DJ‐1 Alleviates Angiotensin II‐Induced Endothelial Progenitor Cell Damage by Activating the PPARγ/HO‐1 Pathway

There is evidence that angiotensin II (Ang II) may impair the functions of endothelial progenitor cells (EPCs). It was revealed that DJ‐1 could resist oxidative stress. In this study, we investigated whether DJ‐1 could protect EPCs against Ang II‐induced cell damage. The proliferation and migration of EPCs were strongly reduced in the Ang II group and were increased by overexpression of DJ‐1. Western blotting indicated that the increased expression of the senescence marker β‐galactosidase and decreased expression of adhesion molecules (ICAM‐1, VCAM‐1) induced by Ang II were reversed after Ad‐DJ‐1 transfection. The reduced angiogenic capacity of EPCs caused by Ang II was also improved after Ad‐DJ‐1 transfection. Moreover, Ang II significantly increased the levels of reactive oxygen species (ROS), malondialdehyde (MDA), and inflammatory cytokines (TNF‐α and IL‐1β), reduced the levels of superoxide dismutase (SOD), glutathione (GSH), and these were reversed by Ad‐DJ‐1 transfection. Expression of peroxisome proliferator‐activated receptor‐γ (PPARγ) and heme oxygenase (HO‐1) was increased by DJ‐1. Therefore, HO‐1 siRNA were constructed and transfected into EPCs, and the results showed that HO‐1 siRNA transfection inhibited the effects of DJ‐1 on EPC function. Thus, our study implies that DJ‐1 may protect EPCs against Ang II‐induced dysfunction by activating the PPARγ/HO‐1. J. Cell. Biochem. 119: 392–400, 2018. © 2017 Wiley Periodicals, Inc.     

ClC-3 deficiency prevents apoptosis induced by angiotensin II in endothelial progenitor cells via inhibition of NADPH oxidase

Endothelial progenitor cells (EPCs) play an important role in postnatal neovascularization and re-endothelialization in response to tissue ischemia and endothelial injury. It is reported that the circulating EPCs number is decreased during hypertension. However, the detailed mechanism is still unclear. Our previous studies have shown that ClC-3 chloride channel is up-regulated with the development of hypertension. This study aims to test whether ClC-3 participates in EPC apoptosis under the condition of increased oxidative stress in angiotensin II (Ang II)-induced hypertension. The results showed that stimulation with 10^?6mol/L Ang II significantly up-regulated the endogenous ClC-3 expression and increased intracellular reactive oxygen species (ROS) generation in EPCs of wild type mice, accompanied by an enhanced NADPH oxidase activity and the expression of gp91^phox (NOX-2), a key catalytic subunit of NADPH oxidase. However, these effects of Ang II were significantly reduced in EPCs of ClC-3^?/? mice. Compared with control, treatment with Ang II induced EPCs apoptosis in wild type mice, concomitantly with declined Bcl-2/Bax ratio, depressed mitochondrial membrane potential and activation of poly(ADP-ribose) polymerase, which was remarkably prevented by both ClC-3 knockout and NADPH oxidase inhibitor apocynin. In addition, the role of ClC-3 deficiency in protecting EPCs against Ang II-induced oxidative stress and apoptosis was further confirmed in Ang II-infused hypertensive mice in vivo. In conclusion, ClC-3 deficiency inhibited Ang II-induced EPC apoptosis via suppressing ROS generation derived from NADPH oxidase.     

Cytoprotective effect of dieckol on human endothelial progenitor cells (hEPCs) from oxidative stress-induced apoptosis

Abstract

Although endothelial progenitor cells (EPCs) have been used to promote revascularization after peripheral or myocardial ischemia, excess amounts of reactive oxygen species (ROS) are often involved in senescence and apoptosis of EPCs, thereby causing defective neovascularization and reduced or failed recovery. Here, we examined the cytoprotective effect of Ecklonia cava-derived antioxidant dieckol (DK) on oxidative stress-induced apoptosis in EPCs to improve EPC bioactivity for vessel repair. Although H₂O₂ (10 ^{? 3} M) increased the intracellular ROS level in EPCs, DK (10ug/ml) pretreatment suppressed the H₂O₂-induced ROS increase and drastically reduced the ratios of apoptotic cells. H₂O₂-induced ROS increased the phosphorylation of p38 MAPK and JNK; this was inhibited by DK pretreatment. H₂O₂ treatment increased the phosphorylation of NF-κB, which was blocked by pretreatment with SB 203580, a p38 MAPK inhibitor, or SP 600125, a JNK inhibitor. H₂O₂ decreased the cellular levels of Bcl-2 and c-IAPs, cellular inhibitors of apoptosis proteins, but increased caspase-3 activation. However, all these effects were inhibited by pretreatment with DK. Injection of DK-mixed EPCs (DK + EPCs) into myocardial ischemic sites in vivo induced cellular proliferation and survival of cells at the ischemic sites and, thereby, enhanced the secretion of angiogenic cytokines at the ischemic sites. These results show that DK + EPC exhibit markedly enhanced anti-apoptotic and antioxidative capabilities, unlike that shown by EPCs alone; thus, they contribute to improved repair of ischemic myocardial injury through cell survival and angiogenic cytokine production.     

Regulation of plasticity and biological features of endothelial progenitor cells by MSC-derived SDF-1

Bone marrow (BM) is a source of mesenchymal stromal cells (MSCs) and endothelial progenitor cells (EPCs). MSCs provide a specific niche in the BM and biological features of EPCs may be changed with this niche. Stromal cell-derived factor 1 (SDF-1) secreted from primary BM-MSCs and biological features of this niche on EPC development are still yet to be understood. The aim of this study was to evaluate the role of SDF-1 produced by MSCs on EPC development. We applied the CRISPR/Cas9 system for the knock-out of the SDF-1 gene in BM-derived MSCs. BM-derived EPCs were then cocultured with MSCs^SDF-1−/− or MSCs^SDF-1+/+ to identify the role of MSC-derived SDF-1α on proliferation, migration and angiogenic activity of EPCs. Next, pre-expanded EPCs were harvested and co-transplanted with MSCs^SDF-1−/− or MSCs^SDF-1+/+ into sublethally irradiated mice to analyze the potency of these cells for marrow reconstitution. Our results revealed that proliferation, colony formation, migration and angiogenic activity of EPCs was significantly increased after coculture with MSCs^SDF-1+/+. We also found that co-transplantation of EPCs with MSCs^SDF-1+/+, in contrast to MSCs^SDF-1−/−, into irradiated mice resulted in marrow repopulation and hematologic recovery, leading to improved survival of transplanted mice. In conclusions, MSC-derived SDF-1 niche plays an important role in the development of EPCs and this niche is essential for bone marrow repopulation by these cells and can enhance the efficiency of EPC therapy for ischemic diseases.     

Sitagliptin‐mediated preservation of endothelial progenitor cell function via augmenting autophagy enhances ischaemic angiogenesis in diabetes

Recently, the dipeptidyl peptidase‐4 (DPP‐4) inhibitor sitagliptin, a major anti‐hyperglycaemic agent, has received substantial attention as a therapeutic target for cardiovascular diseases via enhancing the number of circulating endothelial progenitor cells (EPCs). However, the direct effects of sitagliptin on EPC function remain elusive. In this study, we evaluated the proangiogenic effects of sitagliptin on a diabetic hind limb ischaemia (HLI) model in vivo and on EPC culture in vitro. Treatment of db/db mice with sitagliptin (Januvia) after HLI surgery efficiently enhanced ischaemic angiogenesis and blood perfusion, which was accompanied by significant increases in circulating EPC numbers. EPCs derived from the bone marrow of normal mice were treated with high glucose to mimic diabetic hyperglycaemia. We found that high glucose treatment induced EPC apoptosis and tube formation impairment, which were significantly prevented by sitagliptin pretreatment. A mechanistic study found that high glucose treatment of EPCs induced dramatic increases in oxidative stress and apoptosis; pretreatment of EPCs with sitagliptin significantly attenuated high glucose‐induced apoptosis, tube formation impairment and oxidative stress. Furthermore, we found that sitagliptin restored the basal autophagy of EPCs that was impaired by high glucose via activating the AMP‐activated protein kinase/unc‐51‐like autophagy activating kinase 1 signalling pathway, although an autophagy inhibitor abolished the protective effects of sitagliptin on EPCs. Altogether, the results indicate that sitagliptin‐induced preservation of EPC angiogenic function results in an improvement of diabetic ischaemia angiogenesis and blood perfusion, which are most likely mediated by sitagliptin‐induced prevention of EPC apoptosis via augmenting autophagy.     

Zoledronate Attenuates Angiogenic Effects of Angiotensin II-Stimulated Endothelial Progenitor Cells via RhoA and MAPK Signaling

Background

New vessel formation plays a pivotal role in the pathogenesis of neovascular-related diseases. Endothelial progenitor cells (EPCs) were found to contribute to neovascular-related diseases and interference with EPC neovascularization may be a novel target for these diseases. Zoledronate (Zol) was reported to exhibit anti-angiogenic effect. Basing on these evidences, we proposed that Zol may affect EPC function to exert novel anti-angiogenic effect. In this study, we therefore investigated the effects of Zol on multiple aspects of EPC function and explored the underlying mechanisms involved.

Methodology/Principal Findings

EPCs were cultured from bone marrow derived mononuclear cells. The potential effects of Zol on Angiotensin II (Ang II)-stimulated EPC proliferation, migration, adhesion, in vitro tube formation were investigated. The results showed that Ang II (1 µM) enhanced EPC migration, adhesion, in vitro tube formation but had no effect on cell proliferation. Zol (75 and 100 µM) inhibited proliferation of EPCs and 50 µM geranylgeranyol (GGOH) could reverse the decrease of EPC proliferation. We found for the first time that Zol (50–100 µM) dose dependently attenuated migration, adhesion, and in vitro tube formation of EPCs stimulated by Ang II. GGOH could reverse the attenuation of EPC function induced by Zol. However, Zol did not induce EPC apoptosis. In addition, the underlying mechanisms were determined. The results revealed that Zol markedly down-regulated active RhoA stimulated by Ang II and inhibited the phosphorylation of Erk1/2 and JNK. Moreover, RhoA silencing resulted in a notable inhibition of EPC in vitro tube formation, suggesting that RhoA suppression played a pivotal role in Zol antiangiogenic effect.

Conclusions/Significance

These findings suggested that Zol attenuated the promotion of EPC function stimulated by Ang II and exhibited novel antiangiogenic effect via RhoA and MAPK signaling. Thus, Zol may be served as a novel therapeutic agent for neovascular-related diseases treatment.     

The effect of autologous endothelial progenitor cell transplantation combined with extracorporeal shock-wave therapy on ischemic skin flaps in rats

BackgroundEndothelial progenitor cells (EPCs) have been used to revascularize ischemic tissues, but only limited effect can be achieved. Extracorporeal shock-wave therapy (ESWT) is a promising angiogenic strategy. We hypothesized that EPC transplantation combined with ESWT would greatly benefit the survival of ischemic skin flaps.MethodsSixty-four male Sprague-Dawley rats were divided into 4 groups (n = 16 in each group): group 1 (serving as sham control), group 2 (treated with subcutaneous EPC implantation, 1.0 × 10⁶ cells), group 3 (treated with ESWT, 300 impulses at 0.10 mJ/mm²) and group 4 (treated with EPCs implantation combined with ESWT). Ischemic skin flaps were made on the backs of rats and treated accordingly. Blood flow of skin flaps was measured periodically after operation, and flap survival rates were compared. Tissue samples were harvested at 2 weeks postoperatively from each group.ResultsThe survival rate of skin flaps in group 4 was 87.5 ± 10.23%, which was statistically significantly higher than other groups. Histologic examination showed that the capillary density was higher in the dual-treatment group than in the two single-treatment groups. Compared with groups 2 and 3, blood perfusion increased significantly in group 4. A drastic increase of vWF+ cells was observed in the ischemic skin flaps on immunofluorescence staining in group 4. The expressions of chemotactic factors and angiogenic factors were higher in group 4.ConclusionsCombined treatment with EPCs and ESWT is superior to either EPCs or ESWT alone in improving the survival of ischemic skin flaps in rats.     

Kaposi’s sarcoma-associated herpesvirus infection of endothelial progenitor cells impairs angiogenic activity <Emphasis Type="Italic">in vitro</Emphasis>

A recent study reported that endothelial progenitor cells (EPCs0 are one of the reservoirs of Kaposi’s sarcoma associated herpesvirus (KSHV). Although EPCs are closely linked to angiogenesis and vasculogenesis, little is known about the angiogenic potential of KSHV in EPCs. In this study, we used EPCs isolated from human umbilical cord blood to show that early infection by KSHV in vitro impaired the neovascularization of EPCs in matrigel. Our results suggest that KSHV may disrupt the angiogenic potential of EPCs and that the disseminated infection of KSHV could be associated with EPC dysfunction.     

Concomitant overexpression of triple antioxidant enzymes selectively increases circulating endothelial progenitor cells in mice with limb ischaemia

Endothelial progenitor cells (EPCs) are a group of heterogeneous cells in bone marrow (BM) and blood. Ischaemia increases reactive oxygen species (ROS) production that regulates EPC number and function. The present study was conducted to determine if ischaemia‐induced ROS differentially regulated individual EPC subpopulations using a mouse model concomitantly overexpressing superoxide dismutase (SOD)1, SOD3 and glutathione peroxidase. Limb ischaemia was induced by femoral artery ligation in male transgenic mice with their wild‐type littermate as control. BM and blood cells were collected for EPCs analysis and mononuclear cell intracellular ROS production, apoptosis and proliferation at baseline, day 3 and day 21 after ischaemia. Cells positive for c‐Kit⁺/CD31⁺ or Sca‐1⁺/Flk‐1⁺ or CD34⁺/CD133⁺ or CD34⁺/Flk‐1⁺ were identified as EPCs. ischaemia significantly increased ROS production and cell apoptosis and decreased proliferation of circulating and BM mononuclear cells and increased BM and circulating EPCs levels. Overexpression of triple antioxidant enzymes effectively prevented ischaemia‐induced ROS production with significantly decreased cell apoptosis and preserved proliferation and significantly increased circulating EPCs level without significant changes in BM EPC populations, associated with enhanced recovery of blood flow and function of the ischemic limb. These data suggested that ischaemia‐induced ROS was differentially involved in the regulation of circulating EPC population.     

Angiogenic activity of endothelial progenitor cells through angiopoietin-1 and angiopoietin-2

Angiogenesis is a regulated process involving the proliferation, migration, and remodeling of different cell types particularly mature endothelial cells and recently discovered progenitor cells, named as endothelial progenitor cells (EPCs). Up to now, many attempts have been made to understand the dynamic balance of pro- and anti-angiogenic factors on EPCs on different milieu. It has been accepted that Ang-1, -2 and Tie-1, -2 signaling play a key role on angiogenesis pathways in endothelial lineage cells. In the current experiment, the angiogenic/angio-modulatory potency of Ang-1 and -2 was investigated on isolated EPCs. Freshly isolated EPCs were exposed to different concentrations of Ang-1 and -2 (25 and 50?ng/ml) over a course of 7 and 14 days. Corroborating to our results, a superior effect of Ang-1 on angiogenic properties, including an increased concentration of vascular endothelial growth factor, in vitro tubulogenesis, EPC migratory, Tie-2 expression and clonogenicity, was determined. A large amount of positive mature endothelium markers was achieved in EPCs being-exposed to Ang-1 peptide. Nonetheless, the number of CD133 positive cells increased in the presence of Ang-2. Collectively, we conclude that Ang-1 potentially induces functional and mature vascular-like behavior in EPCs more than Ang-2.     

FGF21 promotes ischaemic angiogenesis and endothelial progenitor cells function under diabetic conditions in an AMPK/NAD+-dependent manner

Diabetic vascular complications are closely associated with long-term vascular dysfunction and poor neovascularization. Endothelial progenitor cells (EPCs) play pivotal roles in maintaining vascular homeostasis and triggering angiogenesis, and EPC dysfunction contributes to defective angiogenesis and resultant diabetic vascular complications. Fibroblast growth factor 21 (FGF21) has received substantial attention as a potential therapeutic agent for diabetes via regulating glucose and lipid metabolism. However, the effects of FGF21 on diabetic vascular complications remain unclear. In the present study, the in vivo results showed that FGF21 efficiently improved blood perfusion and ischaemic angiogenesis in both type 1 and type 2 diabetic mice, and these effects were accompanied by enhanced EPC mobilization and infiltration into ischaemic muscle tissues and increases in plasma stromal cell–derived factor-1 concentration. The in vitro results revealed that FGF21 directly prevented EPC damage induced by high glucose, and the mechanistic studies demonstrated that nicotinamide adenine dinucleotide (NAD⁺) was dramatically decreased in EPCs challenged with high glucose, whereas FGF21 treatment significantly increased NAD⁺ content in an AMPK-dependent manner, resulting in improved angiogenic capability of EPCs. These results indicate that FGF21 promotes ischaemic angiogenesis and the angiogenic ability of EPCs under diabetic conditions by activating the AMPK/NAD⁺ pathway.     

miRNome traits analysis on endothelial lineage cells discloses biomarker potential circulating microRNAs which affect progenitor activities

Background

Endothelial progenitor cells (EPCs) play a fundamental role in not only blood vessel development but also post-natal vascular repair. Currently EPCs are defined as early and late EPCs based on their biological properties and their time of appearance during in vitro culture. Both EPC types assist angiogenesis and have been linked to ischemia-related disorders, including coronary artery disease (CAD).

Results

We found late EPCs are more mobile than early EPCs and matured endothelial cells (ECs). To pinpoint the mechanism, microRNA profiles of early EPCs late EPCs, and ECs were deciphered by small RNA sequencing. Obtained signatures made up of both novel and known microRNAs, in which anti-angiogenic microRNAs such as miR-221 and miR-222 are more abundant in matured ECs than in late EPCs. Overexpression of miR-221 and miR-222 resulted in the reduction of genes involved in hypoxia response, metabolism, TGF-beta signalling, and cell motion. Not only hamper late EPC activities in vitro, both microRNAs (especially miR-222) also hindered in vivo vasculogenesis in a zebrafish model. Reporter assays showed that miR-222, but not miR-221, targets the angiogenic factor ETS1. In contrast, PIK3R1 is the target of miR-221, but not miR-222 in late EPCs. Clinically, both miR-221-PIK3R1 and miR-222-ETS1 pairs are deregulated in late EPCs of CAD patients.

Conclusions

Our results illustrate EPCs and ECs exploit unique miRNA modalities to regulate angiogenic features, and explain why late EPC levels and activities are reduced in CAD patients. These data will further help to develop new plasma biomarkers and therapeutic approaches for ischemia-related diseases or tumor angiogenesis.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-802) contains supplementary material, which is available to authorized users.     

Procyanidin B2 improves endothelial progenitor cell function and promotes wound healing in diabetic mice via activating Nrf2

One of the major reasons for the delayed wound healing in diabetes is the dysfunction of endothelial progenitor cells (EPCs) induced by hyperglycaemia. Improvement of EPC function may be a potential strategy for accelerating wound healing in diabetes. Procyanidin B2 (PCB2) is one of the major components of procyanidins, which exhibits a variety of potent pharmacological activities. However, the effects of PCB2 on EPC function and diabetic wound repair remain elusive. We evaluated the protective effects of PCB2 in EPCs with high glucose (HG) treatment and in a diabetic wound healing model. EPCs derived from human umbilical cord blood were treated with HG. The results showed that PCB2 significantly preserved the angiogenic function, survival and migration abilities of EPCs with HG treatment, and attenuated HG-induced oxidative stress of EPCs by scavenging excessive reactive oxygen species (ROS). A mechanistic study found the protective role of PCB2 is dependent on activating nuclear factor erythroid 2-related factor 2 (Nrf2). PCB2 increased the expression of Nrf2 and its downstream antioxidant genes to attenuate the oxidative stress induced by HG in EPCs, which were abolished by knockdown of Nrf2 expression. An in vivo study showed that intraperitoneal administration of PCB2 promoted wound healing and angiogenesis in diabetic mice, which was accompanied by a significant reduction in ROS level and an increase in circulating EPC number. Taken together, our results indicate that PCB2 treatment accelerates wound healing and increases angiogenesis in diabetic mice, which may be mediated by improving the mobilization and function of EPCs.     

Chronic administration of NMU into the paraventricular nucleus stimulates the HPA axis but does not influence food intake or body weight

C-reactive protein (CRP), a predictor of future cardiovascular diseases, has been reported to damage the vascular wall by inducing endothelial dysfunction and inflammation. This proatherogenic CRP was speculated to have a role in attenuating angiogenic functions of human endothelial progenitor cells (EPCs), possibly impairing vascular regeneration and increasing cardiovascular vulnerability to ischemic injury. Herein, we investigated the direct effect of CRP on angiogenic activity and gene expression in human EPCs. Incubation of EPCs with human recombinant CRP significantly inhibited EPC migration in response to vascular endothelial growth factor, possibly by decreasing the expression of endothelial nitric oxide synthase and subsequent nitric oxide production. In addition, CRP-treated EPCs showed the reduced adhesiveness onto an endothelial cell monolayer. When assayed for the gene expression of arteriogenic chemo-cytokines, CRP substantially decreased their expression levels in EPC, in part due to the upregulation of suppressors of cytokine signaling proteins. These results suggest that CRP directly attenuates the angiogenic and possibly arteriogenic functions of EPCs. This CRP-induced EPC dysfunction may impair the vascular regenerative capacity of EPCs, thereby leading to increased risk of cardiovascular diseases.     

Actin Stabilization by Jasplakinolide Affects the Function of Bone Marrow-Derived Late Endothelial Progenitor Cells

Background

Bone marrow-derived endothelial progenitor cells (EPCs), especially late EPCs, play a critical role in endothelial maintenance and repair, and postnatal vasculogenesis. Although the actin cytoskeleton has been considered as a modulator that controls the function and modulation of stem cells, its role in the function of EPCs, and in particular late EPCs, remains poorly understood.

Methodology/Principal Finding

Bone marrow-derived late EPCs were treated with jasplakinolide, a compound that stabilizes actin filaments. Cell apoptosis, proliferation, adhesion, migration, tube formation, nitric oxide (NO) production and endothelial NO synthase (eNOS) phosphorylation were subsequently assayed in vitro. Moreover, EPCs were locally infused into freshly balloon-injured carotid arteries, and the reendothelialization capacity was evaluated after 14 days. Jasplakinolide affected the actin distribution of late EPCs in a concentration and time dependent manner, and a moderate concentration of (100 nmol/l) jasplakinolide directly stabilized the actin filament of late EPCs. Actin stabilization by jasplakinolide enhanced the late EPC apoptosis induced by VEGF deprivation, and significantly impaired late EPC proliferation, adhesion, migration and tube formation. Furthermore, jasplakinolide attenuated the reendothelialization capacity of transplanted EPCs in the injured arterial segment in vivo. However, eNOS phosphorylation and NO production were increased in late EPCs treated with jasplakinolide. NO donor sodium nitroprusside (SNP) rescued the functional activities of jasplakinolide-stressed late EPCs while the endothelial NO synthase inhibitor L-NAME led to a further dysfunction induced by jasplakinolide in late EPCs.

Conclusions/Significance

A moderate concentration of jasplakinolide results in an accumulation of actin filaments, enhancing the apoptosis induced by cytokine deprivation, and impairing the proliferation and function of late EPCs both in vitro and in vivo. NO donor reverses these impairments, suggesting the role of NO-related mechanisms in jasplakinolide-induced EPC downregulation. Actin cytoskeleton may thus play a pivotal role in regulating late EPC function.     

Endothelial Progenitor Cell Function,Apoptosis, and Telomere Length in Overweight/Obese Humans

Excess adiposity is associated with increased cardiovascular morbidity and mortality. Endothelial progenitor cells (EPCs) play an important role in vascular repair. We tested the hypothesis that increased adiposity is associated with EPC dysfunction, characterized by diminished capacity to release angiogenic cytokines, increased apoptotic susceptibility, reduced cell migration, and shorter telomere length. A total of 67 middle‐aged and older adults (42–67 years) were studied: 25 normal weight (normal weight; BMI: 18.5–24.9 kg/m²) and 42 overweight/obese (overweight/obese; BMI: 25.0–34.9 kg/m²). Cells with phenotypic EPC characteristics were isolated from peripheral blood. EPC release of vascular endothelial growth factor (VEGF) and granulocyte colony–stimulating factor (G‐CSF) was determined in the absence and presence of phytohemagglutinin (10 µg/ml). Intracellular active caspase‐3 and cytochrome c concentrations were determined by immunoassay. Migratory activity of EPCs in response to VEGF (2 ng/ml) and stromal cell–derived factor‐1α (SDF‐1α; 10 ng/ml) was determined by Boyden chamber. Telomere length was assessed by Southern hybridization. Phytohemagglutinin‐stimulated release of VEGF (90.6 ± 7.6 vs. 127.2 ± 11.6 pg/ml) and G‐CSF (896.1 ± 77.4 vs. 1,176.3 ± 126.3 pg/ml) was ～25% lower (P < 0.05) in EPCs from overweight/obese vs. normal weight subjects. Staurosporine induced a ～30% greater (P < 0.05) increase in active caspase‐3 in EPCs from overweight/obese (2.8 ± 0.2 ng/ml) compared with normal weight (2.2 ± 0.2) subjects. There were no significant differences in EPC migration to either VEGF or SDF‐1α. Telomere length did not differ between groups. These results indicate that increased adiposity adversely affects the ability of EPCs to release proangiogenic cytokines and resist apoptosis, potentially compromising their reparative potential.     

Role of integrin-linked kinase for functional capacity of endothelial progenitor cells in patients with stable coronary artery disease

Number and function of endothelial progenitor cells (EPCs) are down-regulated in patients with coronary artery disease (CAD). Integrin-linked kinase (ILK) is a signal and adaptor protein that regulates survival of mature endothelial cells and vascular development.Here we show that EPC dysfunction in patients with CAD is paralleled by down-regulation of ILK while restoration of ILK expression rescues the migratory defect of CAD-EPCs. Human EPCs transduced with dominant-negative ILK (DN-ILK) display significantly reduced expression of CD34⁺/VEGFR-2⁺, DiI-Ac-LDL uptake, and Ulex europaeus lectin binding. Mechanistically, DN-ILK-transfected EPCs are characterized by decreased proliferation, while proliferation is increased in wild-type ILK-transfected EPCs. These effects are paralleled by changes in cyclin D1 expression, colony forming units, and cytoskeletal rearrangement. Functionally, ILK is necessary and sufficient for SDF-1-triggered migration and adhesion in EPCs.These data extend current knowledge about the role of ILK in EPC biology and implicate ILK as a therapeutic target in CAD.