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1.
J Vasc Res ; 46(4): 278-89, 2009.
Article in English | MEDLINE | ID: mdl-19033690

ABSTRACT

Neointima formation participates in the pathophysiology of atherosclerosis and restenosis. Proliferation and migration of vascular smooth muscle cells (VSMC) are initial responses to vascular injury. The aim of the present study was to assess the effect of gliotoxin, an inhibitor of nuclear factor (NF)-kappaB, on migration and proliferation of cultured rat VSMC and neointimal formation in injured rat vessels. In cultured VSMC, gliotoxin inhibited the nuclear translocation of the p65 subunit of NF-kappaB in response to inflammatory stimuli. In addition, gliotoxin inhibited VSMC migration and proliferation in response to platelet-derived growth factor-BB. This was associated with a rapid rearrangement of the F-actin and vimentin cytoskeleton. Furthermore, gliotoxin inhibited endothelial cell nuclear translocation of p65, cell surface expression of adhesion molecules such as VCAM-1, ICAM-1 and E-selectin, and monocytic cell adhesion to a cytokine-activated endothelial monolayer. In the rat carotid artery balloon catheter injury model, the systemic administration of gliotoxin for 10 days decreased neointimal hyperplasia and luminal stenosis by up to 90% and decreased the expression of proliferating cell nuclear antigen in the vessel wall by up to 70%, depending on the dose. These observations suggest that gliotoxin favorably regulates the response to vascular injury through actions on VSMC. However, further studies evaluating the therapeutic benefit of gliotoxin in restenosis after balloon angioplasty are required.


Subject(s)
Cardiovascular Agents/pharmacology , Carotid Artery Injuries/prevention & control , Cell Movement/drug effects , Cell Proliferation/drug effects , Endothelium, Vascular/drug effects , Gliotoxin/pharmacology , Muscle, Smooth, Vascular/drug effects , Actins/metabolism , Active Transport, Cell Nucleus , Angioplasty, Balloon/adverse effects , Animals , Becaplermin , Carotid Artery Injuries/etiology , Carotid Artery Injuries/metabolism , Carotid Artery Injuries/pathology , Cell Adhesion/drug effects , Cell Adhesion Molecules/metabolism , Cells, Cultured , Disease Models, Animal , Endothelium, Vascular/injuries , Endothelium, Vascular/metabolism , Endothelium, Vascular/pathology , Hyperplasia , Male , Monocytes/metabolism , Muscle, Smooth, Vascular/injuries , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/pathology , Platelet-Derived Growth Factor/metabolism , Proliferating Cell Nuclear Antigen/metabolism , Proto-Oncogene Proteins c-sis , Rats , Rats, Sprague-Dawley , Recurrence , Time Factors , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolism , Vimentin/metabolism
2.
J Am Coll Cardiol ; 50(25): 2399-403, 2007 Dec 18.
Article in English | MEDLINE | ID: mdl-18154965

ABSTRACT

OBJECTIVES: We aimed to study the prevalence of Fabry disease (FD) in patients with hypertrophic cardiomyopathy (HCM). BACKGROUND: There are limited and controversial data about the prevalence of FD in patients with HCM. METHODS: We screened the plasma alpha-galactosidase A activity from 508 unrelated patients with HCM (328 men, 180 women, ages 58 +/- 16 years). Patients with low activity (0% to 30% of the normal control in men, and 0% to 50% in women) underwent genetic study of the GLA gene. RESULTS: We found low plasma activity in 15 patients (3%). Three men had GLA mutations (0.9%): S238N (novel) in 2 and E358del (described) in 1. Two women had described mutations (1.1%): L89P and A143T. Three unrelated men had the D313Y variant previously associated with enzyme pseudo-deficiency. Two women had polymorphisms that did not segregate with the disease in their families. Five women (activity 39% to 47%) had no sequence variants. The familial studies allowed the diagnosis of 14 carriers: 6 women without Fabry manifestations, 3 women with cardiomyopathy, 2 men with renal and cardiac disease, 1 man with microhematuria, 1 woman with first-degree atrioventricular block, and a 32-year-old woman with only renal disease. CONCLUSIONS: By means of a screening based on genotyping of patients with low plasma enzymatic activity, the prevalence of FD in our population of HCM is 1% (0.9% in men and 1.1% in women). This diagnosis is relevant, because it allows the identification of disease carriers that might benefit from enzyme replacement therapy.


Subject(s)
Cardiomyopathy, Hypertrophic/epidemiology , Fabry Disease/epidemiology , Cardiomyopathy, Hypertrophic/diagnosis , Cardiomyopathy, Hypertrophic/genetics , Cohort Studies , Cross-Sectional Studies , DNA Mutational Analysis , Fabry Disease/diagnosis , Fabry Disease/genetics , Female , Genetic Carrier Screening , Genetic Predisposition to Disease/genetics , Genetic Testing , Humans , Male , alpha-Galactosidase/genetics
3.
Methods Mol Biol ; 357: 181-98, 2007.
Article in English | MEDLINE | ID: mdl-17172688

ABSTRACT

Endothelial cells form a continuous monolayer lining the inside face of all blood vessels, and present the ability to selectively control vascular permeability. The endothelium is involved in a wide variety of normal physiological and pathological processes. The endothelial dysfunction occurs under activation conditions, with the acquisition of many new functional, inflammatory, and immune properties, and as a consequence, endothelial cells display many different transcription profiles. We describe here the isolation and culture of the most useful model of human umbilical vein endothelial cells, and undertake the proteomic analysis under both basal quiescent condition and activated by stimulation with a proinflammatory cytokine. Series of two-dimensional electrophoresis have allowed us to detect a total of close to 600 polypeptide spots using 4.0-7.0 pH range in both culture conditions. We have selected 233 proteins by cross-matching the gels, and found that 70% showed an increase and 30% a decrease of expression levels in activated cells. Subsequent identification of 35 altered peptides is made by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry, as well as a study of posttranslational modifications. These global findings may contribute to understand the effects of pathological stimuli and the mechanisms that regulate vascular diseases.


Subject(s)
Endothelial Cells/metabolism , Proteome/analysis , Cells, Cultured , Electrophoresis, Gel, Two-Dimensional , Endothelial Cells/cytology , Endothelial Cells/immunology , Humans , Inflammation Mediators/analysis , Proteome/immunology , Proteomics/methods , Reproducibility of Results , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
4.
Eur J Pharmacol ; 548(1-3): 53-63, 2006 Oct 24.
Article in English | MEDLINE | ID: mdl-16973154

ABSTRACT

Statins are powerful agents for lowering plasma cholesterol levels, which act by inhibition of the 3-hydroxy-3-methylglutaryl-CoA reductase. Evidence suggests that some of the beneficial effects may depend on their anti-inflammatory properties, due to their ability to suppress the synthesis of isoprenoids. The present study analyzes the effects of short-term simvastatin exposure on monocyte migration, cell adhesion, and endothelial cytoskeleton. We demonstrate that simvastatin completely inhibited the migration of THP-1 monocytic cells after 24 h of incubation, being prevented by coincubation with mevalonate (MVA) and geranylgeranylpyrophosphate (GGPP), but not by farnesylpyrophosphate (FPP). Simvastatin decreased chemotaxis to 70% after one hour of incubation; surprisingly neither MVA, GGPP nor FPP were able to restore the effects of the drug. Simvastatin also significantly reduced the adhesion of monocytes to interleukin-1beta (IL-1beta)-activated endothelium to 80% after preincubation for 24 h. This effect was completely reversed by coincubation with MVA and GGPP, and partially with FPP. Unexpectedly, simvastatin increased adhesion molecules expression VCAM-1 and ICAM-1 on cytokine-stimulated endothelial cells. Examination of the actin cytoskeleton on IL-1beta-activated endothelial cells showed that both 4 and 24 h of incubation with simvastatin produced a complete disappearance of F-actin, being completely restored by MVA and partially by GGPP and FPP after 24 h of coincubation. We suggest that cytoskeleton disorganization in endothelial cells is important for inhibiting monocyte adhesion, altering the adhesion molecules function. Taken together, these results strongly support the beneficial anti-inflammatory properties of statins, contributing to the overall clinical effects.


Subject(s)
Anticholesteremic Agents/pharmacology , Endothelial Cells/drug effects , Monocytes/drug effects , Simvastatin/pharmacology , Actins/drug effects , Actins/physiology , Cell Adhesion/drug effects , Cell Line , Cell Movement/drug effects , Chemokine CCL2/pharmacology , Cytoskeleton/drug effects , Endothelial Cells/physiology , Humans , Interleukin-1beta/pharmacology , Mevalonic Acid/pharmacology , Monocytes/physiology , Polyisoprenyl Phosphates/pharmacology , Sesquiterpenes
5.
Atherosclerosis ; 184(1): 53-62, 2006 Jan.
Article in English | MEDLINE | ID: mdl-15922347

ABSTRACT

Triterpenoids are natural compounds that are found in a large variety of plants and vegetarian foods, and are used for medicinal purposes in many Asian countries. Pentacyclic triterpenes, such as ursolic acid, have been reported to exhibit anticancer and anti-inflammatory properties. The present study was designed to assess the effects of ursolic acid in the migration and proliferation of vascular smooth muscle cells (VSMC), and in a vascular injury model. The exposure of VSMC to ursolic acid results in a chemotaxis inhibition, in a reduction of the expression of proliferating cell nuclear antigen (PCNA) and in a disorganization of beta-tubulin and vimentin cytoskeletal proteins. Administration of ursolic acid in the rat carotid balloon catheter injury model shows a significant inhibition of neointimal hyperplasia. Thus, we have demonstrated that daily doses of 6 mg/kg body weight for 10 days reduce both the ratio of intimal to medial areas and the degree of stenosis by 80%, and suppress the expression of PCNA in both neointima and media. These results suggest that pentacyclic triterpenes may be of potential therapeutic value in vascular injury, and a possible treatment strategy for the prevention of the progression of atherosclerosis and restenosis after angioplasty.


Subject(s)
Carotid Artery Injuries/pathology , Carotid Artery, Common , Cyclooxygenase Inhibitors/therapeutic use , Triterpenes/therapeutic use , Tunica Intima/pathology , Animals , Carotid Artery Injuries/drug therapy , Carotid Artery Injuries/metabolism , Cell Proliferation/drug effects , Cells, Cultured , Disease Models, Animal , Follow-Up Studies , Hyperplasia/metabolism , Hyperplasia/pathology , Hyperplasia/prevention & control , Immunohistochemistry , In Vitro Techniques , Male , Muscle, Smooth, Vascular/drug effects , Muscle, Smooth, Vascular/metabolism , Muscle, Smooth, Vascular/pathology , Neovascularization, Pathologic/pathology , Neovascularization, Pathologic/prevention & control , Proliferating Cell Nuclear Antigen/biosynthesis , Proliferating Cell Nuclear Antigen/drug effects , Rats , Rats, Sprague-Dawley , Tubulin/drug effects , Tubulin/metabolism , Tunica Intima/drug effects , Tunica Intima/metabolism , Vimentin/drug effects , Vimentin/metabolism , Ursolic Acid
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