The development of small-caliber grafts still represents a challenge in the field of vascular prostheses. Among other factors, the mechanical properties mismatch between natural vessels and artificial devices limits the efficacy of state-of-the-art materials. In this paper, a novel nanocomposite graft with an internal diameter of 6 nun is proposed. The device is obtained through spray deposition using a semi-interpenetrating polymeric network combining poly(ether)urethane and polydimethilsyloxane. The inclusion of BaTiO3 nanoparticles endows the scaffold with piezoelectric properties, which may be exploited in the future to trigger beneficial biological effects. Graft characterization demonstrated a good nanoparticle dispersion and an overall porosity that was not influenced by the presence of nanoparticles. Graft mechanical properties resembled (or even ameliorated) the ones of natural vessels: both doped and non-doped samples showed a Young's modulus of similar to 700 kPa in the radial direction and similar to 900 kPa in the longitudinal direction, an ultimate tensile strength of similar to 1 MPa, a strain to failure of similar to 700%, a suture retention force of similar to 1.7 N and a flexural rigidity of similar to 2.5 x 10(-5) N m(2). The two grafts differed in terms of burst strength that resulted similar to 800 kPa for the control non-doped samples and similar to 1100 kPa for the doped ones. The graft doped with BaTiO3 nanoparticles showed a d(33) coefficient of 1.91 pm/V, almost double than the non-doped control. The device resulted highly stable, with a mass loss smaller than 2% over 3 months and an excellent biocompatibility.

Platelets contain abundant growth factors and cytokines that have a positive influence on the migration and proliferation of different cell types by modulating its physiopathological processes. As it is known that human umbilical cord blood platelet lysate (UCB-PL) contains a supraphysiological concentration of growth factors, in the present study, we investigated its effectiveness in wound-healing processes. Human UCB-PL was obtained by the freeze/thaw of platelet concentrate (1.1 × 109 platelets/L), and its effect was evaluated on human or mouse endothelial cells, monocytes, fibroblasts, and keratinocytes in different concentrations. Human UCB-PL was observed to have high levels of pro-angiogenic growth factor than peripheral blood platelet-rich plasma. Among the cell lines, different concentrations of human UCB-PL were necessary to influence their viability and proliferation. For L929 cells, 5% of total volume was necessary, while for human umbilical vein endothelial cell, it was 10%. Cell migration on monocytes was increased with respect to the positive control, and scratch closure on keratinocytes was increased with respect to serum-free medium with only 10% of human UCB-PL. We concluded that the human UCB-PL may be useful to produce a large amount of standard platelet concentrates sufficient for several clinical-scale expansions avoiding inter-individual variability, which can also be used as a functional tool for clinical regenerative application for wound healing.

Endothelial progenitor cells (EPCs) contribute to ischemic tissue repair by paracrine secretion up-regulated by hypoxia. In this study we use novel nanoparticles (NPs) as carriers for a controlled release of EPC secretome (CM) to improve their angiogenic properties. The in vivo effect in ischemic hindlimb rat model was evaluated, comparing hypoxic EPC-CM-NPs with hypoxic EPC-CM alone. A proteomic characterization of hypoxic CM and the in vitro effect on endothelial cells (HUVECs) were also performed. Up to 647 protein, 17 of which with angiogenic properties, were upregulated by hypoxia. Moreover, hypoxic EPC-CM significantly promoted capillary-like structures on Matrigel. A significant increase of blood perfusion in ischemic limbs at 2 weeks with EPCCM-loaded NPs as compared to both EPC-CM and control and a significant increase of capillary formation were observed. The use of EPC-CM-NPs significantly improved neoangiogenesis in vivo, underlining the advantages of controlled release in regenerative medicine.

As the endothelium still represents the ideal surface for cardiovascular devices, different endothelialization strategies have been attempted for biocompatibility and nonthrombogenicity enhancement. Since endothelial progenitor cells (EPCs) could accelerate endothelialization, preventing thrombosis and restenosis, the aim of this study was to use oligonucleotides (ONs) to biofunctionalize stents for EPC binding. In order to optimize the functionalization procedure before its application to cobalt–chromium (Co/Cr) stents, discs of the same material were preliminarily used. Surface aminosilanization was assessed by infrared spectroscopy and scanning electron microscopy. A fluorescent endothelial-specific ON was immobilized on aminosilanized surfaces and its presence was visualized by confocal microscopy. Fluorescent ON binding to porcine blood EPCs was assessed by flow cytometry. Viability assay was performed on EPCs cultured on unmodified, nontargeting ON or specific ON-coated discs; fluorescent staining of nuclei and F-actin was then performed on EPCs cultured on unmodified or specific ON-coated discs and stents. Disc biofunctionalization significantly increased EPC viability as compared to both unmodified and nontargeting ON-coated surfaces; cell adhesion was also significantly increased. Stents were successfully functionalized with the specific ON, and EPC binding was confirmed by confocal microscopy. In conclusion, stent biofunctionalization for EPC binding was successfully achieved in vitro , suggesting its use to obtain in vivo endothelialization, exploiting the natural regenerative potential of the human body. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 3284–3292, 2015.

Endothelialprogenitorcells(EPCs)aremobilizedfrombonemarrowinto peripheral blood, contributing to the revascularization of ischemicareas, to endothelial repair and to the physiological maintenance ofvascularization. EPC mobilization and homing have been primarilylinked to ischemia and inflammation presence [1]. EPCs are directlycorrelated with endothelial function and inversely correlated withcardiovascular risk factors and atherosclerosis progression [2].EPClevelshavealsobeencorrelatedtoprognosisevaluationincardiovascu-lar disease [3].Regarding EPC correlation with coronary artery disease (CAD)presence and severity, an inverse relationship with CAD severity, inde-pendent of traditional risk factors, was demonstrated by EPC colonycounting [4],whileahighnumberofEPCsassociatedwithCADandcorre-latedwithstenosisseveritywereshownby flowcytometry [5].Recentlyanew protocol, adapted from the standardized ISHAGE protocol for hema-topoieticstemcells,hasbeendevelopedforEPClevelevaluationtoenablecomparison of clinical and laboratory data [6].While thepresence of circulatingEPCshasbeenwidely evaluated indifferent diseases, few studies tried to evaluate the presence of EPCs inhuman vital myocardium.TheaimofourstudywastoinvestigateEPClevelsbothinperipheralbloodand inmyocardium in thesame patients atthesame time, evalu-atingthecorrelationwith CADpresence. In bothsampleswequantifiedCD34

No scientific evidence concerning the presence and role of endothelial progenitor cells (EPCs) in human vital myocardium exists. This study aimed to investigate the correlation between peripheral and myocardial EPC number and coronary artery disease (CAD) severity. 12 consecutive CAD patients, 5

Background Platelets are rich in mediators able to positively affect cell activity in wound healing. Aim of this study was to characterize the effect of different concentrations of human pooled allogeneic platelet lysate on human cells involved in the different phases of wound healing (inflammatory phase, angiogenesis, extracellular matrix secretion and epithelialization). Methodology/Principal Findings Platelet lysate effect was studied on endothelial cells, monocytes, fibroblasts and keratinocytes, in terms of viability and proliferation, migration, angiogenesis, tissue repair pathway activation (ERK1/2) and inflammatory response evaluation (NFκB). Results were compared both with basal medium and with a positive control containing serum and growth factors. Platelet lysate induced viability and proliferation at the highest concentrations tested (10% and 20% v/v). Whereas both platelet lysate concentrations increased cell migration, only 20% platelet lysate was able to significantly promote angiogenic activity (p<0.05 vs. control), comparably to the positive control. Both platelet lysate concentrations activated important inflammatory pathways such as ERK1/2 and NFκB with the same early kinetics, whereas the effect was different for later time-points. Conclusion/Significance These data suggest the possibility of using allogeneic platelet lysate as both an alternative to growth factors commonly used for cell culture and as a tool for clinical regenerative application for wound healing.

Age represents a significant risk factor for the onset and progression of cardiovascular disease, with the increase in life expectancy in developed countries going in parallel with increased incidence of such pathologies. Treatment strategies alternative or additive to pharmacological treatments are needed. The relationship between aging and progenitor cell-mediated repair is of great interest. Endothelial progenitor cells (EPC) mediate repair mechanisms for endothelial regeneration and maintenance, but they are subject to age-associated changes affecting negatively their number and/or function.Aim of this review is to examine the impact of age on EPC-mediated vascular repair, with a focus on the metabolic pathways involved and on the therapeutic targets with potential for attenuating this effect.

Purpose: Platelet lysate (PL) contains growth factors, cytokines and chemokines useful for angiogenesis and tissue repair. Polymer-based nanoparticles (NP) could be used as PL carriers for a controlled release in ischemia. Our aim was to characterize the in vitro (viability, tissue repair and inflammatory response activation) and in vivo (injection in rat ischemic hindlimb and Laser Doppler Perfusion Imaging, LDPI) effect of PL-loaded NP. Methods: Human umbilical vein endothelial cells (HUVEC) were obtained from healthy donors. Experiments were performed at p3, with a pool from 3 donors. PL obtained by thrombapheresis from healthy donors was resuspended (6x108 thrombocytes/ml), subjected to 3 freeze-thaw cycles and stored at -80°C. Two polymers, Poly(Gly-co-HPMA) (fast drug release, e.g. few days) and PLGA (slow drug release, e.g. several weeks), were used to obtain NP for different application. HUVEC were incubated for 48h at 37°C with PL-loaded NP (corresponding to 1.5% PL), with 5% FBS and EGF to assess cell viability. ERK1/2 and NFkB were detected by FACE ELISA, monitoring proteins activated by phosphorylation. HUVEC were treated with PL-loaded NP for 30 and 60' (ERK 1/2) or 1, 24 and 48h (NFkB). Unilateral hindlimb ischemia was induced in adult male Wistar rats (n=24) by surgical excision of the femoral vessels. Blood flow was monitored by LDPI at different time points, up to 21d. After 7d, intramuscular injections of PL, PL-loaded fast NP and saline were performed using fibrin for enhancing engraftment; 6 rats received no treatment. Blood flow was monitored by LDPI. Results: PL-loaded NP did not affect cell viability. PL-loaded fast NP induced ERK1/2 activation at 60', unlike PL-loaded slow NP. PL-loaded slow NP activated NFkB at 48 hours, unlike PL-loaded fast NP. After a short postoperative period of depressed perfusion (p<0.005), the rats showed moderate hindlimb ischemia from d7 onwards, with constant decreased perfusion. Rats receiving only fibrin were comparable to untreated rats. At 7d after intramuscular injection, PL-loaded fast NP significantly increased blood flow (1.01±0.15 ischemic/contralateral hind limb perfusion ratio) in the ischemic hindlimb vs. PL (0.78±0.12, p<0.05). Conclusions: The nanosystems showed cytocompatibility and a differential capability to activate important inflammatory pathways such as ERK 1/2 and NFkB, involved in angiogenesis and tissue repair. Release of PL from loaded NP was effective for blood flow enhancement in a model of chronic, moderate hindlimb ischemia, underlining the advantages of using such drug-controlled release in regenerative medicine.

INTRODUCTION:Conventional therapy for venous thromboembolism or acute coronary syndrome involves the administration of glycoanticoagulants (heparins) or oligosaccharides (fondaparinux). We evaluated the effects of such drugs on angiogenesis and vasculogenesis-like models. MATERIALS AND METHODS:Human umbilical vein endothelial cells or human endothelial progenitor cells were treated with bemiparin, fondaparinux or unfractionated heparin, at concentrations reflecting the doses used in clinical practice. After 24h, cell viability, proliferation, tubule formation and angiogenic molecular mechanisms, such as activation of the serine/threonine kinase AKT, were assessed. In vivo angiogenesis was studied using a Matrigel sponge assay in mice. RESULTS:Bemiparin gave a significant decrease of in vitro angiogenesis as shown by the reduction of endothelial cell tubule network, while both fondaparinux and unfractionated heparin did not show any significant effect. In assays of Matrigel sponge invasion in mice, unfractionated heparin was able to stimulate angiogenesis and, conversely, bemiparin inhibited angiogenesis. Furthermore, both bemiparin and fondaparinux caused a significant reduction in an in vitro vasculogenesis-like model, as demonstrated by the decrease of tubule network after co-seeding of endothelial progenitor cells and human umbilical vein endothelial cells. In addition, unfractionated heparin but not bemiparin was able to increase AKT phosphorylation. CONCLUSIONS:In in vitro experiments, bemiparin was the only drug to show an anti-angiogenic and vasculogenic-like effect, unfractionated heparin showed only a trend to increase in angiogenesis assay and fondaparinux affected only the vasculogenesis-like model. Notably, the in vivo experiments corroborated these data. Such results are important for the choice of a patient-tailored therapy.

Background Foam sclerotherapy has been proven to be a safe and effective treatment for superficial venous insufficiency, but transient visual and neurologic disturbances continue to be reported. These side effects have been theorized to be related to the presence of air or gases in the sclerosing foam that results in “bubble” migration into the cerebral circulation. We present a differing hypothesis that significant amounts of endothelin are released from the treated veins, amounts capable of causing these complications. Material and Methods We tested the release of endothelin 1 (ET-1) in 12 rats after sclerotherapy with sodium tetradecyl sulfate (STS) in liquid and foam preparations. In 11 human subjects, we measured ET-1 in systemic circulation and in a draining vein after foam sclerotherapy with polidocanol. Results Rats treated with STS showed a significant increase in ET-1 levels 1 and 5 minutes after foam sclerotherapy. Patients treated with foam sclerotherapy showed a marked increase in ET-1 levels that correlated significantly with local ET-1 levels. Conclusions Evidence of ET-1 release represents a plausible relationship explaining neurologic and visual disturbances reported after sclerotherapy.