The Potential Role of MiRs-139-5p and -454-3p in Endoglin-Knockdown-Induced Angiogenic Dysfunction in HUVECs.

Hereditary hemorrhagic telangiectasia (HHT) is a rare genetic disease characterized by aberrant angiogenesis and vascular malformations. Mutations in the transforming growth factor beta co-receptor, endoglin (ENG), account for approximately half of known HHT cases and cause abnormal angiogenic activity in endothelial cells (ECs). To date, how ENG deficiency contributes to EC dysfunction remains to be fully understood. MicroRNAs (miRNAs) regulate virtually every cellular process. We hypothesized that ENG depletion results in miRNA dysregulation that plays an important role in mediating EC dysfunction. Our goal was to test the hypothesis by identifying dysregulated miRNAs in ENG-knockdown human umbilical vein endothelial cells (HUVECs) and characterizing their potential role in EC function. We identified 32 potentially downregulated miRNAs in ENG-knockdown HUVECs with a TaqMan miRNA microarray. MiRs-139-5p and -454-3p were found to be significantly downregulated after RT-qPCR validation. While the inhibition of miR-139-5p or miR-454-3p had no effect on HUVEC viability, proliferation or apoptosis, angiogenic capacity was significantly compromised as determined by a tube formation assay. Most notably, the overexpression of miRs-139-5p and -454-3p rescued impaired tube formation in HUVECs with ENG knockdown. To our knowledge, we are the first to demonstrate miRNA alterations after the knockdown of ENG in HUVECs. Our results indicate a potential role of miRs-139-5p and -454-3p in ENG-deficiency-induced angiogenic dysfunction in ECs. Further study to examine the involvement of miRs-139-5p and -454-3p in HHT pathogenesis is warranted.

Antibody functionalized intravascular devices combined with genetically engineered endothelial colony-forming cells for targeted drug delivery: A proof-of-concept study.

This study was designed to test the ability of ex vivo antibody-coated intravascular devices to capture genetically engineered pig endothelial colony-forming cells (ECFCs) as proof of concept for their potential for in vivo targeted drug delivery. Human α-calcitonin gene-related peptide (α-CGRP) was chosen as the therapeutic molecule as it is unsuitable for systemic administration due to its potent peripheral arterial vasodilatory effect and short half-life in blood, requiring local delivery to yield therapeutic benefit in a particular vascular bed. H-2Kk, a murine leukocyte surface antigen, served as the selection marker for genetically modified ECFCs. H-2Kk antibody was immobilized on electropolished cobalt-chromium (CC) discs, CC stents and ePTFE grafts through dopamine self-polymerization. The functionalized surface was integral and smooth, lacked or had significantly reduced chemical signals specific for substrates. Pig bone marrow-derived ECFCs transfected with a plasmid constructed for H-2Kk and α-CGRP expression produced H-2Kk on cell surface and biologically active α-CGRP in culture medium. H-2Kk antibody-coated substrates bound H-2Kk ECFCs but not control ECFCs in vitro. Bare or only dopamine-coated substrates did not bind H-2Kk ECFCs. These data suggest that implantation of antibody functionalized devices combined with injection of genetically modified ECFCs could be potentially applied for targeted drug delivery.

Exploring Endothelial Colony-Forming Cells to Better Understand the Pathophysiology of Disease: An Updated Review.

Endothelial cell (EC) dysfunction has been implicated in a variety of pathological conditions. The collection of ECs from patients is typically conducted postmortem or through invasive procedures, such as surgery and interventional procedures, hampering efforts to clarify the role of ECs in disease onset and progression. In contrast, endothelial colony-forming cells (ECFCs), also termed late endothelial progenitor cells, late outgrowth endothelial cells, blood outgrowth endothelial cells, or endothelial outgrowth cells, are obtained in a minimally invasive manner, namely, by the culture of human peripheral blood mononuclear cells in endothelial growth medium. ECFCs resemble mature ECs phenotypically, genetically, and functionally, making them excellent surrogates for ECs. Numerous studies have been performed that examined ECFC function in conditions such as coronary artery disease, diabetes mellitus, hereditary hemorrhagic telangiectasia, congenital bicuspid aortic valve disease, pulmonary arterial hypertension, venous thromboembolic disease, and von Willebrand disease. Here, we provide an updated review of studies using ECFCs that were performed to better understand the pathophysiology of disease. We also discuss the potential of ECFCs as disease biomarkers and the standardized methods to culture, quantify, and evaluate ECFCs and suggest the future direction of research in this field.

MicroRNA-132-3p, Downregulated in Myeloid Angiogenic Cells from Hereditary Hemorrhagic Telangiectasia Patients, Is Enriched in the TGFß and PI3K/AKT Signalling Pathways.

BACKGROUND: Hereditary hemorrhagic telangiectasia (HHT) is a rare, autosomal dominant genetic disorder characterized by life-threatening vascular dysplasia. Myeloid angiogenic cells (MACs), alternatively called early endothelial progenitor cells or circulating angiogenic cells, do not directly incorporate into developing blood vessels, but augment angiogenesis in a paracrine manner. MAC dysfunction has been reported in HHT. MicroRNAs (miRNAs) regulate cellular function by modulating gene expression post-transcriptionally. To date, the role of miRNAs in HHT MAC dysfunction has not been documented. OBJECTIVE: The goal of this study was to comparatively profile miRNAs in HHT patient and control MACs to identify dysregulated miRNAs that may be responsible for the observed MAC dysfunction in HHT. METHODOLOGY/RESULTS: Twenty-three dysregulated miRNAs (twenty-one upregulated and two downregulated) in HHT MACs were identified with a TaqMan miRNA microarray. Pathway enrichment analysis showed that the dysregulated miRNAs were significantly enriched in pathways involved in HHT pathogenesis, such as the transforming growth factor ß (TGFß), phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), and Hippo signalling pathways. Furthermore, miR-132-3p was determined to be significantly reduced in HHT MACs compared with controls by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Bioinformatic analysis revealed that miR-132-3p is significantly enriched in the TGFß and PI3K/AKT signalling pathways, targeting SMAD4, an effector of the TGFß signalling pathway and RASA1, a negative regulator of the PI3K/AKT signalling pathway, respectively. CONCLUSION: MiRNA dysregulation, specifically reduced expression of miR-132-3p, in HHT MACs was identified. The dysregulated miRNAs are significantly enriched in the TGFß, PI3K/AKT, and Hippo signalling pathways. These data suggest that alteration in miRNA expression may impair these pathways and contribute to MAC dysfunction in HHT.

Endoglin deficiency impairs VEGFR2 but not FGFR1 or TIE2 activation and alters VEGF-mediated cellular responses in human primary endothelial cells.

Hereditary hemorrhagic telangiectasia (HHT) is a genetic disease characterized by vascular dysplasia. Mutations of the endoglin (ENG) gene that encodes a co-receptor of the transforming growth factor ß1 signaling pathway cause type I HHT. ENG is primarily expressed in endothelial cells (ECs), but its interaction with other key angiogenic pathways to control angiogenesis has not been well addressed. The aim of this study is to investigate ENG interplay with VEGFR2, FGFR1 and TIE2 in primary human ECs. ENG was knocked-down with siRNA in human umbilical vein ECs (HUVECs) and human lung microvascular ECs (HMVEC-L). Gene expression was measured by RT-qPCR and Western blotting. Cell signaling pathway activation was analyzed by detecting phosphor-ERK and phosphor-AKT levels. Cell migration and apoptosis were assessed using the Boyden chamber assay and the CCK-8 Kit, respectively. Loss of ENG in HUVECs led to significantly reduced expression of VEGFR2 but not TIE2 or FGFR1, which was also confirmed in HMVEC-L. HUVECs lacking ENG had significantly lower levels of active Rac1 and a substantial reduction of the transcription factor Sp1, an activator of VEGFR2 transcription, in nuclei. Furthermore, VEGF- but not bFGF- or angiopoietin-1-induced phosphor-ERK and phosphor-AKT were suppressed in ENG deficient HUVECs. Functional analysis revealed that ENG knockdown inhibited cell migratory but enhanced anti-apoptotic activity induced by VEGF. In contrast, bFGF, angiopoietin-1 and -2 induced HUVEC migration and anti-apoptotic activities were not affected by ENG knockdown. In conclusion, ENG deficiency alters the VEGF/VEGFR2 pathway, which may play a role in HHT pathogenesis.

Non-Coding RNAs and Hereditary Hemorrhagic Telangiectasia.

Non-coding RNAs (ncRNAs) are functional ribonucleic acid (RNA) species that include microRNAs (miRs), a class of short non-coding RNAs (∼21-25 nucleotides), and long non-coding RNAs (lncRNAs) consisting of more than 200 nucleotides. They regulate gene expression post-transcriptionally and are involved in a wide range of pathophysiological processes. Hereditary hemorrhagic telangiectasia (HHT) is a rare disorder inherited in an autosomal dominant fashion characterized by vascular dysplasia. Patients can develop life-threatening vascular malformations and experience severe hemorrhaging. Effective pharmacological therapies are limited. The study of ncRNAs in HHT is an emerging field with great promise. This review will explore the current literature on the involvement of ncRNAs in HHT as diagnostic and pathogenic factors.

MicroRNA profiling of human myeloid angiogenic cells derived from peripheral blood mononuclear cells.

Human myeloid angiogenic cells (MACs), also termed early endothelial progenitor cells, play an important role in neovascularization and vascular repair. MicroRNAs (miRNAs) are a class of naturally occurring, noncoding, short (∼22 nucleotides), single-stranded RNAs that regulate gene expression post-transcriptionally. MiRNAs have been shown to regulate MAC function. A miRNA signature of MACs was described approximately a decade ago, and many new miRNAs have been discovered in recent years. In this study, we aimed to provide an up-to-date miRNA signature for human MACs. MACs were obtained by culture of human peripheral blood mononuclear cells in endothelial medium for 7 days. Using qPCR array analysis we identified 72 highly expressed miRNAs (CT value < 30) in human MACs. RT-qPCR quantification of select miRNAs revealed a strong correlation between the CT values detected by the array analysis and RT-qPCR, suggesting the miRNA signature generated by the qPCR array assay is accurate and reliable. Experimentally validated target genes of the 10 most highly expressed miRNAs were retrieved. Only a few of the targets and their respective miRNAs have been studied for their role in MAC biology. Our study therefore provides a valuable repository of miRNAs for future exploration of miRNA function in MACs.

MicroRNA signature of human blood mononuclear cells.

MicroRNAs (miRNAs) regulate a wide range of cellular processes and functions. Blood mononuclear cells (BMNCs) participate in the immune response, inflammatory reaction and angiogenesis. In 2010, a total of 157 miRNAs were quantified by RT-qPCR and a miRNA signature was determined for human peripheral BMNCs. With the advent of technologies such as RNA sequencing, many new miRNAs have been identified. This study was designed to provide an up-to-date miRNA signature for human BMNCs. Peripheral BMNCs were isolated by Ficoll density gradient centrifugation. Using the qPCR array assay, we identified 108 highly expressed miRNAs (Ct value < 30) in human BMNCs. Further validation of the array results by quantifying select miRNAs with RT-qPCR revealed a strong correlation between Ct values derived from array analysis and RT-qPCR, suggesting the array results presented in this study are accurate and reliable. Of note, the function of the majority of the highly expressed miRNAs we have identified has not yet been studied. Our findings may help direct further studies of the regulatory roles of miRNAs in BMNC function.

Decreased levels of miR-28-5p and miR-361-3p and increased levels of insulin-like growth factor 1 mRNA in mononuclear cells from patients with hereditary hemorrhagic telangiectasia 1.

Hereditary hemorrhagic telangiectasia (HHT) is a rare vascular disorder inherited in an autosomal dominant manner. Patients with HHT can develop vascular dysplasias called telangiectasias and arteriovenous malformations (AVMs). Our objective was to profile and characterize micro-RNAs (miRNAs), short noncoding RNAs that regulate gene expression posttranscriptionally, in HHT patient-derived peripheral blood mononuclear cells (PBMCs). PBMCs, comprised mostly of lymphocytes and monocytes, have been reported to be dysfunctional in HHT. A total of 40 clinically confirmed HHT patients and 22 controls were enrolled in this study. PBMCs were isolated from 16 mL of peripheral blood and purified for total RNA. MiRNA expression profiling was conducted with a human miRNA array analysis. Select dysregulated miRNAs and miRNA targets were validated with reverse transcription-quantitative polymerase chain reaction. Of the 377 miRNAs screened, 41 dysregulated miRNAs were identified. Both miR-28-5p and miR-361-3p, known to target insulin-like growth factor 1 (IGF1), a potent angiogenic growth factor, were found to be significantly downregulated in HHT patients. Consequently, IGF1 mRNA levels were found to be significantly elevated. Our research successfully identified miRNA dysregulation and elevated IGF1 mRNA levels in PBMCs from HHT patients. This novel discovery represents a potential pathogenic mechanism that could be targeted to alleviate clinical manifestations of HHT.

Myeloid angiogenic cells exhibit impaired migration, reduced expression of endothelial markers, and increased apoptosis in idiopathic pulmonary arterial hypertension 1.

Idiopathic pulmonary arterial hypertension (IPAH) is a rare and devastating condition. There is no known cure for IPAH, and current treatment options are not always effective. Autologous myeloid angiogenic cells (MACs) have been explored as a novel therapy for IPAH, but preliminary data from clinical trials show limited beneficial effects. A complete understanding of IPAH MAC function remains elusive. This study was designed to comprehensively compare cell function between IPAH MACs and healthy control MACs. MACs were procured through the culture of peripheral blood mononuclear cells in endothelial selective medium for 7 days. Compared with healthy MACs, IPAH MACs exhibited (1) significantly lower levels of endothelial markers as shown by fluorescence microscopy; (2) a markedly higher rate of apoptosis under both normal culture condition and serum starvation as shown by the TUNEL assay; (3) significantly decreased migration towards vascular endothelial growth factor as shown by a modified Boyden chamber migration assay; and (4) similar vascular endothelial growth factor and endothelial nitric oxide synthase mRNA levels as shown by reverse transcription quantitative PCR. In conclusion, various aspects of IPAH MAC function are impaired. To achieve greater therapeutic benefits, pharmacologic and (or) genetic manipulations to improve IPAH MAC function, particularly to promote cell survival and migration, are warranted.

The Neurorepellent Slit2 Inhibits Postadhesion Stabilization of Monocytes Tethered to Vascular Endothelial Cells.

The secreted neurorepellent Slit2, acting through its transmembrane receptor, Roundabout (Robo)-1, inhibits chemotaxis of varied cell types, including leukocytes, endothelial cells, and vascular smooth muscle cells, toward diverse attractants. The role of Slit2 in regulating the steps involved in recruitment of monocytes in vascular inflammation is not well understood. In this study, we showed that Slit2 inhibited adhesion of monocytic cells to activated human endothelial cells, as well as to immobilized ICAM-1 and VCAM-1. Microfluidic live cell imaging showed that Slit2 inhibited the ability of monocytes tethered to endothelial cells to stabilize their actin-associated anchors and to resist detachment in response to increasing shear forces. Transfection of constitutively active plasmids revealed that Slit2 inhibited postadhesion stabilization of monocytes on endothelial cells by preventing activation of Rac1. We further found that Slit2 inhibited chemotaxis of monocytes toward CXCL12 and CCL2. To determine whether Slit2 and Robo-1 modulate pathologic monocyte recruitment associated with vascular inflammation and cardiovascular disease, we tested PBMC from patients with coronary artery disease. PBMC from these patients had reduced surface levels of Robo-1 compared with healthy age- and sex-matched subjects, and Slit2 failed to inhibit chemotaxis of PBMC of affected patients, but not healthy control subjects, toward CCL2. Furthermore, administration of Slit2 to atherosclerosis-prone LDL receptor-deficient mice inhibited monocyte recruitment to nascent atherosclerotic lesions. These results demonstrate that Slit2 inhibits chemotaxis of monocytes, as well as their ability to stabilize adhesions and resist detachment forces. Slit2 may represent a powerful new tool to inhibit pathologic monocyte recruitment in vascular inflammation and atherosclerosis.

Angiographic and clinical outcomes after implantation of drug eluting stents in bifurcation lesions with crush or kissing stent technique.

BACKGROUND: Long-term outcome after bifurcation stenting with drug-eluting stents (DES) for obstructive coronary artery disease is poorly understood. In this study, we report 6-9-month angiographic follow-up and long-term clinical outcomes after implantation of drug-eluting stents by crush and kissing stent technique for coronary bifurcation lesions. METHODS: Consecutive patients undergoing bifurcation stenting with DES by crush or kissing stent technique were enrolled in a prospective registry. Angiographic follow-up was obtained at 6-9 months and clinical follow-up completed for a median of 38 months. RESULTS: A total of 86 patients participated in the study. Bifurcation stenting by crush technique was performed in 73 (85%) and by kissing stent in 13 (15%) patients. Stenting of left main bifurcation was applied in 24 (28%) patients. Angiographic follow-up was completed in 75 (87%) patients and showed restenosis in the main for 8 (11%) and side branch for 20 (27%) patients. Clinical follow-up was available for a median duration of 38 months. During follow-up, 2 (2%) patients died, 4 (5%) experienced myocardial infarction (MI), and 11 (13%) underwent target vessel revascularization (TVR) with an overall major adverse cardiac event (MACE) rate of 16%. In left main cohort, angiographic restenosis occurred in 9 (37%) patients, and 3 (12%) patients required TVR. There were no deaths or stent thrombosis. A comparison of crush and kissing stent technique showed significantly higher angiographic restenosis with crush (26% vs 13% in kissing stent patients, P = 0.046) and 95% of restenosis in crush group involved ostium of the side branch. There was no difference in clinical outcomes between the crush and kissing stent groups. Final kissing balloon dilatation (FKB) was successful in 65 (89%) patients in the crush group and associated with a significant reduction in MACE (8% in FKB successful vs 37% in FKB unsuccessful, P = 0.04) during follow-up. CONCLUSION: Bifurcation stenting with crush or kissing stent technique is safe and associated with a low rate of TLR and MACE on long-term follow-up. Crush stenting is associated with a significantly higher rate of side branch restenosis compared to kissing stent technique. FKB is associated with significant reduction in MACE during follow-up.

Characterization of clopidogrel hypersensitivity reactions and management with oral steroids without clopidogrel discontinuation.

OBJECTIVES: The purpose of this study was to characterize clopidogrel hypersensitivity and describe its successful management with oral steroids without clopidogrel discontinuation. BACKGROUND: Hypersensitivity reactions to clopidogrel are poorly understood and present difficulty in management. METHODS: Patients diagnosed with clopidogrel hypersensitivity after percutaneous coronary intervention underwent evaluation and received oral prednisone without clopidogrel discontinuation. Cutaneous testing was performed after completion of clopidogrel therapy for diagnosis and assessment of cross-reactivity. RESULTS: Sixty-two patients representing 1.6% of the percutaneous coronary intervention population developed clopidogrel hypersensitivity during the study period. The mean age was 62 ± 11 years, 71% of patients were male, and 35% reported prior adverse drug reaction. Clopidogrel hypersensitivity manifested as generalized exanthema in 79%, localized skin reaction in 16%, and angioedema or urticaria in 5% of patients. Biopsy of affected areas demonstrated a lymphocyte-mediated delayed hypersensitivity reaction. Complete resolution of hypersensitivity reaction was observed in 61 patients (98%) with a short course of oral prednisone. Cutaneous testing confirmed delayed hypersensitivity reaction to clopidogrel in 34 (81%) and immediate hypersensitivity in 3 of 42 patients (7%) tested. Allergenic cross-reactivity was observed for ticlopidine in 10 (24%), prasugrel in 7 (17%), and both ticlopidine and prasugrel in 3 patients (7%). Histological examination showed lymphocyte-mediated hypersensitivity in abnormal patch test areas. CONCLUSIONS: Clopidogrel hypersensitivity is manifested as generalized exanthema and is caused by a lymphocyte-mediated delayed hypersensitivity in most patients. This can be managed with oral steroids without clopidogrel discontinuation. Allergenic cross-reactivity with ticlopidine, prasugrel, or both is present in a significant number of patients with clopidogrel hypersensitivity.

Characterization of operator learning curve for transradial coronary interventions.

BACKGROUND: Transradial percutaneous coronary intervention (TR-PCI) improves clinical outcomes compared to the transfemoral (TF) approach. However, inadequate training and experience has limited widespread adoption by interventional cardiologists. METHODS AND RESULTS: Clinical and procedural characteristics for TR-PCI were prospectively collected from 1999 to 2008. To identify minimum case volume for optimum clinical benefit, single-vessel TR-PCI cases were chronologically ranked and stratified into 1 to 50, 51 to 100, 101 to 150 and 151 to 300 case volume groups for operators starting the TR approach at the study institution. Cases by operators with a >300 TR-PCI case volume comprised the control group. TR-PCI failure rates, contrast use, guide usage, and fluoroscopy time were compared among groups. A total of 1672 patients underwent TR-PCI by 28 operators. TR-PCI failure occurred in 4% and was higher in the 1 to 50 case volume group compared to the 51 to 100 (P=0.007) and control (P=0.01) groups. Contrast use was greater in the 1 to 50 group (180±79 mL) compared to the 151 to 300 (157±75 mL, P=0.02) and control (168±79 mL, P=0.05) groups. Fluoroscopy time was higher in the 1 to 50 group (15±10 minutes) compared to the 101 to 150 (13±10 minutes, P=0.04) and control (12±9 minutes, P=0.02) groups. Reasons for TR-PCI failure included spasm (38%), subclavian tortuousity (16%), poor guide support (16%), failed access (10%), and radial loop (7%). Case volume was significantly correlated with TR-PCI failure (ß=-0.0076, P=0.0028), and odds of failure was reduced by 32% for each 50 increments in case volume. CONCLUSIONS: TR-PCI success depends on operator experience, and a case volume of ≥50 cases is required to achieve outcomes comparable to experienced operators. These findings have implications both for PCI operators looking to expand their skills and for defining standards for training.

Nitric oxide synthase gene transfer restores activity of circulating angiogenic cells from patients with coronary artery disease.

Circulating angiogenic cells (CACs), represent a potential new therapeutic tool for the treatment of cardiovascular diseases, but their regenerative function is impaired in patients with coronary artery disease (CAD) and cardiac risk factors. The objective of this study is to assess the effect of lentiviral overexpression of endothelial nitric oxide synthase (eNOS) on the activity of CACs from patients with CAD and cardiac risk factors. In vitro and in vivo assays were employed to evaluate the regenerative capacity of the cells compared to CACs derived from healthy volunteers. Lentiviral eNOS transduction of cells from CAD patients significantly improved chemotactic migration compared with sham transduction, and increased the ability of CACs to induce angiogenic tube formation when cocultured with human umbilical vein endothelial cells (HUVECs) on Matrigel. In addition, eNOS transduction restored the ability of patient-derived CACs to enhance neovascularization and improve ischemic hind limb perfusion, approaching the efficacy of cells from healthy donors. These data indicate that CAC dysfunction seen in high-risk patients can be partially reversed by eNOS overexpression, suggesting that ex vivo gene delivery may improve the efficacy of autologous cell therapy for cardiovascular disease.

Comparison of abciximab and eptifibatide on angiographic and clinical outcomes in rescue percutaneous coronary intervention for failed fibrinolytic therapy.

BACKGROUND: Adjunctive administration of the glycoprotein IIb/IIIa platelet receptor antagonist (GPA), abciximab, improves outcomes in patients undergoing rescue percutaneous coronary intervention (PCI). However, it is unknown if other GPAs provide a similar benefit in this setting. OBJECTIVE: We sought to compare angiographic and clinical outcomes of patients receiving abciximab or eptifibatide as an adjunct to rescue PCI. METHODS: In this prospective, nonrandomized study, consecutive patients who underwent rescue PCI and received adjunctive preprocedural GPA comprised the study population. Thrombolysis in myocardial infarction (TIMI) flow, corrected TIMI frame count (CTFC) and myocardial blush grade (MBG) were determined before and immediately after rescue PCI. Residual ST-segment elevation at 90-120 minutes and peak creatine kinase (CK) values for 48 hours after PCI were recorded. Major adverse cardiac events (MACE) including death, reinfarction and target vessel revascularization (TVR) were determined at discharge, 1 and 6 months. RESULTS: A total of 241 patients were included in the study. 162 patients received abciximab and 79 received eptifibatide. There were no differences in baseline clinical and angiographic characteristics between groups. Post-PCI TIMI flow was similar but post-PCI CTFC was significantly lower (17 +/- 10 vs. 22 +/- 18; p = 0.01) and post-PCI MBG significantly higher (2.8 +/- 0.5 vs. 2.6 +/- 0.6; p = 0.01) in the abciximab group. Patients in the abciximab group had less ST-segment elevation (1.0 +/- 0.9 vs. 1.5 +/- 1.0 mm; p = 0.003) and lower peak CK (2,484 +/- 2,176 vs. 2,650 +/- 2,798 U/L; p = 0.001) after PCI. On multivariate analyses, abciximab administration (OR = 0.50, CI = 0.26, 0.96; p = 0.03), pre-PCI TIMI 3 flow (OR = 0.22, CI = 0.05, 0.99; p = 0.04) and female gender (OR = 0.24, CI = 0.08, 0.66; p = 0.006) were positive and cardiogenic shock (OR = 2.76, CI = 1.16, 6.58; p = 0.02) was a negative predictor of normal epicardial perfusion post PCI. Abciximab administration (OR = 0.46, CI = 0.24, 0.87; p = 0.02) and pre-PCI CTFC < 25 (OR = 0.09, CI = 0.02, 0.31, 0.0001) were positive predictors and cardiogenic shock (OR = 3.96, CI = 1.55, 10.12; p = 0.004) was a negative predictor of normal myocardial perfusion post-PCI as determined by CTFC. Abciximab administration (OR = 0.31, CI = 0.15, 0.63; p = 0.001) and pre-PCI MBG 3 (OR = 0.07, CI = 0.02, 0.23; p < 0.0001) were positive predictors of normal myocardial perfusion post-PCI as determined by MBG. In-hospital, 1- and 6-month clinical events and MACE rates did not differ between groups. CONCLUSIONS: In the setting of rescue PCI, adjunctive administration of abciximab resulted in greater improvement in angiographic and electrical estimates of myocardial perfusion and smaller infarct size compared to eptifibatide. These findings suggest that all GPA may not provide equal benefit in rescue PCI.

Rationale and design of Enhanced Angiogenic Cell Therapy in Acute Myocardial Infarction (ENACT-AMI): the first randomized placebo-controlled trial of enhanced progenitor cell therapy for acute myocardial infarction.

BACKGROUND: Despite the widespread use of pharmacological and/or interventional reperfusion therapies, recovery of cardiac function in myocardial infarction (MI) patients is often modest or even absent. Unlike classical pharmacological treatments, the use of progenitor cells could potentially restore functional tissue in regions that otherwise would form only scar. However, a major limitation of autologous cell therapy is the deleterious influence of age and cardiac risk factors on progenitor cell activity. TRIAL DESIGN: The ENACT-AMI trial is a phase IIb, double-blind, randomized placebo-controlled trial, using transplantation of autologous early endothelial progenitor cells (EPCs) for patients who have suffered large MI. Circulating mononuclear cells (MNCs) are obtained by apheresis and subjected to differential culture for 3 days to select a population of highly regenerative, endothelial-like, culture modified MNCs (E-CMMs), often referred to as "early EPCs." A total of 99 patients will be randomized to placebo (Plasma-Lyte A), autologous E-CMMs, or E-CMMs transfected with human endothelial nitric oxide synthase delivered by coronary injection into the infarct-related artery. The primary efficacy end point is change from baseline to 6 months in global left ventricular ejection fraction by cardiac MRI; secondary endpoints include regional wall motion, wall thickening, infarct volume, time to clinical worsening, and quality of life. CONCLUSIONS: This will be the first clinical trial to include a strategy designed to enhance the function of autologous progenitor cells by overexpressing endothelial nitric oxide synthase, and the first to use combination gene and cell therapy for the treatment of cardiac disease.

Mechanism and predictors of failed transradial approach for percutaneous coronary interventions.

OBJECTIVES: The study aimed to determine the mechanism and predictors of procedural failure in patients undergoing percutaneous coronary intervention (PCI) from the transradial approach (TR). BACKGROUND: Transradial approach PCI reduces vascular complications compared with a transfemoral approach (TF). However, the mechanism and predictors of TR-PCI failure have not been well-characterized. METHODS: The study population consisted of patients undergoing TR-PCI by low-to-intermediate volume operators with traditional TF guide catheters. Baseline characteristics, procedure details, and clinical outcomes were prospectively collected. Univariate and multivariate analyses were performed to determine independent predictors of TR-PCI failure. RESULTS: A total of 2,100 patients underwent TR-PCI and represented 38% of PCI volume. Mean age was 64 +/- 12 years, and 17% were female. Vascular complications occurred in 22 (1%), and TR-PCI failure was observed in 98 (4.7%) patients. The mechanism of TR-PCI failure included inability to advance guide catheter to ascending aorta in 50 (51%), inadequate guide catheter support in 35 (36%), and unsuccessful radial artery puncture in 13 (13%) patients. The PCI was successful in 94 (96%) patients with TR-PCI failure by switching to TF. On multivariate analysis, age >75 years (odds ratio [OR]: 3.86; 95% confidence interval [CI]: 2.33 to 6.40, p = 0.0006), prior coronary artery bypass graft surgery (OR: 7.47; 95% CI: 3.45 to 16.19, p = 0.0002), and height (OR: 0.97; 95% CI: 0.95 to 0.99, p = 0.02) were independent predictors of TR-PCI failure. CONCLUSIONS: Transradial approach PCI can be performed by low-to-intermediate volume operators with standard equipment with a low failure rate. Age >75 years, prior coronary artery bypass graft surgery, and short stature are independent predictors of TR-PCI failure. Appropriate patient selection and careful risk assessment are needed to maximize benefits offered by TR-PCI.

VEGF gene therapy fails to improve perfusion of ischemic myocardium in patients with advanced coronary disease: results of the NORTHERN trial.

Despite the promise of proangiogenic gene therapy most clinical trials have failed to show benefit for the primary end point analysis. The NOGA angiogenesis Revascularization Therapy: assessment by RadioNuclide imaging (NORTHERN) trial was a double-blind, placebo-controlled study of intramyocardial vascular endothelial growth factor (VEGF165) gene therapy versus placebo, involving seven sites across Canada, designed to overcome major limitations of previous proangiogenic gene therapy trials. A total of 93 patients with refractory Canadian Cardiovascular Society (CCS) class 3 or 4 anginal symptoms were randomized to receive 2,000 microg of VEGF plasmid DNA or placebo (buffered saline) delivered via the endocardial route using an electroanatomical NOGA guidance catheter. There was no difference between the VEGF-treated and the placebo groups in the primary end point of change in myocardial perfusion from baseline to 3 or 6 months, assessed by single photon emission tomography (SPECT) imaging, although a significant reduction in the ischemic area was seen in both groups. Also, similar improvements in exercise treadmill time and anginal symptoms were seen in the VEGF and the placebo groups at 3 and 6 months, although again there were no differences between these groups. Despite the intramyocardial administration of a high "dose" of plasmid DNA using a percutaneous guidance catheter system, there was no benefit of VEGF gene therapy at 3 or 6 months for any of the end points studied.

Endothelial progenitor cell therapy for the treatment of coronary disease, acute MI, and pulmonary arterial hypertension: current perspectives.

Since their identification in 1997, bone marrow derived endothelial progenitor cells (EPCs) have been studied for their role in the endogenous maintenance and repair of endothelium and their potential regenerative capacity beyond the endothelium. In particular, EPCs have been tested in cell therapy approaches with the aim of developing novel therapies for conditions currently lacking effective treatment options. In this review, we discuss the scientific background and clinical experience using EPC delivery or mobilization for the treatment of post-angioplasty restenosis, acute myocardial infarction and pulmonary arterial hypertension. Although these approaches are safe, efficacy has yet to be proven in large randomized clinical trials. Unfortunately, the biology of EPCs is still poorly understood. The success of future clinical trials depends on a better understanding of EPC biology and intelligent design.