumor necrosis factor-α from these cells. Moreover, the exposure of EPCs to BaP induced ROS generation and the activation of NF-κB. Experiments with EPCs pre-treated with pyrrolidine dithiocarbamate, an inhibitor of NF-κB, revealed that the BaP-mediated inhibition of proliferation, migration, adhesion and angiogenesis of EPCs is mainly regulated by NF-κB. Thus, tobacco smoke may induce oxidant-mediated stress responses in EPCs and impair their function via the activation of the NF-κB pathway.

Concepts: Cancer, Oxidative stress, Oxidative phosphorylation, Reactive oxygen species, Endothelial progenitor cell, Tobacco smoking, Umbilical cord, Cord blood

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Endothelial progenitor cells (EPCs ) have reduced expression of eNOS, this may decrease their antithrombogenic property when used as seeding cells for small caliber vascular graft. The aim of this study is to investigate whether overexpression of eNOS in EPCs can increase its antithrombogenic property and regulate tissue factor (TF) level.

Concepts: Gene expression, Endothelial progenitor cell, Progenitor cell, Knitting

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Endothelial progenitor cells (EPCs) can be isolated from human bone marrow or peripheral blood and reportedly contribute to neovascularization. Aptamers are 40-120-mer nucleotides that bind to a specific target molecule, as antibodies do. To utilize apatmers for isolation of EPCs, in the present study, we successfully generated aptamers that recognize human CD31, an endothelial cell marker. CD31 aptamers bound to human umbilical cord blood-derived EPCs and showed specific interaction with human CD31, but not with mouse CD31. However, CD31 aptamers showed non-specific interaction with CD31-negative 293FT cells and addition of polyanionic competitor dextran sulfate eliminated non-specific interaction without affecting cell viability. From the mixture of EPCs and 293FT cells, CD31 aptamers successfully isolated EPCs with 97.6% purity and 94.2% yield, comparable to those from antibody isolation. In addition, isolated EPCs were decoupled from CD31 aptamers with a brief treatment of high concentration dextran sulfate. EPCs isolated with CD31 aptamers and subsequently decoupled from CD31 aptamers were functional and enhanced the restoration of blood flow when transplanted into a murine hindlimb ischemia model. In this study, we demonstrated isolation of foreign material-free EPCs, which can be utilized as a universal protocol in preparation of cells for therapeutic transplantation.

Concepts: DNA, Blood, Bone marrow, Blood vessel, Endothelial progenitor cell, Ischemia, Umbilical cord, Aptamer

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We have previously shown that estrogen (estradiol; E2) supplementation enhances voluntary alcohol consumption in ovariectomized (OVX) female rodents and that increased alcohol consumption impairs ischemic hind limb vascular repair. However, the effect of E2-induced alcohol consumption on post-infarct myocardial repair and on the phenotypic/functional properties of endothelial progenitor cells (EPCs) is not known. Additionally, the molecular signaling of alcohol-estrogen interactions remains to be elucidated. This study examined the effect of E2-induced increases in ethanol consumption on post-infarct myocardial function/repair. OVX female mice, implanted with 17β-E2 or placebo pellets were given access to alcohol for 6 weeks and subjected to acute myocardial infarction (AMI). Left ventricular functions were consistently depressed in mice consuming ethanol compared to those receiving only E2. Alcohol consuming mice also displayed significantly increased infarct size and reduced capillary density. Ethanol consumption also reduced E2-induced mobilization and homing of EPCs to injured myocardium compared to the E2 alone group. In vitro, exposure of EPCs to ethanol suppressed E2-induced proliferation, survival, migration and markedly altered E2-induced estrogen receptor-dependent cell survival signaling and gene expression. Furthermore, ethanol-mediated suppression of EPC biology was eNOS-dependent as eNOS-null mice displayed an exaggerated response to post-AMI LV functions. These data suggest that E2-modulation of alcohol consumption, and the ensuing EPC dysfunction, may negatively compete with the beneficial effects of estrogen on post-infarct myocardial repair.

Concepts: Gene, Alcohol, Myocardial infarction, Atherosclerosis, Endothelial progenitor cell, Progenitor cell, Infarction, Necrosis

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In the microcirculation, pericytes are believed to function as mesenchymal stromal cells (MSCs). We hypothesized that the vasa vasorum harbor progenitor cells within the adventitia of human aorta. Pericytes, endothelial progenitor cells, and other cell subpopulations were detected among freshly isolated adventitial cells using flow cytometry. Purified cultured pericytes were enriched for the MSC markers CD105 and CD73 and depleted of the endothelial markers von Willebrand factor and CD31. Cultured pericytes were capable of smooth muscle lineage progression including inducible expression of smooth muscle myosin heavy chain, calponin, and α-smooth muscle actin, and adopted a spindle shape. Pericytes formed spheroids when cultured on Matrigel substrates and peripherally localized with branching endothelial cells in vitro. Our results indicate that the vasa vasorum form a progenitor cell niche distinct from other previously described progenitor populations in human adventitia. These findings could have important implications for understanding the complex pathophysiology of human aortic disease.

Concepts: Protein, Mesenchymal stem cell, Cell biology, Endothelial progenitor cell, Actin, Myosin, Muscle contraction, Von Willebrand factor

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Although nitric oxide (NO) signaling promotes differentiation and maturation of endothelial progenitor cells, its role in the differentiation of mesenchymal stem cells (MSCs) into endothelial cells remains controversial. We tested the role of NO signaling in MSCs derived from WT mice and mice homozygous for a deletion of S-nitrosoglutathione reductase (GSNOR(-/-)), a denitrosylase that regulates S-nitrosylation. GSNOR(-/-) MSCs exhibited markedly diminished capacity for vasculogenesis in an in vitro Matrigel tube-forming assay and in vivo relative to WT MSCs. This decrease was associated with down-regulation of the PDGF receptorα (PDGFRα) in GSNOR(-/-) MSCs, a receptor essential for VEGF-A action in MSCs. Pharmacologic inhibition of NO synthase with L-N(G)-nitroarginine methyl ester (L-NAME) and stimulation of growth hormone-releasing hormone receptor (GHRHR) with GHRH agonists augmented VEGF-A production and normalized tube formation in GSNOR(-/-) MSCs, whereas NO donors or PDGFR antagonist reduced tube formation ∼50% by murine and human MSCs. The antagonist also blocked the rescue of tube formation in GSNOR(-/-) MSCs by L-NAME or the GHRH agonists JI-38, MR-409, and MR-356. Therefore, GSNOR(-/-) MSCs have a deficient capacity for endothelial differentiation due to downregulation of PDGFRα related to NO/GSNOR imbalance. These findings unravel important aspects of modulation of MSCs by VEGF-A activation of the PDGFR and illustrate a paradoxical inhibitory role of S-nitrosylation signaling in MSC vasculogenesis. Accordingly, disease states characterized by NO deficiency may trigger MSC-mediated vasculogenesis. These findings have important implications for therapeutic application of GHRH agonists to ischemic disorders.

Concepts: DNA, Stem cell, Mesenchymal stem cell, Bone marrow, Endothelial progenitor cell, Endothelium, Nitric oxide, Growth hormone releasing hormone

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Diabetes mellitus (DM) causes dysfunction of endothelial progenitor cells (EPCs), resulting in impaired wound healing. EPC therapy is a potential substitute to the current treatments of chronic wounds. Since EPCs isolated from diabetic patients are dysfunctional, and therefore pose an obstacle in their efficacious employment in autologous cell therapy, a strategy to rescue them prior to transplantation would be expected to improve the efficacy of autologous cell therapy multi-fold. With compromised ROS scavenging ability being the main cause of EPC Dysfunction (EPCD), ROS scavengers are likely to reverse or rescue EPC dysfunction. Therefore, in the present study, we evaluated the potential of curcumin in reversing DM-induced EPCD. We found that in vitro treatment of bone marrow EPCs from diabetic mice (D-EPC) with curcumin restored their functionality, as judged by colony formation, tubule formation and migration assays. Most importantly, autologous transplantation of curcumin-treated D-EPCs onto diabetic wounds also resulted in accelerated wound healing. Furthermore, curcumin-treated diabetic mice exhibited improved wound healing, as compared to their vehicle-treated diabetic counterparts, underscoring the efficacy of curcumin in vivo as well. The levels and activity of MnSOD in D-EPCs treated in vitro with curcumin or those isolated from curcumin-treated diabetic mice were comparable to those in non-diabetic EPCs. Addition of methyl mercury chloride to inhibit MnSOD activity during curcumin treatment abolished the salutary effects of curcumin. Our data demonstrate that curcumin reverses DM-induced EPC dysfunction by boosting MnSOD expression and activity, and emphasises its potential for use in autologous cell therapy for diabetic wound management.

Concepts: Diabetes mellitus, Wound healing, Endothelial progenitor cell, Progenitor cell, In vivo, In vitro, Wound, Chronic wound

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Two decades after the first report on endothelial progenitor cells (EPC), their key role in postnatal vasculogenesis and vascular repair is well established. The therapeutic potential of EPC and their growing use in clinical trials calls for the development of more robust, reproducible and safer methods for the in vitro expansion and maintenance of these cells. Despite many limitations associated with its usage, fetal bovine serum (FBS) is still widely applied as a cell culture supplement. While different approaches aiming at establishing FBS-free culture have been developed for many cell types, adequate solutions for endothelial cells, and for EPC in particular, are still scarce, possibly due to the multiple challenges that have to be faced when culturing these cells. In this review, we provide a brief overview on the therapeutic relevance of EPC, and critically analyze the available literature on FBS-free endothelial cell culture methods, including xeno-free, serum-free and chemically defined systems.

Concepts: Angiogenesis, Developmental biology, Cell biology, Endothelial progenitor cell, Cell culture, Endothelium