[show abstract][hide abstract] ABSTRACT: The aims of this study were the development, characterization and bioevaluation of a novel biocompatible, resorbable and bio-active wound dressing prototype, based on anionic polymers (sodium alginate - AlgNa, carboximethylcellulose - CMC) and magnetic nanoparticles loaded with usnic acid (Fe3O4@UA). The antimicrobial activity was tested against Staphylococcus aureus grown in biofilms. The biocompatibility testing model included an endothelial cell line from human umbilical vein and human fetal progenitor cells derived from the amniotic fluid, that express a wide spectrum of surface molecules involved in different vascular functions and inflammatory response, and may be used as skin regenerative support. The obtained results demonstrated that CMC/Fe3O4@UA and AlgNa/Fe3O4@UA are exhibiting structural and functional properties that recommend them for further applications in the biomedical field. They could be used alone or coated with different bio-active compounds as Fe3O4@UA for the development of novel, multifunctional porous materials used in tissues regeneration, as antimicrobial substances releasing devices, providing also a mechanical support for the eukaryotic cells adhesion, and exhibiting the advantage of low cytotoxicity on human progenitor cells. The great antimicrobial properties exhibited by the newly synthesized nano-bioactive coatings, are recommending them as successful candidates for improving implanted devices surfaces used in regenerative medicine.
International journal of pharmaceutics 08/2013; · 2.96 Impact Factor
[show abstract][hide abstract] ABSTRACT: AIM: The purpose of this project was to evaluate the influence of circulating endothelial progenitor cells (EPCs) and platelet microparticles (PMPs) on blood platelet function in experimental hypertension associated with hypercholesterolemia. METHODS: GOLDEN SYRIAN HAMSTERS WERE DIVIDED IN SIX GROUPS: (i) control, C; (ii) hypertensive-hypercholesterolemic, HH; (iii) 'prevention', HHin-EPCs, HH animals fed a HH diet and treated with EPCs; (iv) 'regression', HHfin-EPCs, HH treated with EPCs after HH feeding; (v) HH treated with PMPs, HH-PMPs, and (vi) HH treated with EPCs and PMPs, HH-EPCs-PMPs. RESULTS: Compared to HH group, the platelets from HHin-EPCs and HHfin-EPCs groups showed a reduction of: (i) activation, reflected by decreased integrin 3β, FAK, PI3K, src protein expression; (ii) secreted molecules as: SDF-1, MCP-1, RANTES, VEGF, PF4, PDGF and (iii) expression of pro-inflammatory molecules as: SDF-1, MCP-1, RANTES, IL-6, IL-1β; TFPI secretion was increased. Compared to HH group, platelets of HH-PMPs group showed increased activation, molecules release and proteins expression. Compared to HH-PMPs group the combination EPCs with PMPs treatment induced a decrease of all investigated platelet molecules, however not comparable with that recorded when EPC individual treatment was applied. CONCLUSION: EPCs have the ability to reduce platelet activation and to modulate their pro-inflammatory and anti-thrombogenic properties in hypertension associated with hypercholesterolemia. Although, PMPs have several beneficial effects in combination with EPCs, these did not improve the EPC effects. These findings reveal a new biological role of circulating EPCs in platelet function regulation, and may contribute to understand their cross talk, and the mechanisms of atherosclerosis.
PLoS ONE 01/2013; 8(1):e52058. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: This study aimed to (i) employ our newly designed model, the hypertensive-hypercholesterolemic hamster (HH), in order to find out whether a correlation exists between circulating microparticles (MPs), endothelial progenitor cells (EPCs) and their contribution to vascular dysfunction and (ii) to assess the effect of irbesartan treatment on HH animals (HHI).
The results showed that compared with the control (C) group, HH displayed: (i) a significant increase in plasma cholesterol and triglyceride concentration, and an augmentation of systolic and diastolic arterial blood pressure, and of heart rate; (ii) a marked elevation of MPs and a significant decrease in EPCs; (iii) structural modifications of the arterial wall correlated with altered protein expression of MMP2, MMP9, MMP12, TIMP1, TIMP2 and collagen type I and III; (iv) a considerably altered reactivity of the arterial wall closely correlated with MPs and EPC adherence; and (v) an inflammatory process characterized by augmented expression of P-Selectin, E-Selectin, von Willebrand factor, tissue factor, IL-6, MCP-1 and RANTES. Additionally, the experiments showed the potential of irbesartan to correct all altered parameters in HH and to mobilize EPCs by NO, chemokines and adhesion molecule-dependent mechanisms.
Hypertension associated with hypercholesterolemia is accompanied by structural modifications and expression of pro-inflammatory molecules by the vessel wall, the alteration of vascular tone, enhanced release of MPs and reduced EPCs; the ratio between the latter two may be considered as a marker of vascular dysfunction. Irbesartan, which exhibits a pharmacological control on the levels of MPs and EPCs, has the potential to restore homeostasis of the arterial wall.
Journal of Thrombosis and Haemostasis 02/2012; 10(4):680-91. · 6.08 Impact Factor
[show abstract][hide abstract] ABSTRACT: To test the involvement of histone deacetylases (HDACs) activity in endothelial lineage progression, we investigated the effects of HDAC inhibitors on endothelial progenitors cells (EPCs) derived from umbilical cord blood (UCB). Adherent EPCs, that expressed the endothelial marker proteins (PCAM-1, CD105, CD133, and VEGFR(2)) revealed by flow cytometry were treated with three HDAC inhibitors: Butyrate (BuA), Trichostatin A (TSA), and Valproic acid (VPA). RT-PCR assay showed that HDAC inhibitors down-regulated the expression of endothelial genes such as VE-cadherin, CD133, CXCR4 and Tie-2. Furthermore, flow cytometry analysis illustrated that HDAC inhibitors selectively reduce the expression of VEGFR(2), CD117, VE-cadherin, and ICAM-1, whereas the expression of CD34 and CD45 remained unchanged, demonstrating that HDAC is involved in endothelial differentiation of progenitor cells. Real-Time PCR demonstrated that TSA down-regulated telomerase activity probably via suppression of hTERT expression, suggesting that HDAC inhibitor decreased cell proliferation. Cell motility was also decreased after treatment with HDAC inhibitors as shown by wound-healing assay. The balance of acethylation/deacethylation kept in control by the activity of HAT (histone acetyltransferases)/HDAC enzymes play an important role in differentiation of stem cells by regulating proliferation and endothelial lineage commitment.
International Journal of Molecular Sciences 01/2012; 13(11):15074-85. · 2.46 Impact Factor
[show abstract][hide abstract] ABSTRACT: Promise of cell therapy has advanced the use of adult stem cells towards the development of novel approaches to promote regeneration
of injured endothelium. The aim of this study was to stimulate endothelial progenitor cells (EPCs) with lectin isolated from
Solanum tuberosum (potato) shoot and Calendula officinalis (marigold) extracts, in order to increase EPCs proliferation and gene expression of molecules with roles in chemotaxis and adhesion
for a better attachment to injured vascular tissue. EPCs were differentiated from umbilical cord blood-derived mononuclear
cells and characterized by light microscopy, flow cytometry, and vascular tube-like structures formation on Matrigel. Cell
proliferation was determined by MTS assay, and gene expression of molecules involved in EPCs adhesion (VCAM-1, VE-cadherin,
ICAM-1, PECAM-1, P-selectin) and chemotaxis was determined (CXCR4, Tie-2) by RT-PCR. For the assessment of cell motility,
wound-healing assay was employed. Both potato shoot lectin and marigold extracts stimulated EPCs proliferation in a concentration dependent manner and were able to increase expression of adhesion
and chemotactic molecules. Marigold flower extract proved to be more efficient. This study demonstrates the usefulness of potato lectin and marigold extracts to increase EPCs proliferation and modulate gene expression of chemotactic and adhesion molecules, which may facilitate
EPCs attachment to injured endothelium.
Central European Journal of Biology 01/2011; 6(3):330-341. · 0.82 Impact Factor
[show abstract][hide abstract] ABSTRACT: Wharton's jelly (WJ) is a rich source of multiple-lineage differentiating cells, recently proposed for cell replacement therapy. However, their ability to integrate into the cardiac tissue has not been elucidated, yet. We employed in vitro cardiac transplantation models to investigate the capacity of a novel population of human WJ-derived mesenchymal stem cells (nMSCs) to integrate into both living and ischemic cardiac tissue. NMSCs were characterized for the expression of stem/progenitor cell genes and proteins, as well as for multi-lineage differentiation potential. To assess their integration properties, nMSCs were cocultured with either living or ischemic embryonic murine ventricular slices. Immunohistochemical analyses were performed on cryosections of cocultured preparations to allow human cells tracking within the cocultures. Results showed that nMSCs shared MSC and endothelial colony-forming cell characteristics at gene, protein, and functional levels. NMSCs were markedly chemoattracted towards the ventricular slices, integrating robustly into the depth of both living and ischemic cardiac tissue. In conclusion, the functional ability of WJ-derived cells to populate the cardiac tissue could be validated in vitro. The transplantation models described could be further used to depict the mechanisms of WJ-derived cells integration into the cardiac tissue, contributing to optimization of reliable cell therapies for cardiac repair.
Cellular Physiology and Biochemistry 01/2011; 28(1):63-76. · 3.42 Impact Factor
[show abstract][hide abstract] ABSTRACT: The umbilical cord blood derived endothelial progenitor cells (EPCs) contribute to vascular regeneration in experimental models of ischaemia. However, their ability to participate in cardiovascular tissue restoration has not been elucidated yet. We employed a novel coculture system to investigate whether human EPCs have the capacity to integrate into living and ischaemic cardiac tissue, and participate to neovascularization. EPCs were cocultured with either living or ischaemic murine embryonic ventricular slices, in the presence or absence of a pro-angiogenic growth factor cocktail consisting of VEGF, IGF-1, EGF and bFGF. Tracking of EPCs within the cocultures was performed by cell transfection with green fluorescent protein or by immunostaining performed with anti-human vWF, CD31, nuclei and mitochondria antibodies. EPCs generated vascular tube-like structures in direct contact with the living ventricular slices. Furthermore, the pro-angiogenic growth factor cocktail reduced significantly tubes formation. Coculture of EPCs with the living ventricular slices in a transwell system did not lead to vascular tube-like structures formation, demonstrating that the direct contact is necessary and that the soluble factors secreted by the living slices were not sufficient for their induction. No vascular tubes were formed when EPCs were cocultured with ischaemic ventricular slices, even in the presence of the pro-angiogenic cocktail. In conclusion, EPCs form vascular tube-like structures in contact with living cardiac tissue and the direct cell-to-cell interaction is a prerequisite for their induction. Understanding the cardiac niche and micro-environmental interactions that regulate EPCs integration and neovascularization are essential for applying these cells to cardiovascular regeneration.
Journal of Cellular and Molecular Medicine 10/2010; 15(9):1914-26. · 4.75 Impact Factor
[show abstract][hide abstract] ABSTRACT: The aim of this study was to determine the effect of simultaneous hypertension and hypercholesterolemia on platelet activation, nitric oxide (NO) production and oxidative stress, and to evaluate the role of irbesartan, an angiotensin II type 1 receptor antagonist.
Golden Syrian hamsters were divided into three groups: controls, C (fed a standard diet); hypertensive-hypercholesterolemic, HH (fed a diet enriched in 3% cholesterol, 15% butter and 8% NaCl, for 4 months); and hypertensive-hypercholesterolemic treated with irbesartan, HHI (fed as HH group, plus irbesartan 10 mg kg(-1) per day, for 4 months).
Compared with the C group, platelets isolated from the HH group showed: morphological modifications; increased integrin β3 exposure and protein expression of P-selectin, FAK, PI3K, Akt and Src; reduced eNOS protein expression and NO production; higher generation of ROS, mostly produced by NADPH-oxidase, cyclooxygenase-1 (COX-1) and 12-lipoxygenase; and enhanced NAD(P)H oxidase activity and protein expression of gp91phox and p22phox subunits, 12-lipoxygenase, COX-1, cPLA(2) and PKC. Compared with the HH group, the treatment with irbesartan (HHI group) significantly attenuates the changes in all the molecules tested, reduces platelet aggregation, and improves intraplatelet redox balance.
Experimental hypertension associated with hypercholesterolemia produces major changes in morphology, signaling mechanisms and oxidative stress in blood platelets. These changes were significantly diminished by irbesartan administration, which functions as an antioxidant on platelets.
Journal of Thrombosis and Haemostasis 10/2010; 9(1):173-84. · 6.08 Impact Factor
[show abstract][hide abstract] ABSTRACT: Chronic venous insufficiency (CVI) results when the veins in the legs no longer pump blood back to the heart effectively. Microparticles (MPs) are small membrane vesicles released by several circulating and vascular cells upon activation or apoptosis.
The purpose of this study was to assess the subpopulations of circulating endothelial (EMPs) and platelet microparticles (PMPs) in CVI, and to disclose their contribution in mediating dysfunction of human peripheral venules.
Human peripheral venules were explanted during leg surgery on patients with CVI and on control subjects (C); concurrently, blood samples were collected and circulating MPs isolated. The techniques used were: flow cytometry, fluorescence and electron microscopy, myograph technique and western-blotting technique.
The results showed that compared with controls, patients with CVI had: (i) a marked elevation of circulating EMPs and PMPs; (ii) a structural modification of the venous wall consisting of activation of endothelial and smooth muscle cells, an abundance of intermediary filaments and synthesis of hyperplasic-multilayered basal lamina; (iii) a significantly altered reactivity of the venous wall, closely associated with EMPs and PMPs adherence; (iv) altered contractile response to noradrenaline, acetylcholine, 5-hydroxytryptamine and KCl, and an impeded relaxation in response to sodium nitroprusside; and (iv) a substantially increased protein expression of tissue factor (TF) and of P-Selectin both in the venular vascular wall and on the surface of EMPs and PMPs.
The findings indicate that CVI is accompanied by an enhanced release of EMPs and PMPs that contribute to altered dysfunctional response of the venous wall.
Journal of Thrombosis and Haemostasis 07/2009; 7(9):1566-75. · 6.08 Impact Factor
[show abstract][hide abstract] ABSTRACT: External bile duct fistulas are inherent postoperative complications that usually appear after biliary tract surgery, traumatic bile duct injuries and liver surgery for hepatic hydatid disease or liver transplant. The management is highly individualized, while the success and long-term results of endoscopic and surgical techniques are conflicting. The study included 32 cases with external bile duct fistulas managed by endoscopic retrograde cholangiography (ERC) with sphincterotomy and/or stent placement, including "rendez-vous" procedures in 2 cases. The causes of the external fistula were represented by cholecystectomy with/without retained common bile duct stones or strictures (22 cases), cholecystectomy and drainage of a subphrenic abscess caused by severe acute pancreatitis (1 case) and surgical interventions for hepatic hydatid disease (9 cases). Due to the prospective protocol of the study we were able to apply an individualized endoscopic treatment: sphincterotomy with proper relief of the bile duct obstruction (stone extraction) or sphincterotomy with large-size (10 Fr) stent placement for large-sized bile duct defects. The results consisted in closure of the fistula in 3.5 +/- 1.7 days for the subgroup of patients with sphincterotomy alone. Among the patients with stent insertion, fistulas healed slower in 14 +/- 3.5 days. There were no complications after endoscopic treatment; however the stent could not be passed in one patient that required subsequent surgery. In conclusion, endoscopic intervention is the treatment of choice for small external biliary fistulas complicating biliary tract surgery or liver surgery for hepatic hydatid disease. When the fistula is large, the placement of a 10 Fr endoprosthesis becomes necessary, while failure of endoscopic treatment leads to surgery with hepatico-jejunal anastomosis.