Lack of a relationship between circulating gamma-glutamyltransferase levels and carotid intima media thickness in hypertensive and diabetic patients

Marco Nuti, Paolo Spontoni, Chrysanthos Grigoratos, Giulia Dell'Omo, Alberto Balbarini, Roberto PedrinelliDipartimento Cardio Toracico e Vascolare, Università di Pisa, Pisa, ItalyBackground: By increasing the intracellular prooxidant burden, gamma-glutamyltransferase (GGT) may accelerate atherosclerotic vascular disease. That noxious influence may be reflected by circulating enzyme levels, a correlate of cardiovascular risk factors, and a predictor of incident events. To evaluate this hypothesis, we tested the association between circulating GGT and common carotid intima-media thickness (CIMT), a surrogate index of systemic atherosclerotic involvement, in a large and well-characterized group of patients at risk of cardiovascular disease (CVD).Patients: This study analyzed 548 patients with hypertension and/or diabetes and a widely prevalent history of CVD. Subjects with known hepatic disease and abnormal GGT values were excluded.Methods: CIMT (B-mode ultrasonography) values were the mean of four far-wall measurements at both common carotids. Metabolic syndrome (MetS) was diagnosed according to National Cholesterol Education Program-Adult Treatment Panel III criteria. Due to inherent sex-related differences in GGT levels, the data were analyzed separately in males and females in samples dichotomized by the median.Results: The age-adjusted CIMT values did not differ by GGT levels in males or females. In contrast, the carotid wall was consistently thicker in patients with a history of CVD and MetS independent of age and concurrent GGT values. In both sexes, GGT was associated with key components of the MetS such as triglycerides, fasting plasma glucose, and body mass index.Conclusion: The data collected in this mixed group of hypertensive and/or diabetic patients with widely prevalent history of CVD do not support the concept of a direct pathophysiological link between GGT levels within reference limits and atherosclerotic involvement.Keywords: gamma-glutamyltransferase, carotid intima-media thickness, atherosclerosis, metabolic syndrom

An uncommon clinical condition: chronic thrombosis of the inferior vena cava. A case report and review of literature

The lifetime incidence of deep vein thrombosis (DVT) is approximately 0.1% in general population and even more uncommon in subjects below 40 years of age. Thrombosis of the inferior vena cava (IVC) is an exceptionally rare clinical condition, with etiological factors similar to lower limb DVT. We present a case of post-traumatic chronic obstruction of the IVC in 41 years-old man, caused by a prolonged squatted position, while he was working as a bricklayer. We visited the patient fifteen years after the onset of the first clinical setting showing a severe post thrombotic syndrome, as a consequence of the already diagnosticated thrombosis, involving predominantly the right inferior leg. Thrombophilia screening tests showed patient to be a heterozygous carrier of methylenetetrahydrofolate reductase (MTHFR) gene mutation. Computed tomography (CT) scan confirmed the thrombotic obstruction of the infrahepatic IVC, both common iliac veins, right external and internal iliac veins, with multiple collateral pathways. Because of thrombosis extension, inherited prothrombotic condition and the young age of the patient, we decided to continue life-long oral anticoagulant therapy

Influence of the Occlusion Site

Background: Previous findings suggest that transient myocardial ischemia and reperfusion may elicit changes in the autonomic balance. In this study, a spectral analysis of heart rate variability was used to assess the modifications of sympathovagal balance induced by coronary angioplasty and their relationship with the occlusion site. Methods: We studied 23 patients (17M, 6F, age 58 ± 10 years) with left anterior descending and 19 patients (15M, 4F, age 56 ± 9 years) with right coronary artery stenosis. Spectral analysis of heart rate variability was performed, by autoregressive model, in basal conditions and during each balloon inflation. At least two inflations of 90–120 seconds were performed in each patient. Results: In patients with left anterior descending artery stenosis, the first occlusion induced marked changes in the autonomic balance, which moved toward a sympathetic predominance. The low frequency component of the spectrum and the low-to-high frequency ratio increased from 59 ± 10 normalized units (NU) to 75 ± 10 NU (P < 0.001) and from 2.4 ± 1.4 to 7.3 ± 4.7 (P < 0.001) respectively, while the high frequency component decreased from 30 ± 11 NU to 14 ± 7 NU (P < 0.001). These changes showed a progressive attenuation during repetitive occlusions, and were significantly correlated with the entity of myocardial ischemia assessed by the ST-segment shift measured on the intracoronary electrocardiographic lead. On the contrary, in patients with right coronary artery stenosis the first occlusion was ineffective with regard to the spectral parameters whereas the third occlusion induced a significant increase in the high frequency component (from 31 ± 9 NU to 41 ± 10 NU, P < 0.01) and decrease in the low-to-high frequency ratio (from 2.1 ± 0.9 to 1.3 ± 0.5, P < 0.05) suggesting a vagal activation. The entity of vagal activation was not correlated with the ST-segment shift. Conclusions: Our data indicate that repetitive coronary occlusions induce significant changes in the autonomic balance. The direction and the time course of these changes are related to the occlusion site

Fibrin gel: a new scaffold for cardiovascular applications

Aims: Peripheral blood endothelial progenitor cells (EPC) are promising therapies for irreversible myocardial damage, heart failure and peripheral ischemia disease. Natural biopolymers as fibrin are appealing in tissue engineering, because fibrin is biocompatible and bioresorbable. In vitro studies indicate that fibrin can support the growth migration and proliferation of several cells types. Up to date numerous studies have proved the potential of fibrin based injectable cell delivery systems. No studies are available with fibrin as scaffold for EPC. The goal of this study was to investigate if fibrin is a suitable matrix for EPC culture as compared with fibronectin and if different concentrations of fibrinogen (Fb) and thrombin (Th) can influence fibrin structure and EPC behaviour. Methods: Fibrin (Kedrion S.p.a. Lucca, Italy) was prepared mixing Fb (final 4.5-9-18-36 mg/ml) and Th (final 6-12.5-25-50 U/ml). The scaffolds were maintained for 1 hour at 37?C, 5% CO2 before cell seeding. The ultrastructure of fibrin was investigated by scanning electron microscopy (SEM), cryogenic SEM (CRYO-SEM) and atomic force microscopy (AFM) that allow the hydratating analysis of the sample, to evaluate fibre diameter and density. EPC were obtained from peripheral blood of healthy donors and cultured for 1 week on fibrin at the concentration of 1x106 cell/ml in endothelial growth medium. EPC seeded on fibronectin were used as control. Metabolic cell activity on the different scaffolds was assessed after 7 and 14 days by WST1 while cell viability by confocal microscopy (Calcein AM incorporation). Results: Fibrin polymerization rate ranged between 17 and 68 seconds and increased at higher Fb or Th concentrations. Both AFM and SEM analysis revealed a nanometric fibrous structure, with a decrease in fiber diameter with higher fibrinogen concentrations (4.5 mg/ml: 166?4 nm. vs. 36 mg/ml: 119?3 nm, p<0.005, n=5). Different concentrations of Th didn\u27t affect fibre diameter and density. CRYO-SEM suggested a reticulate structure with mesh-size up to 10?m. WST1 assay showed that EPC metabolic activity was better with lower fibrinogen concentrations (4.5 mg/ml: 0.890?0.134 a.u. vs. 36 mg/ml 0.234?0.046 a.u., p<0.05, n=5), while Th had no significant effect. Calcein staining demonstrated that EPC were viable at 14 days and even organised in cluster. Conclusions: Fibrin combines important properties of an ideal biological scaffold, like the nanometric structure, important for the growth and migration of cells. Fibrin is also an ideal scaffold for EPC but the ratio between fibrinogen and thrombin is important for cell viability

Development of a new technology for 3-D nanostructured scaffolds with potential cardiovascular applications

Aims The in situ release and maintaining of cells to promote revascularization is a new goal of cardiovascular therapy. Endothelial progenitor cells (EPC) may contribute to the process of vascular repair. Medical devices realized according to tissue engineering are composed by a cellular component and by an artificial component, usually made of a biocompatible polymer. Scaffolds may be coated with bio-polymers like fibrin to enhance cell adhesion and growth. Aim of this study was to realize nanocomposite 3D scaffolds composed by a synthetic polymer coated with fibrin to support EPC growth and to promote in vivo angiogenesis. Methods 3D PEtU-PDMS scaffolds were studied in vitro for their biocompatibility (viability and proliferation tests; citokine release). In vivo biocompatibility was studied by intramuscular implant in a rabbit model. The scaffolds were fabricated by spray-phase inversion technique. 25U/mL thrombin was sprayed during the fabrication process. The composite scaffold was then incubated o.n. at 37?C with 18mg/mL fibrinogen. The scaffold morphology was analysed by stereo-microscopy and by scanning electron microscopy (SEM). EPC obtained from peripheral blood were cultured for 1 week on the scaffolds at the concentration of 1x106 cell/ml. Fibronectin coating was used as a control. Cell viability was assessed by confocal laser (Calcein-AM incorporation). To test in vivo angiogenesis, EPC-seeded scaffolds were subcutaneously implanted into the back of rats for 14 days. After harvesting, the scaffolds were examined histologically and immunohistochemically to evaluate inflammatory response and neovascularization. Results In vitro and in vivo biocompatibility data demonstrated absence of any citotoxic effect, immunocompatibility and a slight inflammatory reaction without any sign of encapsulation and implant rejection. Morphological analyses showed an homogeneus fibrin coating of the scaffolds, tightly bound and interconnected to the PEtU-PDMS surface. SEM showed the presence of a well organized layer of fibres in a nm scale (mean diameter ~140nm). Cell viability and phenotype were not affected when EPC were seeded on PEtU-PDMS/fibrin scaffolds. The histological observation of explanted scaffolds revealed a slightly inflammatory response and a significant increased numbers of neovessels in tissues surrounding the EPC-seeded scaffold as compared to the scaffold without cells. Conclusions Our data suggest that PEtU-PDMS/fibrin scaffold obtained with a new spray manufacturing technology can support in vitro EPC growth and promote in vivo neovascularisation. Further studies are currently under way in an ischemic hindlimb rat model

3-D Fibrin Scaffold Improves Stemness of Human Peripheral Blood Endothelial Progenitor Cells

Aims Fibrin is a natural biopolymer appealing for cell-based regenerative therapies, because it can support growth, migration and differentiation of different cell types. Endothelial progenitor cells (EPC) represent a very interesting alternative cell source for mature endothelial cells; the fact that can easily isolated from the peripheral blood, thereby eliminating donor morbidity, makes them ideal in applications in the field of regenerative medicine. We have demonstrated that fibrin can support EPC viability and growth. Aim of this study was to evaluate if fibrin can affect EPC differentiation and stem cell markers expression. Methods Fibrin was prepared mixing commercially available (Kedrion S.p.A. Lucca, Italy) fibrinogen (9 mg/ml) and thrombin (25 U/ml). Clot ultrastructure was investigated by scanning electron microscopy (SEM) and cryogenic SEM (CRYO-SEM) to measure fibre diameter and density. Clot elasticity was evaluated by atomic force microscopy (AFM), measuring the tip-sample force by cantilever displacement. EPC were obtained from peripheral blood and cultured on fibrin at the concentration of 1x106cell/cm2. Fibronectin coating was used as a control. Metabolic activity was assessed after 7 and 14 days by WST1 assay and viability by confocal microscopy (calcein incorporation). The expression of both endothelial (CD31, KDR, vWF, Ve-Cadherin) and stem cell markers (nanog, oct-4) was assessed by flow cytometry, confocal microscopy and Real Time RT-PCR. Results SEM analysis revealed a nanometric fibrous structure, with mean fiber diameter of 165?4 nm and mean density of 95.9?0.2 %. CRYO-SEM suggested a reticulate structure with mesh-size up to 10 ?m. Fibrin clot elasticity was 1.78 MPa, as in literature. WST1 assay showed that fibrin increased EPC metabolic activity as compared to fibronectin (fibrin: 0.606?0.056 a.u. vs. fibronectin: 0.311?0.067). Calcein staining demonstrated that EPC were still viable at 14 days. Flow cytometry showed the expression of endothelial markers (CD31=41.8?8.4%; vWF=32.3?3.0%; KDR=89.3?3.7%; VE-Cadherin=41.2?3.8%), confirmed also by confocal microscopy and Real Time RT-PCR. Interestingly, nanog and oct-4 (embryonic stem cell markers) expression was significantly greater on fibrin (p<0.001) as compared to fibronectin. Conclusions These findings suggest that fibrin it is not only a suitable scaffold for EPC growth and viability but also induces EPC differentiation. The observation that Nanog, known as the most important marker of stemness, is maintained longer than on fibronectin, may offer a surplus value to stem cell-based therapies

Circulating endothelial progenitor cells and large artery structure and function in young subjects with uncomplicated Type 1 Diabetes

<p>Abstract</p> <p>Background</p> <p>Carotid intima-media thickness (IMT), indices of large artery stiffness and measures of endothelium function may be used as markers of early atherosclerosis in type 1 diabetes mellitus (T1DM). The aim of the present study was to compare the indices of large artery structure and function as well as endothelial function and regenerating capacity between adolescents with T1DM and healthy control of similar age. In addition, the associations of different vascular measures with endothelial progenitor cells (EPCs), glyco-metabolic control and serum levels of advanced glycation endproducts (AGEs), soluble receptors for AGEs (sRAGE) and adiponectin were evaluated.</p> <p>Methods</p> <p>Sixteen uncomplicated young T1DM patients (mean age 18 ± 2 years, history of disease 11 ± 5 years, HbA1c 7.7 ± 1.1%) and 26 controls (mean age 19 ± 2 years) were studied. A radiofrequency-based ultrasound system (Esaote MyLab 70) was used to measure carotid IMT and wave speed (WS, index of local stiffness), applanation tonometry (PulsePen) was applied to obtain central pulse pressure (PP) and augmentation index (AIx), and carotid-femoral pulse wave velocity (PWV, Complior) was used as index of aortic stiffness. Peripheral endothelium-dependent vasodilation was determined as reactive hyperemia index (RHI, EndoPAT). Circulating EPCs, glycometabolic profile, AGEs (autofluorescence method), sRAGE and adiponectin were also measured.</p> <p>Results</p> <p>After adjusting for age, sex and blood pressure, T1DM adolescents had significantly higher carotid IMT (456 ± 7 vs. 395 ± 63 μm, p < 0.005), carotid WS (p < 0.005), PWV (p = 0.01), AIx (p < 0.0001) and central PP (p < 0.01) and lower EPCs (p = 0.02) as compared to controls. RHI was reduced only in diabetic patients with HbA1c ≥7.5% (p < 0.05). In the overall population, EPCs were an independent determinant of carotid IMT (together with adiponectin), while fasting plasma glucose was an independent determinant of carotid WS, AIx and central PP.</p> <p>Conclusions</p> <p>Our findings suggest that young subjects with relatively long-lasting T1DM have a generalized preclinical involvement of large artery structure and function, as well as a blunted endothelium regenerating capacity. Hyperglycemia and suboptimal chronic glycemic control seem to deteriorate the functional arterial characteristics, such as large arteries stiffness, wave reflection and peripheral endothelium-dependent vasodilation, whereas an impaired endothelium regenerating capacity and adiponectin levels seem to influence arterial structure.</p

Coronary artery anomalies and their clinical relevance

Coronary artery anomalies (CAAs) represent one of the most confusing topic in cardiology and affect approximately 1% of the general population. Altough some anomalies seem to be only anatomical curiosities, others may sometimes have fatal consequences. This review describes the anatomical characteristics of main CAAs and focuses on the pathophysiological mechanisms by which CAAs may cause a pathological state. The last section describes these therapeutical options of this congenital disorders

Impact of depression on circulating endothelial progenitor cells in patients with acute coronary syndromes

Aims: Depression has been identified as a risk factor for an adverse prognosis and reduced survival in patients with acute coronary syndrome (ACS). The number of endothelial progenitor cells (EPCs) is an independent predictor of clinical outcomes in patients with ACS. The aim was to evaluate the impact of depression on EPC levels in patients with ACS. Methods: Out of 74 ACS patients [23 non-ST-segment elevation myocardial infarction (NSTEMI), 48 STEMI], 36 had a diagnosis of major depressive episode (MDE) according to Diagnostic and Statistical Manual of Mental Disorders 4th edition (DSM-IV) criteria at the time of the inclusion in the study. Control groups were as follows: 15 healthy individuals and 18 patients with current MDE without a history of cardiovascular diseases. EPCs were defined as CD34RCD133RKDRR and evaluated by flow cytometry. All patients underwent standardized cardiological and psychopathological evaluations. Parametric and nonparametric statistical tests were performed wherever appropriate. Results: ACS patients with MDE showed a significant decrease in circulating EPC number compared with ACS patients without MDE (P &lt;0.001). The ACS study population was then subdivided into STEMI and NSTEMI groups, and inside each group again patients with MDE showed a significant decrease in circulating CD34RCD133RKDRR EPCs compared with others (P &lt;0.001). Conclusion: We showed that ACS patients with MDE have a reduced number of circulating CD34RCD133RKDRR cells compared with ACS patients without MDE, suggesting that the presence of MDE reduces the response of bone marrow to acute ischemic events. Considering the reparative role of EPCs in ACS patients, we suppose that patients with MDE might be protected less than patients without MDE

HAS-1 genetic polymorphism in sporadic abdominal aortic aneurysm

The hyaluronan synthase 1 (HAS-1) gene encodes a plasma membrane protein that synthesizes hyaluronan (HA), an extracellular matrix molecule. Accumulating evidence emphasizes the relevance of HA metabolism in an increasing number of processes of clinical interest, including abdominal aortic aneurysm (AAA). The existence of aberrant splicing variants of the HAS-1 gene could partly explain the altered extracellular matrix architecture and influence various biological functions, resulting in progressive arterial wall failure in the development of AAA. In the present study, we assessed the hypothesis that HAS-1 genetic 833A/G polymorphism could be associated with the risk of AAA by performing a case-control association study, involving AAA patients and healthy matched donors