Sylvie Briand

AngiomiR-126 expression and secretion from circulating CD34(+) and CD14(+) PBMCs: role for proangiogenic effects and alterations in type 2 diabetics.

Several peripheral blood mononuclear cell (PBMC)-derived cell populations can promote angiogenesis, and differences in CD34(+) or CD14(+) surface expression have been used to separate PBMC subpopulations in this respect. AngiomiRs, microRNAs regulating angiogenesis, are key regulators of angiogenic processes. The present study examines differential angiomiR expression/secretion from CD34(+)/CD14(+), CD34(+)/CD14(-), CD34(-)/CD14(+), and CD34(-)/CD14(-) PBMC subsets and their relevance for different proangiogenic properties. Notably, both circulating human CD34(+)/14(+) and CD34(+)/14(-) PBMC subsets and their supernatants exerted more potent proangiogenic effects compared with CD34(-) PBMC subsets. MiR-126 was identified as most differentially expressed angiomiR in CD34(+) compared with CD34(-) PBMC subsets, determined by miR-array and RT-PCR validation. Modulation of miR-126 by anti-miR-126 or miR-mimic-126 treatment resulted in significant loss or increase of proangiogenic effects of CD34(+) PBMCs. MiR-126 levels in supernatants of CD34(+) PBMC subsets were substantially higher compared with CD34(-) PBMC subsets. MiR-126 was secreted in microvesicles/exosomes, and inhibition of their release impaired CD34(+) PBMCs proangiogenic effects. Notably, high-glucose treatment or diabetes reduced miR-126 levels of CD34(+) PBMCs, associated with impaired proangiogenic properties that could be rescued by miR-mimic-126 treatment. The present findings provide a novel molecular mechanism underlying increased proangiogenic effects of CD34(+) PBMCs, that is, angiomiR-126 expression/secretion. Moreover, an alteration of angiomiR-126 expression in CD34(+) PBMCs in diabetes provides a novel pathway causing impaired proangiogenic effects.

AngiomiR-126 expression and secretion from circulating CD34+ and CD14+ PBMCs: role for pro-angiogenic effects and alterations in type-2 diabetics

Several peripheral blood mononuclear cell (PBMC)-derived cell populations can promote angiogenesis, and differences in CD34+ or CD14+ surface-expression have been used to separate PBMC-subpopulations in this respect. AngiomiRs, microRNAs regulating angiogenesis, have been identified as key regulators of angiogenic processes. The present study examines differential angiomiR-expression/secretion from CD34+/CD14+; CD34+/CD14-; CD34-/CD14+; CD34-/CD14- PBMC-subsets and their relevance for different pro-angiogenic properties. Notably, both circulating human CD34+/14+ and CD34+/14- PBMC-subsets and their supernatants exerted more potent pro-angiogenic effects as compared to CD34-PBMC-subsets. MiR-126 was identified as most differentially expressed angiomiR in CD34+ as compared to CD34-PBMC-subsets, determined by miR-array and RT-PCR validation. Modulation of miR-126 by anti-miR-126 or miR-mimic-126 treatment resulted in significant loss or increase of pro-angiogenic effects of CD34+PBMCs. MiR-126 levels in supernatants of CD34+PBMC-subsets were substantially higher as compared to CD34- PBMC-subsets. MiR-126 was secreted in microvesicles/exosomes, and inhibition of their release impaired CD34+PBMCs pro-angiogenic effects. Notably, high-glucose treatment or diabetes reduced miR-126-levels of CD34+PBMCs, associated with impaired pro-angiogenic properties that could be rescued by miR-mimic-126 treatment. The present findings provide a novel molecular mechanism underlying increased pro-angiogenic effects of CD34+PBMC-subpopulations, i.e. angiomiR-126 expression/secretion. Moreover, an alteration of angiomiR-126-expression in CD34+PBMC in diabetes provides a new pathway causing impaired pro-angiogenic effects.

Loss of AngiomiR-126 and 130a in Angiogenic Early Outgrowth Cells From Patients With Chronic Heart Failure Role for Impaired In Vivo Neovascularization and Cardiac Repair Capacity

Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection.Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection. # Clinical Perspective {#article-title-54}

Deletion of the Activated Protein-1 Transcription Factor JunD Induces Oxidative Stress and Accelerates Age-Related Endothelial Dysfunction

Background— Reactive oxygen species are major determinants of vascular aging. JunD, a member of the activated protein-1 family of transcription factors, is emerging as a major gatekeeper against oxidative stress. However, its contribution to reactive oxygen species homeostasis in the vasculature remains unknown. Methods and Results— Endothelium-dependent vasorelaxation was impaired in young and old JunD−/− mice (6 and 22 months old) compared with age-matched wild-type mice. JunD−/− mice displayed an age-independent decline in endothelial nitric oxide release and endothelial nitric oxide synthase activity and increased mitochondrial superoxide formation and peroxynitrite levels. Furthermore, vascular expression and activity of the free radical scavengers manganese and extracellular superoxide dismutase and aldehyde dehydrogenase 2 were reduced, whereas the NADPH oxidase subunits p47phox, Nox2, and Nox4 were upregulated. These redox changes were associated with premature vascular aging, as shown by reduced telomerase activity, increased &bgr;-galactosidase–positive cells, upregulation of the senescence markers p16INK4a and p53, and mitochondrial disruption. Interestingly, old wild-type mice showed a reduction in JunD expression and transcriptional activity resulting from promoter hypermethylation and binding with tumor suppressor menin, respectively. In contrast, JunD overexpression blunted age-induced endothelial dysfunction. In human endothelial cells, JunD knockdown exerted a similar impairment of the O2−/nitric oxide balance that was prevented by concomitant NADPH inhibition. In parallel, JunD expression was reduced in monocytes from old versus young healthy subjects and correlated with mRNA levels of scavenging and oxidant enzymes. Conclusions— JunD provides protection in aging-induced endothelial dysfunction and may represent a novel target to prevent reactive oxygen species–driven vascular aging.

Deletion of the AP-1 Transcription Factor JunD Induces Oxidative Stress and Accelerates Age-Related Endothelial Dysfunction

Background— Reactive oxygen species are major determinants of vascular aging. JunD, a member of the activated protein-1 family of transcription factors, is emerging as a major gatekeeper against oxidative stress. However, its contribution to reactive oxygen species homeostasis in the vasculature remains unknown. Methods and Results— Endothelium-dependent vasorelaxation was impaired in young and old JunD−/− mice (6 and 22 months old) compared with age-matched wild-type mice. JunD−/− mice displayed an age-independent decline in endothelial nitric oxide release and endothelial nitric oxide synthase activity and increased mitochondrial superoxide formation and peroxynitrite levels. Furthermore, vascular expression and activity of the free radical scavengers manganese and extracellular superoxide dismutase and aldehyde dehydrogenase 2 were reduced, whereas the NADPH oxidase subunits p47phox, Nox2, and Nox4 were upregulated. These redox changes were associated with premature vascular aging, as shown by reduced telomerase activity, increased &bgr;-galactosidase–positive cells, upregulation of the senescence markers p16INK4a and p53, and mitochondrial disruption. Interestingly, old wild-type mice showed a reduction in JunD expression and transcriptional activity resulting from promoter hypermethylation and binding with tumor suppressor menin, respectively. In contrast, JunD overexpression blunted age-induced endothelial dysfunction. In human endothelial cells, JunD knockdown exerted a similar impairment of the O2−/nitric oxide balance that was prevented by concomitant NADPH inhibition. In parallel, JunD expression was reduced in monocytes from old versus young healthy subjects and correlated with mRNA levels of scavenging and oxidant enzymes. Conclusions— JunD provides protection in aging-induced endothelial dysfunction and may represent a novel target to prevent reactive oxygen species–driven vascular aging.

Increased Proangiogenic Activity of Mobilized CD34+ Progenitor Cells of Patients With Acute ST-Segment-Elevation Myocardial Infarction: Role of Differential MicroRNA-378 Expression.

Objective— Proangiogenic effects of mobilized bone marrow–derived stem/progenitor cells are essential for cardiac repair after myocardial infarction. MicroRNAs (miRNA/miR) are key regulators of angiogenesis. We investigated the differential regulation of angio-miRs, that is, miRNAs regulating neovascularization, in mobilized CD34+ progenitor cells obtained from patients with an acute ST-segment–elevation myocardial infarction (STEMI) as compared with those with stable coronary artery disease or healthy subjects. Approach and Results— CD34+ progenitor cells were isolated from patients with STEMI (on day 0 and day 5), stable coronary artery disease, and healthy subjects (n=27). CD34+ progenitor cells of patients with STEMI exhibited increased proangiogenic activity as compared with CD34+ cells from the other groups. Using a polymerase chain reaction–based miRNA-array and real-time polymerase chain reaction validation, we identified a profound upregulation of 2 known angio-miRs, that are, miR-378 and let-7b, in CD34+ cells of patients with STEMI. Especially, we demonstrate that miR-378 is a critical regulator of the proangiogenic capacity of CD34+ progenitor cells and its stimulatory effects on endothelial cells in vitro and in vivo, whereas let-7b upregulation in CD34+ cells failed to proof its effect on endothelial cells in vivo. Conclusions— The present study demonstrates a significant upregulation of the angio-miRs miR-378 and let-7b in mobilized CD34+ progenitor cells of patients with STEMI. The increased proangiogenic activity of these cells in patients with STEMI and the observation that in particular miR-378 regulates the angiogenic capacity of CD34+ progenitor cells in vivo suggest that this unique miRNA expression pattern represents a novel endogenous repair mechanism activated in acute myocardial infarction.

Loss of AngiomiR-126 and 130a in Angiogenic Early Outgrowth Cells From Patients With Chronic Heart FailureClinical Perspective: Role for Impaired In Vivo Neovascularization and Cardiac Repair Capacity

Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection.Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection. # Clinical Perspective {#article-title-54}

Loss of AngiomiR-126 and 130a in Angiogenic Early Outgrowth Cells From Patients With Chronic Heart FailureClinical Perspective

Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection.Background— MicroRNAs are key regulators of angiogenic processes. Administration of angiogenic early outgrowth cells (EOCs) or CD34+ cells has been suggested to improve cardiac function after ischemic injury, in particular by promoting neovascularization. The present study therefore examines regulation of angiomiRs, microRNAs involved in angiogenesis, in angiogenic EOCs and circulating CD34+ cells from patients with chronic heart failure (CHF) and the role for their cardiac repair capacity. Methods and Results— Angiogenic EOCs and CD34+ cells were isolated from patients with CHF caused by ischemic cardiomyopathy (n=45) and healthy subjects (n=35). In flow cytometry analyses, angiogenic EOCs were largely myeloid and positive for alternatively activated M2 macrophage markers. In vivo cardiac neovascularization and functional repair capacity were examined after transplantation into nude mice with myocardial infarction. Cardiac transplantation of angiogenic EOCs from healthy subjects markedly increased neovascularization and improved cardiac function, whereas no such effect was observed after transplantation of angiogenic EOCs from patients with CHF. Real-time polymerase chain reaction analysis of 14 candidate angiomiRs, expressed in angiogenic EOCs, revealed a pronounced loss of angiomiR-126 and -130a in angiogenic EOCs from patients with CHF that was also observed in circulating CD34+ cells. Anti–miR-126 transfection markedly impaired the capacity of angiogenic EOCs from healthy subjects to improve cardiac function. miR-126 mimic transfection increased the capacity of angiogenic EOCs from patients with CHF to improve cardiac neovascularization and function. Conclusions— The present study reveals a loss of angiomiR-126 and -130a in angiogenic EOCs and circulating CD34+ cells from patients with CHF. Reduced miR-126 expression was identified as a novel mechanism limiting their capacity to improve cardiac neovascularization and function that can be targeted by miR-126 mimic transfection. # Clinical Perspective {#article-title-54}