Early Re-endothelialization Research Articles

AMD3100 and SDF‑1 regulate cellular functions of endothelial progenitor cells and accelerate endothelial regeneration in a rat carotid artery injury model

The present study was conducted to assess the effects of AMD3100 and stromal cell-derived factor 1 (SDF-1) on cellular functions and endothelial regeneration of endothelial progenitor cells (EPCs). The cell proliferation and adhesion capacity of EPCs were evaluated in vitro following treatment with AMD3100 and SDF-1 using a Cell Counting Kit-8 assay. Furthermore, the expression levels of C-X-C motif chemokine receptor 4 (CXCR4) and C-X-C motif chemokine receptor 7 (CXCR7) were detected before and after treatment with AMD3100 and SDF-1 to elucidate their possible role in regulating the cellular function of EPCs. A rat carotid artery injury model was established to assess the influences of AMD3100 and SDF-1 on endothelial regeneration. AMD3100 reduced the proliferation and adhesion capacity of EPCs to fibronectin (FN), whereas it increased the adhesion capacity of EPCs to human umbilical vein endothelial cells (HUVECs). However, SDF-1 stimulated the proliferation and cell adhesion capacity of EPCs to HUVECs and FN. Additionally, the expression levels of CXCR7 but not CXCR4 were upregulated following AMD3100 treatment, whereas the expression levels of both CXCR4 and CXCR7 were upregulated after SDF-1 treatment. In vivo results demonstrated that AMD3100 increased the number of EPCs in the peripheral blood and facilitated endothelial repair at 7 days after treatment. However, local administration of SDF-1 alone did not enhance reendothelialization 7 and 14 days after treatment. Importantly, the combination of AMD3100 with SDF-1 exhibited superior therapeutic effects compared with AMD3100 treatment alone, accelerated reendothelialization 7 days after treatment, and attenuated neointimal hyperplasia at day 7 and 14 by recruiting more EPCs to the injury site. In conclusion, AMD3100 could positively regulate the adhesion capacity of EPCs to HUVECs via elevation of the expression levels of CXCR7 but not CXCR4, whereas SDF-1 could stimulate the proliferation and adhesion capacity of EPCs to FN and HUVECs by elevating the expression levels of CXCR4 and CXCR7. AMD3100 combined with SDF-1 outperformed AMD3100 alone, promoted early reendothelialization and inhibited neointimal hyperplasia, indicating that early reendothelialization attenuated neointimal hypoplasia following endothelial injury.

Development of in vitro endothelialized drug-eluting stent using human peripheral blood-derived endothelial progenitor cells.

We propose in vitro endothelialization of drug-eluting stents (DES) to overcome late stent thrombosis by directly introducing late-outgrowth human endothelial progenitor cells (EPCs) at the target site utilizing abluminal DES. Isolated EPCs were confirmed as late-outgrowth EPCs by flow cytometric analysis. Abluminally paclitaxel-loaded stents were seeded with different cell concentrations and durations to determine optimal seeding conditions, in both uncrimped and crimped configurations. The seeding yield was determined by evaluating the percent coverage of the stent struts' area. The EPC-seeded DES were exposed to arterial shear stress to evaluate the effect of high shear stress on EPCs. To investigate how much paclitaxel elutes during the seeding procedure, a pharmacokinetic analysis was performed. Finally, to validate the proof of concept, EPC-seeded DES were placed on a fibrin matrix with and without smooth muscle cells (SMCs) and cultured for 3 days under perfusion. The seeding procedure resulted in 47% and 26% coverage of the stent surface in uncrimped and crimped conditions, respectively. After the optimal seeding, almost 99% of drug was still available. When EPC-seeded DES were placed on a fibrin matrix and cultured for 3 days, the EPCs confluently covered the stent surface and spread to the surrounding fibrin gel. When EPC-seeded DES were placed on SMC-containing fibrin layers, cells in contact with the struts died. EPCs can be successfully seeded onto DES without losing drug-eluting capability, and EPCs exhibit sufficient proliferative ability. EPC-seeded DES may combine early re-endothelialization ability with the antirestenotic effectiveness of DES.

Endovascular procedures cause transient endothelial injury but do not disrupt mature neointima in Drug Eluting Stents

Extensive application of coronary intravascular procedures has led to the increased need of understanding the injury inflicted to the coronary arterial wall. We aimed to investigate acute and prolonged coronary endothelial injury as a result of guidewire use, repeated intravascular imaging and stenting. These interventions were performed in swine (N = 37) and injury was assessed per coronary segment (n = 81) using an Evans Blue dye-exclusion-test. Scanning electron microscopy and light microscopy were then used to visualize the extent and nature of acute (<4 hours) and prolonged (5 days) endothelial injury. Guidewire and imaging injury was mainly associated with denudation and returned to control levels at 5 days. IVUS and OCT combined (Evans Blue staining 28 ± 16%) did not lead to more acute injury than IVUS alone (33 ± 15%). Stent placement caused most injury (85 ± 4%) and despite early stent re-endothelialization at 5 days, the endothelium proved highly permeable (97 ± 4% at 5 days; p < 0.001 vs acute). Imaging of in-stent neointima at 28 days after stent placement did not lead to neointimal rupture. Guidewire, IVUS and OCT induce acute endothelial cell damage, which does not increase during repeated imaging, and heals within 5 days. Interestingly, endothelial permeability increases 5 days post stenting despite near complete re-endothelialization.

Cilostazol-loaded electrospun three-dimensional systems for potential cardiovascular application: Effect of fibers hydrophilization on drug release, and cytocompatibility

Currently marketed drug-eluting stents are non-selective in their anti-restenotic action. New active substance introduction to polymeric stents and vascular grafts can promote early re-endothelialization, crucial in preventing implant restenosis. Additionally, managing material hydrophobicity by blending synthetic polymers limits adverse effects on bulk properties and controls active substance release. However, the influence of hydrophilic synthetic polymer on human cells in the cardiovascular system remains to be determined. In this report, effects of both poly(ε-caprolactone) (PCL) fibers hydrophilization with Pluronic P123 (P123) and cilostazol (CIL) loading were studied.Physicochemical and mechanical properties of electrospun tubular structures produced from PCL and PCL/P123 fibers with and without CIL were investigated and compared. Release profiles studies and in vitro cell proliferation assays of electrospun materials were conducted. It was found that P123 located near the surface of electrospun fibers increased the rate of CIL release. PCL formulation sustained human umbilical vein endothelial cells (HUVEC) growth for 48 h. Despite improved hydrophilicity, PCL/P123 formulations were found to reduce HUVEC viability. Both PCL and PCL/P123 materials reduced primary aortic smooth muscle cells (PASM) viability after 48 h. In PCL formulations containing CIL, drug release caused a decrease in PASM viability. P123 blending with PCL was found to be as a useful pre-fabrication technique for modulating surface hydrophobicity of electrospun materials and the release profile of incorporated active substance. The cytotoxicity of P123 was evaluated to improve the design of drug-loaded vascular grafts for cardiovascular applications.

TCT-269 Optical Coherence Tomography evaluation of Very Early eNdothelialization after Dual Therapy Drug-eluting Stent with EPC CaptURE (OCT-VENTURE)

The Combo drug-eluting stent (OrbusNeich Medical, Inc, Fort Lauderdale, FL) has an abluminal sirolimus-eluting bioabsorbable polymer and a luminal CD34+ antibody designed to capture endothelial progenitor cells to promote early re-endothelialization. This is the first prospective study to evaluate

P1653The role of thrombus and its composition in early re-endothelialization in injured swine coronary arteries

P1653The role of thrombus and its composition in early re-endothelialization in injured swine coronary arteries

Randomized evaluation of short-term dual antiplatelet therapy in patients with acute coronary syndrome treated with the COMBO dual therapy stent: rationale and design of the REDUCE trial

The optimal duration of dual antiplatelet therapy (DAPT) in acute coronary syndrome (ACS) patients treated with drug eluting stents (DES) is still under debate. Recent meta-analyses on ≤6months versus 12months DAPT suggest that bleeding rates can be reduced, without a higher rate of thrombotic complications. In particular, the COMBO dual therapy stent, being associated with early re-endothelialization, may allow for a reduction of the duration of DAPT without increasing the thrombotic risk, while reducing the risk of bleeding complications. The aim of the REDUCE trial is to demonstrate the non-inferiority of a combined efficacy and safety endpoint of a short-term 3months DAPT strategy as compared to standard 12-month DAPT strategy in ACS patients treated with the COMBO stent. A prospective, multicenter, randomized study designed to enroll 1500 patients with ACS treated with the COMBO stent. Patients will be randomized before discharge in a 1:1 fashion to either 3 or 12months of DAPT. A clinical follow-up is scheduled at 3, 6, 12, and 24months. The primary endpoint is the time to event as defined by the occurrence of one of the following: all cause mortality, myocardial infarction, stent thrombosis, stroke, target vessel revascularization or bleeding (Bleeding Academic Research Council type II, III and V) within 12months. The study has recruited patients since July 2014, and the results are expected in 2017. A reduction of the DAPT duration in ACS patients after PCI without affecting the thrombotic risk is an attractive option with regard to the associated bleeding risk. The REDUCE trial will be the first to investigate the efficacy and safety of a 3-month DAPT strategy compared to a 12-month DAPT strategy in an ACS only population treated with the COMBO stent.

Chitosan/heparin sirolimus drug-eluting stent system improves anticoagulation and promotes early re-endothelialization in porcine coronary artery

Objective To construct a sirolimus loaded drug-eluting stent(DES)using chitosan/heparin coating membrane and to explore its effect on anticoagulation and early re-endothelialization in porcine coronary artery.Methods Chitosan/heparin layer-by-layer self-assembly coating was applied to sirolimus DES,and the asymmetric applicator was used to allow for the stent blood surface to be chitosan/heparin coated and the stent vascular surface to be polylactic acid(PLA)-sirolimus.The experiment was divided into bare metal stent(BMS)group,chitosan/heparin stent group,sirolimus DES group and chitosan/heparin sirolimus DES group.The anticoagulation effect of chitosan/heparin sirolimus DES was tested by arteriovenous shunt model and high load thrombosis model.The effect of chitosan/heparin sirolimus DES on early reendothelialization was tested by 1-week long balloon injury to porcine coronary artery.Results No thrombus was found on the surfaces of chitosan/heparin sirolimus DES and chitosan/heparin stent in the arteriovenous shunt model,while the surfaces of BMS and sirolimus DES were covered with thrombus.No stent thrombosis was found in the high load thrombosis model test of chitosan/heparin sirolimus DES and chitosan/heparin stent within 6hours,and stent thrombosis was found in BMS at(59.0± 8.5)min and in sirolimus DES at(67.0±7.8)min(P0.01).The early re-endothelialization test showed endothelial coverage rates of chitosan/heparin sirolimus DES and chitosan/heparin stent after 7dimplantation were(82.7±16.4)% and(80.7±14.1)%, significantly higher than those of BMS and sirolimus DES([64.3±11]%and[59.8±8.4]%,respectively;P0.01).Conclusion Chitosan/heparin-coated-sirolimus DES has satisfactory anticoagulation property and it can accelerate early re-endothelialization.

Basic Fibroblast Growth Factor as a Potential Stent Coating Material Inducing Endothelial Cell Proliferation

The use of drug-eluting stents has reduced the incidence of in-stent restenosis following percutaneous coronary intervention; however, almost all drugs eluting from polymers on stents induce antiproliferative effects on vascular endothelial and vascular smooth muscle cells. Due to injury of the endothelium and delayed reendothelialization, the risk of thrombosis increases over time. Enhancing rapid reendothelialization after stent placement is important for solving these problems. Basic fibroblast growth factor (bFGF) is one of the most important growth factors involved in vascular lesion formation. In this study, we evaluated the potential of bFGF as a stent coating promoting endothelial cell proliferation. Human umbilical vein endothelial cells (HUVECs) and human aortic smooth muscle cells (HASMCs) were cultured with various doses of bFGF in vitro, and the effects of bFGF on the degree of cell proliferation and migration were monitored. We also investigated the effects of bFGF on the protein expression of endothelial nitric oxide synthase (eNOS) in HUVECs using Western blotting. Cell proliferation and migration were promoted in HUVECs by bFGF in a dose- and time-dependent manner. On the other hand, bFGF stimulation had little effect on the HASMCs. Basic FGF increased the eNOS protein levels in the HUVECs, with a maximum at 10 ng/mL followed by a decline at 100 ng/mL. Basic FGF is a possible candidate stent coating and/or eluting drug material stimulating endothelial cell proliferation and early reendothelialization without excessive vascular smooth muscle cell proliferation.

Muscle-Derived Stem Cells Promote Angiogenesis and Attenuate Intimal Hyperplasia in Different Murine Vascular Disease Models

Muscle-derived stem cells (MDSCs) are known to promote angiogenesis, but have never been studied in vascular diseases. We differentiated MDSCs into endothelial lineage cells in vitro by stimulation with shear stress and vascular endothelial growth factor. Such differentiated MDSCs (diff-MDSC) showed strong angiogenic potential in vitro. When tested in ischemic hindlimbs of mice, diff-MDSCs increased perfusion and decreased necrosis of the ischemic limbs, by promoting new vessel formation and by upregulating genes involved in endothelial expression. Such effects were not observed with native MDSCs (without endothelial stimulation in vitro). Diff-MDSCs were also injected into carotid arteries of rats after balloon denudation of the intima layer to induce intimal hyperplasia. The cell-treated group had significantly reduced intima-to-media thickness ratio compared to control, thus attenuating intimal hyperplasia by early re-endothelialization of the intima layer. Our findings suggest that MDSCs are a potential source of stem cell therapy for treatment of various vascular diseases, by inducing angiogenesis to improve perfusion in sites of ischemia, and by preventing intimal hyperplasia in sites of vessel injury.

Rationale and design of the randomized, multicenter EREMUS trial, a study that investigates how to achieve low restenosis and early reendothelialization after percutaneous coronary interventions

Drug eluting stent (DES) use has improved clinical outcome after percutaneous coronary interventions. However, DES-treated patients may have a higher risk of stent thrombosis, mainly due to uncorrect stent deployment or lack of reendothelialization. Thus, the availabilily of different approaches with comparable efficacy to DES, but higher safety, especially in bleeding-prone patients, have to be investigated. The EREMUS is a multicenter open-label prospective randomized three-arm clinical trial that will investigate the efficacy of a paclitaxel coated balloon + an endothelial progenitor capture stent for the treatment of native coronary lesions, and compare it to a DES strategy, or the sole endothelial progenitor capture stent. An angiographic follow-up with optical coherence tomography analysis will be scheduled 9 months after index procedure. Noninferiority regarding the primary endpoint (late luminal loss) between study group and DES is hypothesized. All patients will undergo clinical follow-up until 24 months from index hospitalization. The EREMUS trial will determine whether a composite approach with a paclitaxel coated balloon + an endothelial progenitor capture stent in coronary lesions at medium-to-high risk of restenosis will achieve similar results compared to DES regarding inhibition of intrastent proliferation; complete stent strut reendothelialization, a safety issue, will also be investigated.

Rationale and study design of the OISTER trial: Optical coherence tomography evaluation of stent struts re-endothelialization in patients with non-ST-elevation acute coronary syndromes – a comparison of the intrEpide tRapidil eluting stent vs. taxus drug-eluting stent implantation

Drug-eluting stents (DES) have been designed to prevent restenosis, but long-term clinical outcome may be offset by an increased risk of stent thrombosis, which is associated with suboptimal stent implantation or delayed re-endothelialization. DES implantation has also been associated with local persistent endothelial dysfunction. Conversely, Trapidil is a potent anti-inflammatory, vasodilatator and antiproliferative drug and several studies have shown anti-restenotic effects, suggesting substantial clinical benefits through the use of Trapidil-eluting DES. This is a longitudinal, single-blind, double-arm, randomized multicenter study. Forty patients with non-ST-elevation acute coronary syndromes who present at the index procedure with multivessel coronary disease in the major epicardial coronary arteries will be enrolled. Patients should present a culprit lesion with stenosis 70% or more associated with another stenosis 70% or more in another coronary artery. Patients will be randomized in a 1: 1 fashion to receive either an Intrepide trapidil-eluting stent or a Taxus paclitaxel-eluting stent on the culprit lesion. After 90 days, the nonculprit lesion will be treated with the stent of the opposite randomization arm and optical coherence tomography (OCT) analysis of the index stented segment will be performed. Follow-up angiography, combined with vasomotor analysis of endothelial function by rapid atrial pacing, will be done at 12 months after the index procedure on both stents. To further characterize the status of the endothelium, serum measurement of vascular endothelial growth factor gradient between the aorta and 15 mm distal to the implanted stent will be performed at 12 months. The primary endpoint of the study is to compare stent struts re-endothelialization at 90 days by OCT. The secondary endpoint is to compare angiographic outcome and coronary endothelial function 12 months after the index procedure and to compare clinical outcome at 1 and 2 years between trapidil-eluting DES versus paclitaxel-eluting DES. We hypothesize that the utilization of trapidil-eluting DES in the setting of acute coronary syndromes will be characterized by a greater early re-endothelialization associated with an antiproliferative effect offering a similar efficacy with a better safety profile compared with first-generation DES.

Molecular Basis of Restenosis and Novel Issues of Drug-Eluting Stents

Restenosis after stent deployment is an overreaction of the wound healing response after vascular injury, and is characterized by the sequence of inflammation, granulation, extracellular matrix remodeling, and smooth muscle cell (SMC) proliferation and migration. In contrast, reendothelialization of at least part of the injured vessel surface, which is essential in the wound healing process, may occur at the site of stenting. Recent advances in drug-eluting stents (DES) have substantially reduced restenosis, but do not contribute to improve long-term prognosis, compared with bare metal stents (BMS). One of the reasons may be that reendothelialization is impaired after DES stenting. Regenerated endothelial cells and proliferated SMCs in the neointima are both in part derived from their progenitor cells, which are mobilized from bone marrow to injured vessel sites and differentiate into both vascular endothelial cells and SMCs. DES inhibits mobilization and differentiation of endothelial and smooth muscle progenitor cells, and thus not only inhibits restenosis but also impairs reendothelialization, which may lead to late stent thrombosis. To improve long-term prognosis in the DES era, adjunctive medical treatments inducing early reendothelialization, but inhibiting SMC proliferation, would be required.

Abstract 3744: Local Transplantation Of Blood-derived Late-outgrowth Progenitor Cells Increased Reendothelialization, Inhibited Smooth Muscle Cell Proliferation And Reduced Neointima Formation After Experimental Carotid Injury

Introduction : Recent studies demonstrate the vasculoprotective effects of cell-based therapies to inhibit limit restenosis. Delivery of autologous late outgrowth endothelial progenitor cells (eEPC) result in early reendothelialization and inhibition of neointimal hyperplasia. Additional anti-inflammatory treatment may further reduce neointima generation. Methods and Results : CD34+ cells were isolated using immunomagnetic beads and cultured in endothelial cell medium to obtain eEPC. Early passage cells were transduced ex vivo by a retroviral vector expression of IL-1ra (eEPC IL-1ra) or empty vector (eEPC pLXSN) and expanded up to 46 population doublings. Expression levels were confirmed by RT-PCR. Athymic nude rats underwent carotid balloon and wire injury immediately followed by local delivery of eEPCs, eEPC IL-1ra, eEPC pLXSN or buffer for 20 minutes (n=22). Elastica van Giesson staining revealed a decrease in the intima/media ratio from 1.53 ± 0.11 to 1.13 ± 0.09 (mean ± SEM) after transplantation of eEPC (p=0.009). However, no additional reduction in neointima generation was observed after transplantation of eEPC IL-1ra. Endothelial-cell specific staining demonstrated an enhancement of reendothelialisation by eEPCs 24 hours after vascular injury. After 2 weeks a reduction of Ki67+ proliferating cells from 37.0 ± 8.5 Ki67+ cells in the control group to 11.67 ± 2.85 Ki67+ cells after eEPC transplantation (P=0.03) was found. Yet, no additional anti-proliferative effects were observed after transplantation of eEPC IL-1ra. Thus, enhancement of reendothelialization through local delivery of eEPCs may be sufficient for the inhibition of neointima generation since additional anti-inflammatory treatment using eEPC IL-1ra had no beneficial effect. Conclusion : Local delivery of blood-derived expanded endothelial progenitor cells after vascular injury contributes to endothelial regeneration, inhibits neointimal smooth muscle cell proliferation and reduces neointima generation. Autologous cell transplantation after vascular injury may be a feasible strategy for promotion of reendotheli-alisation and improvement of vascular regeneration.

High-Dose Granulocyte-Colony Stimulating Factor Promotes Neointimal Hyperplasia in the Early Phase and Inhibits Neointimal Hyperplasia in the Late Phase After Vascular Injury

Granulocyte-colony stimulating factor (G-CSF) affects injured arteries through early endothelialization. Some reports, however, have cautioned that the restenosis rate may increase after G-CSF injection. In the present study, high-dose G-CSF was administered to mice with vascular injury to clarify its effect. Mice were received daily subcutaneous injections of saline or a high dose (300 microg/kg) of G-CSF for 5 days after vascular injury. In the FACS analysis, CD34-/Sca-1-positive progenitor cells were more abundant in the G-CSF group (p<0.05). Neointimal hyperplasia was more evident in the G-CSF group at 1 week (p<0.05), whereas at 4 weeks it was more evident in the control group (p<0.01). TUNEL-positive cells in the arterial wall were more numerous in the G-CSF group at day 1 (p<0.01). CD34-positive cells were observed in the G-CSF group at 1 week. Re-endothelialization appeared earlier in the G-CSF group (at 4 weeks; p<0.01). An increased number of 1A4-positive smooth muscle cells were found in bone marrow cell culture treated with G-CSF. High-dose G-CSF induced neointimal proliferation through excessive inflammation and bone marrow cell mobilization in the early phase. In the late phase, however, it induced early re-endothelialization and thereby inhibited neointimal hyperplasia.

Abstract 1175: Critical Role of Bone Marrow ASC (Apoptosis-associated Speck-like Protein Containing a CARD) in Neointimal Formation After Vascular Injury in Mice

Background: ASC is an adaptor protein that forms inflammasome complex and regulates caspase-1-dependent interleukin (IL)-1β and IL-18 generation; however, the role of ASC in the pathophysiology of cardiovascular diseases remains undefined. Here, we investigated the contribution of ASC to neointimal formation after vascular injury in ASC-deficient (ASC −/− ) mice. Methods and Results: Wire-mediated vascular injury was produced in the femoral artery of ASC −/− and wild-type (WT: C57BL/6) mice. Immunohistochemical analysis revealed that ASC was markedly expressed in the infiltrated macrophages in the neointima. Histological analysis revealed that neointimal formation was significantly reduced in ASC −/− mice as compared to that in the WT mice (I/M ratio, p = 0.007). IL-1β and IL-18 were clearly expressed in the neointimal lesion of the WT mice but showed decreased expression in the lesion of ASC −/− mice. However, TUNEL staining showed no significant difference in apoptotic cells of the media between these mice. Further, no significant differences were observed in early reendothelialization (CD31) and the number of macrophages (F4/80) and smooth muscle cells (SMCs: αSMA) per unit of the neointima. Flow cytometry analysis also showed no difference in the number of peripheral CD34 + /Flk-1 + cells (EPCs) after injury. To determine the contribution of bone marrow cells, we developed 3 types of bone marrow transplantation (BMT Wild+Wild , BMT Wild+ASC−/− , and BMT ASC−/− → Wild ) mice. The neointimal formation in BMT ASC−/−+Wild mice mice was remarkably reduced as compared to that in BMT Wild+Wild (I/M ratio, p = 0.0003) and BMT Wild+ASC−/− mice (p = 0.015), indicating that bone marrow ASC is critical for neointimal formation. In vitro experiments showed that the lipopolysaccharide-induced production of inflammatory cytokines such as IL-1β, IL-6, MCP-1, and IFN-γ, but not TNF-α, in ASC −/− bone marrow cells was significantly decreased. Further, serum-stimulated proliferation activity and MAP kinase (ERK1/2 and p38) activation of ASC −/− SMCs were not impaired. Conclusions: These findings suggest that bone marrow-derived ASC is critical for the neointimal formation after vascular injury and identify ASC as a novel therapeutic target for cardiovascular diseases.

Pathophysiology of stem cells in restenosis.

Recent evidence has shown that vascular function depends not only on cells within the vessels, but is also significantly modulated by circulating cells derived from the bone marrow. A number of studies indicate that an early reendothelialization by circulating endothelial precursors after vascular injury prevents excessive cell proliferation and restenosis. Conversely, other studies concluded that the homing of other cell fractions, consisting mainly of smooth muscle precursors, cause pathological remodelling. Different cell types have been identified and characterized so far as circulating precursors able to participate in vascular repair by homing and differentiating towards endothelial cells or smooth muscle cells. Among these, endothelial precursor cells, smooth muscle progenitor cells, mesenchymal stem cells and others have been described. The origins, the hierarchy, the role and the markers of these different cell populations are still controversial. Nevertheless, different strategies have been developed so far in animal models to induce the mobilization and the recruitment of stem cells to the injury site, based on physical training, hormone injection and application of stem cell-capturing coated stents. It should also be mentioned that the limited data currently available derived from clinical trials provide contrasting results about the effective role of vascular cell precursors in restenosis prevention, thus indicating that conclusions derived from studies in animal models cannot always be directly applied to humans and that caution should be used in the manipulation of circulating progenitor cells for therapeutic strategies.

Vasculoprotective Effects of Erythropoietin

The body of evidence demonstrating the vasculoprotective effects of direct cell-based therapies to inhibit the acute response to vascular injury continues to grow. Delivery of autologous circulation-derived cells capable of assuming an endothelial phenotype to an injured arterial segment results in early reendothelialization and normalization of vascular function.1–3 Current applications of this approach require cell isolation and modification in vitro, followed by direct delivery. However, novel approaches are being developed to avoid the inherent translational concerns of this strategy. Broadly, these new approaches include the mobilization and targeting of cell populations capable of augmenting reendothelialization of injured arteries while limiting neointimal formation. Multiple cytokines have been shown to mobilize cells capable of assuming an endothelial phenotype (broadly known as EPCs). These cytokines include VEGF, G-CSF, GM-CSF, and erythropoietin (Epo). The article by Urao et al4 in this issue of Circulation Research extends the observation that Epo can mobilize EPCs while demonstrating its ability to enhance reendothelialization and inhibit neointimal formation after vascular injury in an NO-dependent manner. Taken together, these findings extend our understanding of the role of Epo as a tissue protectant and apply it to the setting of acute vascular injury. The article by Urao tested the hypothesis that systemic delivery of recombinant human Epo would inhibit neointimal formation in a mouse carotid artery wire injury model.4 Epo (1000 IU/kg of body weight) was administered for 3 days beginning at the time of arterial injury. Morphometric analysis of serial arterial sections performed at 14 days after injury showed a 52% decrease in the neointimal area in the Epo-treated mice compared with the saline injected …

Rosiglitazone facilitates angiogenic progenitor cell differentiation toward endothelial lineage: a new paradigm in glitazone pleiotropy.

Peroxisome proliferator-activated receptor-gamma (PPAR-gamma) agonists inhibit vascular smooth muscle proliferation and migration and improve endothelial function. It is unknown whether PPAR-gamma agonists favorably modulate bone marrow (BM)-derived angiogenic progenitor cells (APCs) to promote endothelial lineage differentiation and early reendothelialization after vascular intervention. C57/BL6 mice, treated with or without rosiglitazone (8 mg/kg per day), a PPAR-gamma agonist, underwent femoral angioplasty. Rosiglitazone treatment attenuated neointimal formation (intima/media ratio: 0.98+/-0.12 [rosiglitazone] versus 3.1+/-0.5 [control]; P<0.001; n=10 per group). Using a BM transplantation model, we identified that 58+/-12% of the cells within the neointima at 4 weeks were derived from the BM. Pure endothelial marker-positive, pure alpha-smooth muscle actin (alphaSMA)-positive, or double-positive APCs could be found both in mouse BM and in human peripheral blood after culture in conditional medium enriched with vascular endothelial growth factor. Rosiglitazone caused a 6-fold (P<0.001) increase in colony formation by human endothelial progenitor cells, promoted the differentiation of APCs toward the endothelial lineage in mouse BM in vivo (0.66+/-0.06% [control] to 0.95+/-0.08% [rosiglitazone]; P<0.05) and in human peripheral blood in vitro (13.2+/-1.5% [control] to 28.4+/-3.3% [rosiglitazone]; P<0.05), and inhibited the differentiation toward the smooth muscle cell lineage. Within the neointima, rosiglitazone also stimulated APCs to differentiate into mature endothelial cells and caused earlier reendothelialization compared with controls (31+/-5 versus 8+/-2 CD31-positive cells per millimeter of neointimal surface on day 14; P<0.01). Similar to embryonic stem cell-derived progenitors, the adult BM and peripheral blood harbor APCs that are at least bipotential and able to differentiate into endothelial and smooth muscle lineages. The PPAR-gamma agonist rosiglitazone promotes the differentiation of these APCs toward the endothelial lineage and attenuates restenosis after angioplasty.

End-to-side and end-to-end vascular anastomoses with a carbon dioxide laser

This study was designed to compare anastomoses performed with a carbon dioxide laser and conventional anastomoses performed with 7-0 polypropylene suture. In each of 80 rabbits, the divided left carotid artery was anastomosed by a continuous suture technique and the right carotid was anastomosed with a carbon dioxide laser. In each of 40 additional rabbits, both end-to-end and end-to-side laser anastomoses were performed on the same carotid artery. The laser technique involved the placement of three stay sutures (end-to-end technique) or four stay sutures (end-to-side technique) of 7-0 polypropylene and an everting laser seal at a power level of 65 mW. The 1-year overall patency rate was 98% (78/80) in laser anastomoses, 79% (63/80) in suture anastomoses, and 95% (38/40) in combined end-to-end and end-to-side laser anastomoses. Microscopic findings in laser anastomoses demonstrated degeneration of collagen and protein in the adventitia and media, but much less intimal injury than in suture anastomoses, with reendothelialization beginning earlier (within 7 days after anastomosis as compared with 2 to 4 weeks). The tissue tensile strength at 1 hour was less in laser anastomoses than in suture anastomoses, but the laser anastomoses still withstood an intraluminal pressure load of 380 mm Hg. Laser anastomosis improved the microscopic and histologic appearance of the intimal layer, allowing for rapid early reendothelialization and resulting in excellent patency rates.