Circulating Angiogenic Cells Function Research Articles

Identification of Macrophage Genotype and Key Biological Pathways in Circulating Angiogenic Cell Transcriptome.

Background Circulating angiogenic cells (CAC) have been identified as important regulators of vascular biology. However, there is still considerable debate about the genotype and function of CAC. Methods and Results Data from publicly available gene expression data sets were used to analyse the transcriptome of in vitro cultured CAC (CACiv). Genes and pathways of interest were further evaluated using qPCR comparing CACiv versus CD14+ monocytic cells. The CACiv transcriptome strongly related to tissue macrophages, and more specifically to regulatory M2c macrophages. The cytokine expression profile of CACiv was predominantly immune modulatory and resembled the cytokine expression of tumor-associated macrophages (TAM). Pathway analysis revealed previously unrecognized biological processes in CACiv, such as riboflavin metabolism and liver X receptor (LXR)/retinoid X receptor (RXR) and farnesoid X receptor (FXR)/retinoid X receptor (RXR) pathways. Analysis of endothelial-specific genes did not show evidence for endothelial transdifferentiation. Conclusions CACiv are genotypically similar to regulatory M2c macrophages and lack signs of endothelial differentiation.

Exercise and Cardiovascular Progenitor Cells.

Autologous stem/progenitor cell-based methods to restore blood flow and function to ischemic tissues are clinically appealing for the substantial proportion of the population with cardiovascular diseases. Early preclinical and case studies established the therapeutic potential of autologous cell therapies for neovascularization in ischemic tissues. However, trials over the past ∼15years reveal the benefits of such therapies to be much smaller than originally estimated and a definitive clinical benefit is yet to be established. Recently, there has been an emphasis on improving the number and function of cells [herein generally referred to as circulating angiogenic cells (CACs)] used for autologous cell therapies. CACs include of several subsets of circulating cells, including endothelial progenitor cells, with proangiogenic potential that is largely exerted through paracrine functions. As exercise is known to improve CV outcomes such as angiogenesis and endothelial function, much attention is being given to exercise to improve the number and function of CACs. Accordingly, there is a growing body of evidence that acute, short-term, and chronic exercise have beneficial effects on the number and function of different subsets of CACs. In particular, recent studies show that aerobic exercise training can increase the number of CACs in circulation and enhance the function of isolated CACs as assessed in ex vivo assays. This review summarizes the roles of different subsets of CACs and the effects of acute and chronic exercise on CAC number and function, with a focus on the number and paracrine function of circulating CD34+ cells, CD31+ cells, and CD62E+ cells. © 2019 American Physiological Society. Compr Physiol 9:767-797, 2019.

Stearic acid at physiologic concentrations induces in vitro lipotoxicity in circulating angiogenic cells

Background and aimsSaturated free fatty acids (SFAs) can induce lipotoxicity in different cells. No studies have investigated the effects of SFA in circulating angiogenic cells (CACs), which play a key role in endothelial repair processes. The aim of the study was to assess the effects of SFAs, specifically stearic acid (SA), on viability and function of CACs and to investigate potential underlying molecular mechanisms. MethodsCACs were isolated from healthy subjects by established methods. CACs were incubated with BSA-complexed stearate (100 μM) to assess the time course (from 8 to 24 h exposure) of the effects on viability and apoptosis (activation of caspases 3/7), angiogenic function (tube formation assay), pro-inflammatory cytokine (IL-1β, IL-6, IL-8, MCP-1 and TNFα) gene expression (qPCR) and secretion (ELISA), activation of MAPK (JNK, p38 and Erk1/2) by Western blot and endoplasmic reticulum (ER) stress marker (CHOP, BIP, ATF4, XBP-1 and sXBP-1) gene expression by qPCR. ResultsStearic acid activates effector caspases in CACs in a dose- and time-dependent manner. SA also impairs CAC function and increases pro-inflammatory molecule (IL-1β, IL-6, IL-8, MCP-1 and TNFα) gene expression and secretion in CACs starting from 3 h of incubation. The activation of JNK by SA mediates pro-inflammatory response, but it may be not necessary for apoptosis. Moreover, SA induces the expression of ER stress markers across the three branches of the ER stress response. ConclusionsIn humans, both function and viability of CACs are exquisitely vulnerable to physiologic concentrations of stearate; lipotoxic impairment of endothelial repair processes may be implicated in vascular damage caused by SFAs.

Investigating the extremes of the continuum of paracrine functions in CD34-/CD31+ CACs across diverse populations.

S100A8 and S100A9 proteins in concentrations secreted by CD34-/CD31+ circulating angiogenic cells (CACs) with impaired function reduce endothelial cell capillary-like network formation. These effects appear to be mediated by Toll-like receptor 4 and are absent with S100A8 and S100A9 in concentrations secreted by healthy CD34-/CD31+ CACs.

IFN-β affects the angiogenic potential of circulating angiogenic cells by activating calpain 1.

Circulating angiogenic cells (CACs) are monocyte-derived cells with endothelial characteristics, which contribute to both angiogenesis and arteriogenesis in a paracrine way. Interferon-β (IFN-β) is known to inhibit these divergent processes in animals and patients. We hypothesized that IFN-β might act by affecting the differentiation and function of CACs. CACs were cultured from peripheral blood mononuclear cells and phenotypically characterized by surface expression of monocytic and endothelial markers. IFN-β significantly reduced the number of CACs by 18-64%. Apoptosis was not induced by IFN-β, neither in mononuclear cells during differentiation, nor after maturation to CACs. Rather, IFN-β impaired adhesion to, and spreading on, fibronectin, which was dependent on α5β1 (VLA-5)-integrin. IFN-β affected the function of VLA-5 in mature CACs, leading to rounding and detachment of cells, by induction of calpain 1 activity. Cell rounding and detachment was completely reversed by inhibition of calpain 1 activity in mature CACs. During in vitro capillary formation, CAC addition and calpain 1 inhibition enhanced sprouting of endothelial cells to a comparable extent, but were not sufficient to rescue tube formation in the presence of IFN-β. We show that the IFN-β-induced reduction of the numbers of in vitro differentiated CACs is based on activation of calpain 1, resulting in an attenuated adhesion to extracellular matrix proteins via VLA-5. In vivo, this could lead to inhibition of vessel formation due to reduction of the locally recruited CAC numbers and their paracrine angiogenic factors.

Effects of TiO2 and Co3O4 Nanoparticles on Circulating Angiogenic Cells

Background and AimSparse evidence suggests a possible link between exposure to airborne nanoparticles (NPs) and cardiovascular (CV) risk, perhaps through mechanisms involving oxidative stress and inflammation. We assessed the effects of TiO2 and Co3O4 NPs in human circulating angiogenic cells (CACs), which take part in vascular endothelium repair/replacement.MethodsCACs were isolated from healthy donors’ buffy coats after culturing lymphomonocytes on fibronectin-coated dishes in endothelial medium for 7 days. CACs were pre-incubated with increasing concentration of TiO2 and Co3O4 (from 1 to 100 μg/ml) to test the effects of NP – characterized by Transmission Electron Microscopy – on CAC viability, apoptosis (caspase 3/7 activation), function (fibronectin adhesion assay), oxidative stress and inflammatory cytokine gene expression.ResultsNeither oxidative stress nor cell death were associated with exposure to TiO2 NP (except at the highest concentration tested), which, however, induced a higher pro-inflammatory effect compared to Co3O4 NPs (p<0.01). Exposure to Co3O4 NPs significantly reduced cell viability (p<0.01) and increased caspase activity (p<0.01), lipid peroxidation end-products (p<0.05) and pro-inflammatory cytokine gene expression (p<0.05 or lower). Notably, CAC functional activity was impaired after exposure to both TiO2 (p<0.05 or lower) and Co3O4 (p<0.01) NPs.ConclusionsIn vitro exposure to TiO2 and Co3O4 NPs exerts detrimental effects on CAC viability and function, possibly mediated by accelerated apoptosis, increased oxidant stress (Co3O4 NPs only) and enhancement of inflammatory pathways (both TiO2 and Co3O4 NPs). Such adverse effects may be relevant for a potential role of exposure to TiO2 and Co3O4 NPs in enhancing CV risk in humans.

Collagen matrix-induced expression of integrin αVβ3 in circulating angiogenic cells can be targeted by matricellular protein CCN1 to enhance their function.

Circulating angiogenic cells (CACs) play an important role in vascular homeostasis and hold therapeutic promise for treating a variety of cardiovascular diseases. However, further improvements are needed because the effects of CAC therapy remain minimal or transient. The regenerative potential of these cells can be improved by culture on a collagen-based matrix through the up-regulation of key integrin proteins. We found that human CAC function was enhanced by using the matricellular protein CCN1 (CYR61/CTGF/NOV family member 1) to target integrin αV and β3, which are up-regulated on matrix. Compared to matrix-cultured CACs, CCN1-matrix CACs exhibited a 2.2-fold increase in cell proliferation, 1.8-fold greater migration toward VEGF, and 1.7-fold more incorporation into capillary-like structures in an angiogenesis assay. In vivo, intramuscular injection of CCN1-matrix-cultured CACs into ischemic hind limbs of CD-1 nude mice resulted in blood flow recovery to 80% of baseline, which was greater than matrix-cultured CACs (66%) and PBS (35%) treatment groups. Furthermore, transplanted CCN1-matrix CACs exhibited greater engraftment (11-fold) and stimulated the up-regulation of survival and angiogenic genes (>3-fold). These findings reveal the importance of cell-matrix interactions in regulating CAC function and also reveal a mechanism by which these may be exploited to enhance cell therapies for ischemic disease.

The dual PPARα/γ agonist aleglitazar increases the number and function of endothelial progenitor cells: implications for vascular function and atherogenesis

Aleglitazar is a dual PPARα/γ agonist but little is known about its effects on vascular function and atherogenesis. Hence, we characterized its effects on circulating angiogenic cells (CAC), neoangiogenesis, endothelial function, arteriogenesis and atherosclerosis in mice. C57Bl/6 wild-type (WT, normal chow), endothelial NOS (eNOS)(-/-) (normal chow) and ApoE(-/-) (Western-type diet) mice were treated with aleglitazar (10 mg·kg(-1) ·day(-1) , i.p.) or vehicle. Aleglitazar enhanced expression of PPARα and PPARγ target genes, normalized glucose tolerance and potently reduced hepatic fat in ApoE(-/-) mice. In WT mice, but not in eNOS(-/-) , aleglitazar up-regulated Sca-1/VEGFR2-positive CAC in the blood and bone marrow and up-regulated diLDL/lectin-positive CAC. Aleglitazar augmented CAC migration and enhanced neoangiogenesis. In ApoE(-/-) mice, aleglitazar up-regulated CAC number and function, reduced markers of vascular inflammation and potently improved perfusion restoration after hindlimb ischaemia and aortic endothelium-dependent vasodilatation. This was associated with markedly reduced formation of atherosclerotic plaques. In human cultured CAC from healthy donors and patients with coronary artery disease with or without diabetes mellitus, aleglitazar increased migration and colony-forming units in a concentration-dependent manner. Furthermore, oxidative stress-induced CAC apoptosis and expression of p53 were reduced, while telomerase activity and expression of phospho-eNOS and phospho-Akt were elevated. Comparative agonist and inhibitor experiments revealed that aleglitazar's effects on CAC migration and colony-forming units were mediated by both PPARα and PPARγ signalling and required Akt. Aleglitazar augments the number, function and survival of CAC, which correlates with improved vascular function, enhanced arteriogenesis and prevention of atherosclerosis in mice.

The role of integrin α2 in cell and matrix therapy that improves perfusion, viability and function of infarcted myocardium

Injectable delivery matrices hold promise in enhancing engraftment and the overall efficacy of cardiac cell therapies; however, the mechanisms responsible remain largely unknown. Here we studied the interaction of a collagen matrix with circulating angiogenic cells (CACs) in a mouse myocardial infarction model. CACs + matrix treatment enhanced CAC engraftment, and improved myocardial perfusion, viability and function compared to cells or matrix alone. Integrin-linked kinase (ILK) was up-regulated in matrix-cultured CACs. Integrin α2β1 blocking prevented ILK up-regulation, significantly reduced the adhesion, proliferation, and paracrine properties of matrix-cultured CACs, and negated the benefits of CACs + matrix therapy in vivo. Furthermore, integrin α5 was essential for the angiogenic potential of CACs on matrix. These findings indicate that the synergistic therapeutic effect of CACs + matrix therapy in MI requires the matrix to enhance CAC function via α2β1 and α5 integrin signaling mechanisms, rather than simply delivering the cells.

Circulating Angiogenic Cell Dysfunction in Patients with Hereditary Hemorrhagic Telangiectasia

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant vascular disorder. Circulating angiogenic cells (CACs) play an important role in vascular repair and regeneration. This study was designed to examine the function of CACs derived from patients with HHT. Peripheral blood mononuclear cells (PBMNCs) isolated from patients with HHT and age- and gender-matched healthy volunteers were assessed for expression of CD34, CD133 and VEGF receptor 2 by flow cytometry. PBMNCs were cultured to procure early outgrowth CACs. Development of endothelial cell (EC) phenotype in CACs was analyzed by fluorescence microscopy. CAC apoptosis was assayed with Annexin V staining, and CAC migration assessed by a modified Boyden chamber assay. mRNA expression of endoglin (ENG), activin receptor-like kinase-1 (ACVLR1 or ALK1) and endothelial nitric oxide synthase (eNOS) in CACs was measured by real time RT-PCR. The percentage of CD34+ cells in PBMNCs from HHT patients was significantly higher than in PBMNCs of healthy controls. CACs derived from patients with HHT not only showed a significant reduction in EC-selective surface markers following 7-day culture, but also a significant increase in the rate of basal apoptosis and blunted migration in response to vascular endothelial growth factor and stromal cell-derived factor-1. CACs from HHT patients expressed significantly lower levels of ENG, ALK1 and eNOS mRNAs. In conclusion, CACs from patients with HHT exhibited various functional impairments, suggesting a reduced regenerative capacity of CACs to repair the vascular lesions seen in HHT patients.

Integrin α5 Mediates the Pro-Angiogenic Potency of Circulating Angiogenic Cells Cultured on Collagen Matrices

Cell therapy using circulating angiogenic cells (CACs) has the ability to promote neovascularization and restore blood flow to ischemic tissue; however, cell retention and engraftment remain significant hurdles. To address these challenges, we have developed a collagen-based matrix to deliver CACs. We previously showed that the function of CACs is improved with culture on the matrix, but the mechanisms responsible remain unknown. The aim of this study was to investigate the involvement of integrins in the matrix-enhanced function of CACs.

Collagen Matrices Enhance Circulating Angiogenic Cell Function Through Increased Sensitivity to Cysteine-Rich Angiogenic Inducer 61 (CYR-61) Via Integrin αV

Revascularization following myocardial infarction using circulating angiogenic cells (CACs) may serve to prevent/limit cardiac damage. We have previously shown that culturing CACs on a collagen matrix enhanced their function and therapeutic potential compared to traditional fibronectin culture. Since integrins are major receptors that link the extracellular environment and intracellular signaling pathways, the aim of this study was to investigate the role of integrins in enhancing CAC function.

Interferon α impairs survival and function of circulating angiogenic cells in vitro: a model of failed endothelial repair in systemic lupus erythematosus

BackgroundPatients with systemic lupus erythematosus have an increased risk of cardiovascular disease. Interferon α may impair endothelial repair mechanisms in lupus. Patients with systemic lupus erythematosus have fewer circulating angiogenic cells (CAC) and endothelial progenitor cells (EPCs). Mixed EPC and CAC populations have been shown to be sensitive to interferon α (IFNα). We aimed to investigate the effects of IFNα2b on an in-vitro model of angiogenesis and vascular repair. MethodsPeripheral blood mononuclear cells from healthy individuals were cultured on human fibronectin for 7 days. CAC phenotype was confirmed by low-density lipoprotein (LDL) uptake, lectin binding, and cell surface marker expression by RT-PCR. Cell survival in response to IFNα2b (0·01–10 ng/mL) was determined by the number of LDL-uptake-positive cells. To study CAC function, supernatant from CACs in the presence or absence of 10ng/mL IFNα2b was added to human aortic endothelial cells on Matrigel. Tubule formation was assessed at 14 h. FindingsCACs expressed markers of endothelial (CD31) and myeloid lineage (CD14, CD45), and strongly expressed the markers CD68, CD163, and CD206 suggesting an alternatively activated (M2) macrophage phenotype. IFNα2b significantly reduced the number of CACs at day 7 in a dose-dependent manner (r2= −0·769, p<0·0001). In co-culture with endothelial cells on Matrigel, CACs co-localised to the endothelial tubules but did not form tubule networks alone. CAC supernatant significantly increased tubule network density in terms of total pixel area (mean 27 781 [SE 1469] vs 36 283 [1804], p=0·0065), number of branches (340·2 [11·0] vs 510·6 [70·2], p=0·0434), junctions (162·4 [5·9 vs 241·1 [28·0], p=0·0104), and closed loops (21·8 [1·9] vs 38·3 [2·2], p=0·005). IFNα2b significantly reduced the number of closed loops (38·2 [2·2] vs 24·1 [3·5], p=0·0094). All other network parameters were reduced by IFNα2b but did not reach statistical significance. InterpretationCACs are of myeloid lineage, and CAC supernatant contains potent angiogenic factors which augment endothelial tubule networks. IFNα2b dramatically reduces the survival of CACs in vitro, resulting in reduced tubule network formation and may be a mechanism by which IFNα promotes vascular damage in systemic lupus erythematosus. FundingNorth West England Medical Research Council Fellowship Scheme in Clinical Pharmacology and Therapeutics.

Pathologic endothelial response and impaired function of circulating angiogenic cells in patients with Fabry disease

Fabry disease is an X-chromosomal recessive deficiency of the lysosomal hydrolase alpha-galactosidase A (alpha-Gal). This results in an accumulation of globotriaosylceramide (GL-3) in a variety of cells often with subsequent functional impairment. Here, the impact of Fabry disease on the biology of circulating angiogenic cells (CACs) and the endothelial response to transient ischemia was investigated. Untreated patients with Fabry disease (n = 26), patients after initiation of alpha-Gal enzyme replacement therapy (ERT) (n = 16) and healthy controls (n = 26) were investigated. Endothelial function was assessed by the EndoPAT2000 device. CAC numbers were assessed by flow-cytometry, CAC function by a modified Boyden chamber assay. Fabry patients showed a pathologic endothelial response, which normalized after ERT. CACs were increased in number, but functionally impaired. Immunofluorescence and electron microscopy identified an accumulation of GL-3 in Fabry CACs. ERT attenuated CAC dysfunction and improved markers of oxidative stress response in Fabry patients via a reduction in GL-3 accumulation in vitro and in vivo. Silencing of alpha-Gal in healthy CACs impaired their migratory capacity underlining a key role of this enzyme for CAC function. CAC supernatant as well as CACs from Fabry patients impaired angiogenesis and migratory capacity of HUVECs providing a mechanistic link between CAC and endothelial dysfunction. CAC adhesion to TNF-α pre-stimulated HUVECs and tube formation was impaired by alpha-Gal knockdown. Fabry patients show a dysfunction of CAC and a pathologic endothelial response. ERT improves CAC and endothelial function and thus may attenuate development of cardiovascular disease in the long term in this patient population.

Abstract 509: Identification of Macrophage Phenotype and Key Biological Pathways in Circulating Angiogenic Cell Transcriptome

Objective: Circulating angiogenic cells (CACs) have been identified as important regulators of vascular biology. However, there is still considerable debate about the phenotype and function of CACs. Methods and results: DNA microarray transcriptome analysis was used to study the CAC phenotype and investigate the key biological pathways in CAC biology. We determined a CAC-specific gene signature, which revealed the close relationship between CACs and members of the macrophage lineage, such as tumor-associated macrophages (TAMs), dendritic cells (DCs), osteoclasts and atherosclerotic plaque-remodeling macrophages. We identified CCL2, CCL17, CCL18 and CCL22 as a chemokinetic fingerprint for CACs and indicated cholesterol transporter pathways (LXR/RXR, FXR/RXR) and riboflavin metabolism as highly important biological pathways in the CAC transcriptome. These findings, together with upregulation of the chemokine receptor CCR7, suggest the involvement of CACs in reverse plaque remodeling. Conclusions: CACs are phenotypically similar to regulatory M2 macrophages and lack signs of endothelial differentiation. However, the expression profile of CACs prompted us to suggest new mechanisms by which these cells could engage in cardiovascular pathophysiology, such as immunomodulation, tissue remodeling, enhancement of cholesterol efflux and vasculoprotection. These findings expand the current knowledge on CAC biology and suggest new indications for CAC therapy in future clinical trials.

Exercise to the rescue

Exercise to the rescue

Improved Function of Circulating Angiogenic Cells Is Evident in Type 1 Diabetic Islet-Transplanted Patients

Circulating angiogenic cells (CACs) are vascular-committed bone marrow-derived cells that are dysfunctional in type 1 diabetes (T1D). Here we studied whether restoration of normoglycemia following islet transplantation is associated with better CAC function. We carried out a cross-sectional study of 18 T1D patients, 14 insulin-independent islet-transplanted patients (ITA) and 14 healthy controls (C) evaluating in vivo and in vitro CACs viability and function. We found that the percentage of CACs in vivo did not differ among the three groups while the number of CAC colonies obtained from T1D, but not from ITA, was reduced compared to C (C = 7.3 ± 1.9, T1D = 0.9 ± 0.4 and ITA = 4.7 ± 1.9; p < 0.05 T1D vs. all). In vitro CAC migration/differentiation were similar, while in vivo an improved angiogenic ability of ITA compared to T1D was shown (capillary density: C = 93.5 ± 22.1, T1D = 19.2 ± 2.8 and ITA = 44.0 ± 10.5, p < 0.05 T1D vs. all). Increased apoptosis and lesser IL-8 secretion were evident in CACs obtained from T1D compared to C and ITA. in vitro addition of anti-hIL-8 reduced the number of colonies obtained from C. Finally, T1D, but not ITA, had a lower endothelial-dependent dilatation (EDD) compared with C. These data suggest that CAC function is altered in T1D and may be improved after islet transplantation.

Nitric Oxide Synthase Expression and Functional Response to Nitric Oxide Are Both Important Modulators of Circulating Angiogenic Cell Response to Angiogenic Stimuli

Circulating angiogenic cells (CACs), also termed endothelial progenitor cells, play an integral role in vascular repair and are functionally impaired in coronary artery disease (CAD). The role of nitric oxide (NO) in CAC function is poorly understood. We hypothesized that CAC migration toward angiogenic signals is modulated by both NO synthase (NOS) expression and functional response to NO. Similar to endothelial cells, CAC chemotaxis to vascular endothelial growth factor (VEGF) was blocked by inhibition of NOS, phosphatidylinositol 3-kinase, or guanylyl cyclase or by treatment with an NO scavenger. Addition of an NO donor (S-nitroso-N-acetylpenicillamine) and the NOS substrate l-arginine increased random cell migration (chemokinesis) and enhanced VEGF-dependent chemotaxis. Healthy CACs expressed endothelial NOS, but endothelial NOS was not detected in CAD patient CACs. Both chemokinesis and chemotaxis to VEGF of patient CACs were decreased compared with healthy CACs but were restored to healthy values by S-nitroso-N-acetylpenicillamine. In parallel, CAD patients exhibited lower flow-mediated vasodilation and plasma NO source nitrite than young, healthy subjects, indicating endothelial dysfunction with reduced NO bioavailability. NOS activity is required for CAC chemotaxis. In CAD patients, impairment of NOS expression and NO bioavailability, rather than response to NO, may contribute to dysfunction of CACs and limit their regenerative capacity.

Exercise training improves function of circulating angiogenic cells in patients with chronic heart failure

Alterations in circulating angiogenic cells (CAC) and endothelial progenitor cells (EPC), known to contribute to endothelial repair, could explain the reversal of endothelial function in response to exercise training. Moreover, training-induced vascular remodeling might affect the acute response of EPC and CAC following a single exercise bout. We studied the impact of exercise training on CAC function and numbers of CD34(+)/KDR(+) EPC in patients with chronic heart failure (CHF) and we assessed the effect of acute exercise on CAC and EPC in sedentary and trained patients. Twenty-one sedentary CHF patients underwent 6-month exercise training and were compared to a non-trained control group (n = 17) and 10 healthy age-matched subjects. At baseline and follow-up, flow-mediated dilation was assessed and graded exercise testing (GXT) was performed. Before and immediately after GXT, CAC migratory capacity was assessed in vitro and circulating CD34(+)/KDR(+) EPC were quantified using flow cytometry. At baseline, CAC migration was significantly impaired in sedentary CHF patients but normalized acutely after GXT. Training corrected endothelial dysfunction, which coincided with a 77% increase in CAC migration (P = 0.0001). Moreover, the GXT-induced improvement detected at baseline was no longer observed after training. Numbers of CD34(+)/KDR(+) EPC increased following 6-month exercise training (P = 0.021), but were not affected by GXT, either prior or post-training. In conclusion, the present findings demonstrate for the first time that exercise training in CHF reverses CAC dysfunction and increases numbers of CD34(+)/KDR(+) EPC, which is accompanied by improvement of peripheral endothelial function. The acute exercise-induced changes in CAC function wane with exercise training, suggesting that repetitive exercise bouts progressively lead to functional endothelial repair.

Exercise acutely reverses dysfunction of circulating angiogenic cells in chronic heart failure

Recruitment of endothelial progenitor cells (EPCs) and enhanced activity of circulating angiogenic cells (CACs) might explain the benefits of exercise training in reversing endothelial dysfunction in chronic heart failure (CHF) patients. We studied baseline EPC numbers and CAC function and the effect of a single exercise bout. Forty-one CHF patients (mild, n = 22; severe, n = 19) and 13 healthy subjects were included. Migratory activity of CACs was evaluated in vitro and circulating CD34+ and CD34+/KDR+ (EPC) cells were quantified by flow cytometry before and after cardiopulmonary exercise testing (CPET). Circulating stromal cell-derived factor-1alpha (SDF-1alpha) and vascular endothelial growth factor (VEGF) concentrations were measured. Both CAC migration as well as CD34+ cell numbers were significantly reduced in CHF, whereas CD34+/KDR+ cells were not different from controls. Endothelial dysfunction was related to impaired CAC migration (r = 0.318, P = 0.023). Cardiopulmonary exercise testing improved CAC migration in severe (+52%, P < 0.005) and mild CHF (+31%, P < 0.005), restoring it to levels similar to controls. Following CPET, SDF-1alpha increased in healthy controls and mild CHF (P < 0.005). Vascular endothelial growth factor, CD34+, and CD34+/KDR+ cell numbers remained unchanged. The present findings reveal a potent stimulus of acute exercise to reverse CAC dysfunction in CHF patients with endothelial dysfunction.