RESUMEN
Varicose veins are the most common venous disorder in humans and are characterized by hemodynamic instability due to valvular insufficiency and orthostatic lifestyle factors. It is unclear how changes in biomechanical signals cause aberrant remodeling of the vein wall. Our previous studies suggest that Notch signaling is implicated in varicose vein arterialization. In the arterial system, mechanoresponsive ETS1 is a transcriptional activator of the endothelial Notch, but its involvement in sensing disrupted venous flow and varicose vein formation has not been investigated. Here, we use human varicose veins and cultured human venous endothelial cells to show that disturbed venous shear stress activates ETS1-NOTCH4/DLL4 signaling. Notch components were highly expressed in the neointima, whereas ETS1 was upregulated in all histological layers of varicose veins. In vitro microfluidic flow-based studies demonstrate that even minute changes in venous flow patterns enhance ETS1-NOTCH4/DLL4 signaling. Uniform venous shear stress, albeit an inherently low-flow system, does not induce ETS1 and Notch proteins. ETS1 activation under altered flow was mediated primarily by MEK1/2 and, to a lesser extent, by MEK5 but was independent of p38 MAP kinase. Endothelial cell-specific ETS1 knockdown prevented disturbed flow-induced NOTCH4/DLL4 expression. TK216, an inhibitor of ETS-family, prevented the acquisition of arterial molecular identity and loss of endothelial integrity in cells exposed to the ensuing altered shear stress. We conclude that ETS1 senses blood flow disturbances and may promote venous remodeling by inducing endothelial dysfunction. Targeting ETS1 rather than downstream Notch proteins could be an effective and safe strategy to develop varicose vein therapies.
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Proteína Proto-Oncogénica c-ets-1 , Receptor Notch4 , Transducción de Señal , Várices , Humanos , Proteína Proto-Oncogénica c-ets-1/metabolismo , Proteína Proto-Oncogénica c-ets-1/genética , Receptor Notch4/metabolismo , Várices/metabolismo , Várices/patología , Proteínas de Unión al Calcio/metabolismo , Proteínas de Unión al Calcio/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Células Endoteliales/metabolismo , Endotelio Vascular/metabolismo , Endotelio Vascular/patología , Masculino , Estrés Mecánico , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Femenino , Células Endoteliales de la Vena Umbilical Humana/metabolismoRESUMEN
Background & objectives: Heart failure (HF) is emerging as a major health problem in India. The profile of HF in India is divergent from elsewhere in the world. While cardiologists must equip themselves with the requisite clinical management tools, scientists and health policymakers would need epidemiological data on HF and information on the resources required to meet the challenges ahead. The aim of this study was to identify the lacunae and to suggest recommendations to improve HF research. Methods: We surveyed a multidisciplinary group of HF experts using a two stage process. An email-based survey was conducted using a structured questionnaire, followed by an online discussion. The experts prioritized the major challenges in convergence research in India and inter-rater agreement values were calculated. In addition, they enlisted potential research gaps and barriers in the domains of epidemiology, diagnostics, management and technology and suggested recommendations to overcome those barriers. Results: The experts identified a paucity of data on HF burden, lack of state-of-the-art diagnostic facilities and trained personnel, overt dependence on imported devices/equipment/reagents, lack of interaction/awareness/information among stakeholders and lack of biobanks, as major barriers in HF research. Three fourths of the experts agreed that lack of interaction among stakeholders was the major challenge with the highest inter-rater agreement in both stages (19 out of 25 and 11 out of 17, respectively). The experts recommended the creation of multidisciplinary taskforces dedicated to population sciences, data sciences, technology development and patient management with short-, intermediate- and long-term strategies. Interpretation & conclusions: The study generated a wish list for advances in HF research and management, and proposed recommendations for facilitating convergence research as a way forward to reduce the burden of HF in India.
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Insuficiencia Cardíaca , Humanos , Insuficiencia Cardíaca/diagnóstico , Insuficiencia Cardíaca/epidemiología , Insuficiencia Cardíaca/terapia , India/epidemiología , Encuestas y CuestionariosRESUMEN
BACKGROUND: Cerebral arteriovenous malformations (cAVM) are a significant cause of intracranial hemorrhagic stroke and brain damage. The arteriovenous junctions in AVM nidus are known to have hemodynamic disturbances such as altered shear stress, which could lead to endothelial dysfunction. The molecular mechanisms coupling shear stress and endothelial dysfunction in cAVMs are poorly understood. We speculated that disturbed blood flow in artery-vein junctions activates Notch receptors and promotes endothelial mesenchymal plasticity during cAVM formation. METHODS: We investigated the expression profile of endothelial mesenchymal transition (EndMT) and cell adhesion markers, as well as activated Notch receptors, in 18 human cAVM samples and 15 control brain tissues, by quantitative real-time PCR (qRT-PCR) and immunohistochemical evaluation. Employing a combination of a microfluidic system, qRT-PCR, immunofluorescence, as well as invasion and inhibitor assays, the effects of various shear stress conditions on Notch-induced EndMT and invasive potential of human cerebral microvascular endothelial cells (hCMEC/d3) were analyzed. RESULTS: We found evidence for EndMT and enhanced expression of activated Notch intracellular domain (NICD3 and NICD4) in human AVM nidus samples. The expression of transmembrane adhesion receptor integrin α9/ß1 is significantly reduced in cAVM nidal vessels. Cell-cell adhesion proteins such as VE-cadherin and N-cadherin were differentially expressed in AVM nidus compared with control brain tissues. Using well-characterized hCMECs, we show that altered fluid shear stress steers Notch3 nuclear translocation and promotes SNAI1/2 expression and nuclear localization. Oscillatory flow downregulates integrin α9/ß1 and VE-cadherin expression, while N-cadherin expression and endothelial cell invasiveness are augmented. Gamma-secretase inhibitor RO4929097, and to a lesser level DAPT, prevent the mesenchymal transition and invasiveness of cerebral microvascular endothelial cells exposed to oscillatory fluid flow. CONCLUSIONS: Our study provides, for the first time, evidence for the role of oscillatory shear stress in mediating the EndMT process and dysregulated expression of cell adhesion molecules, especially multifunctional integrin α9/ß1 in human cAVM nidus. Concomitantly, our findings indicate the potential use of small-molecular inhibitors such as RO4929097 in the less-invasive therapeutic management of cAVMs.
Asunto(s)
Células Endoteliales , Malformaciones Arteriovenosas Intracraneales , Humanos , Células Endoteliales/metabolismo , Malformaciones Arteriovenosas Intracraneales/metabolismo , Receptores Notch/metabolismo , Cadherinas/genética , Cadherinas/metabolismo , Transición Epitelial-MesenquimalRESUMEN
Citrus flavonoids particularly quercetin which is abundant in grapefruit, onion, green tea, berries etc. are known to have a protective effect on oxidative stress. Pancreatic ß cells which synthesize and secrete insulin are prone to oxidative stress induced damage because of low cellular antioxidant enzymes. To delineate the effects of quercetin on pancreatic ß cells we evaluated the protective effect of quercetin on TC6 insulinoma cells subjected to oxidative stress induced by tert-butyl-hydrogen-peroxide (TBHP). Quercetin was found to reduce TBHP induced apoptosis and trigger insulin secretion in response to glucose, in a dose-dependent manner. Quercetin treatment increased mitochondrial biogenesis, caused hypertrophy in pancreatic ß cells and activated mTOR signaling with a transient change in mitochondrial membrane potential and AMP/ATP. Activation of mTOR signaling resulted in enhanced insulin secretion in TC6 cells.
Asunto(s)
Flavonoides/farmacología , Células Secretoras de Insulina/efectos de los fármacos , Insulinoma/tratamiento farmacológico , Estrés Oxidativo/fisiología , Quercetina/farmacología , Serina-Treonina Quinasas TOR/metabolismo , Animales , Antioxidantes/farmacología , Apoptosis/efectos de los fármacos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Células Secretoras de Insulina/metabolismo , Insulinoma/metabolismo , Insulinoma/patología , Ratones , Transducción de Señal , terc-Butilhidroperóxido/farmacologíaRESUMEN
Advancements in systems biology have resulted in the development of network pharmacology, leading to a paradigm shift from "one-target, one-drug" to "target-network, multi-component therapeutics". We employ a chimeric approach involving in-vivo assays, gene expression analysis, cheminformatics, and network biology to deduce the regulatory actions of a multi-constituent Ayurvedic concoction, Amalaki Rasayana (AR) in animal models for its effect in pressure-overload cardiac hypertrophy. The proteomics analysis of in-vivo assays for Aorta Constricted and Biologically Aged rat models identify proteins expressed under each condition. Network analysis mapping protein-protein interactions and synergistic actions of AR using multi-component networks reveal drug targets such as ACADM, COX4I1, COX6B1, HBB, MYH14, and SLC25A4, as potential pharmacological co-targets for cardiac hypertrophy. Further, five out of eighteen AR constituents potentially target these proteins. We propose a distinct prospective strategy for the discovery of network pharmacological therapies and repositioning of existing drug molecules for treating pressure-overload cardiac hypertrophy.
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Cardiomegalia/tratamiento farmacológico , Desarrollo de Medicamentos/métodos , Extractos Vegetales/farmacología , Animales , Cardiomegalia/metabolismo , Cromatografía Liquida , Sinergismo Farmacológico , Humanos , Espectrometría de Masas , Modelos Biológicos , Simulación del Acoplamiento Molecular , Farmacología Clínica/métodos , Mapas de Interacción de Proteínas/efectos de los fármacos , Proteómica , Transducción de Señal/efectos de los fármacos , Biología de Sistemas/métodosRESUMEN
Endothelium of blood vessels is continuously exposed to various hemodynamic forces. Flow-mediated epigenetic plasticity regulates vascular endothelial function. Recent studies have highlighted the significant role of mechanosensing-related epigenetics in localized endothelial dysfunction and the regional susceptibility for lesions in vascular diseases. In this article, we review the epigenetic mechanisms such as DNA de/methylation, histone modifications, as well as non-coding RNAs in promoting endothelial dysfunction in major arterial and venous diseases, consequent to hemodynamic alterations. We also discuss the current challenges and future prospects for the use of mechanoepigenetic mediators as biomarkers of early stages of vascular diseases and dysregulated mechanosensing-related epigenetic regulators as therapeutic targets in various vascular diseases.
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Epigénesis Genética , Hemodinámica , Enfermedades Vasculares/genética , Enfermedades Vasculares/metabolismo , Animales , Aterosclerosis/metabolismo , Biomarcadores/metabolismo , Metilación de ADN , Células Endoteliales/patología , Endotelio Vascular/metabolismo , Humanos , Hipertensión Pulmonar/genética , Hipertensión Pulmonar/metabolismo , Ratones , Procesamiento Proteico-PostraduccionalRESUMEN
We examined the hitherto unexplored role of mitochondrial transporters and iron metabolism in advancing metabolic and mitochondrial dysfunction in the heart during long term pressure overload. We also investigated the link between mitochondrial dysfunction and fluctuation in mitochondrial transporters associated with pressure overload cardiac hypertrophy. Left ventricular hypertrophy (LVH) was induced in 3-month-old male Wistar rats by constriction of the aorta using titanium clips. After sacrifice at the end of 6 and 15 months after constriction, tissues from the left ventricle (LV) from all animals were collected for histology, biochemical studies, proteomic and metabolic profiling, and gene and protein expression studies. LV tissues from rats with LVH had a significant decrease in the expression of ABCB7 and mitochondrial oxidative phosphorylation (mt-OXPHOS) enzymes, an increased level of lipid metabolites, decrease in the level of intermediate metabolites of pentose phosphate pathway and elevated levels of cytoplasmic and mitochondrial iron, reactive oxygen species (ROS) and autophagy-related proteins. Knockdown of ABCB7 in H9C2 cells and stimulation with angiotensin II resulted in increased ROS levels, ferritin, and transferrin receptor expression and iron overload in both mitochondria and cytoplasm. A decrease in mRNA and protein levels of mt-OXPHOS specific enzymes, mt-dynamics and autophagy clearance and activation of IGF-1 signaling were also seen in these cells. ABCB7 overexpression rescued all these changes. ABCB7 was found to interact with mitochondrial complexes IV and V. We conclude that in chronic pressure overload, ABCB7 deficiency results in iron overload and mitochondrial dysfunction, contributing to heart failure.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Hipertrofia Ventricular Izquierda/metabolismo , Sobrecarga de Hierro/metabolismo , Mitocondrias Cardíacas/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Transportadoras de Casetes de Unión a ATP/genética , Animales , Autofagia/genética , Cardiomegalia/genética , Cardiomegalia/metabolismo , Cardiomegalia/fisiopatología , Línea Celular , Expresión Génica , Hipertrofia Ventricular Izquierda/genética , Hipertrofia Ventricular Izquierda/fisiopatología , Hierro/metabolismo , Sobrecarga de Hierro/diagnóstico , Masculino , Mitocondrias Cardíacas/genética , Proteínas de Transporte de Membrana Mitocondrial/genética , Miocardio/citología , Miocardio/metabolismo , Presión , Proteómica/métodos , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Mitochondrial dysfunction is widely recognized as a major factor for the progression of cardiac failure. Mitochondrial uptake of metabolic substrates and their utilization for ATP synthesis, electron transport chain activity, reactive oxygen species levels, ion homeostasis, mitochondrial biogenesis, and dynamics as well as levels of reactive oxygen species in the mitochondria are key factors which regulate mitochondrial function in the normal heart. Alterations in these functions contribute to adverse outcomes in heart failure. Iron imbalance and oxidative stress are also major factors for the evolution of cardiac hypertrophy, heart failure, and aging-associated pathological changes in the heart. Mitochondrial ATP-binding cassette (ABC) transporters have a key role in regulating iron metabolism and maintenance of redox status in cells. Deficiency of mitochondrial ABC transporters is associated with an impaired mitochondrial electron transport chain complex activity, iron overload, and increased levels of reactive oxygen species, all of which can result in mitochondrial dysfunction. In this review, we discuss the role of mitochondrial ABC transporters in mitochondrial metabolism and metabolic switch, alterations in the functioning of ABC transporters in heart failure, and mitochondrial ABC transporters as possible targets for therapeutic intervention in cardiac failure.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Insuficiencia Cardíaca/metabolismo , Mitocondrias Cardíacas/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Miocitos Cardíacos/metabolismo , Animales , Cardiomegalia/metabolismo , Cardiomegalia/patología , Proteínas del Complejo de Cadena de Transporte de Electrón/metabolismo , Insuficiencia Cardíaca/fisiopatología , Homeostasis/fisiología , Humanos , Hierro/metabolismo , Dinámicas Mitocondriales/fisiología , ATPasas de Translocación de Protón Mitocondriales/metabolismo , Modelos Animales , Biogénesis de Organelos , Estrés Oxidativo/fisiología , Ratas , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Given the high prevalence of chronic venous diseases (CVD), defining criteria to screen patients who are in need for intervention is attaining primacy. An important clinical criterion for treating CVD is incompetence of larger veins. We have assessed the association of size of afflicted veins with disease severity in patients with CVD to define an acceptable criterion to identify patients who need intervention. Demographic characteristics and risk factors were recorded from 6350 patients. Based on physical examination and venous duplex ultrasound study, patients were classified into clinical severity, etiology, anatomy, and pathophysiology (CEAP) classes and grouped according to the size of the veins which had varicosities. Patients with reflux in smaller veins (vein size <4 mm diameter) were considered as type I and those with varicosities in truncal veins (>4 mm diameter) as type II. Risk ratio was determined by multivariate regression analysis. About 47.67% of patients in this study were found to have CEAP class 3 disease. Compared with varicose veins of large truncal veins, patients with varicosities in smaller superficial veins had 2.85-fold ( p < 0.01) more risk of edema and 5.71-fold ( p < 0.01) higher prevalence of hyperpigmentation. Varicosities in small superficial veins were associated with higher risk of ulceration (odds ratio 3.93, 95% confidence interval 2.51-6.18) compared with truncal vein reflux. Our study reveals that presence of small varicose veins in patients without truncal saphenous reflux involvement is associated with severe manifestations of venous insufficiency such as edema and skin lesions even in the absence of varicosities in truncal saphenous veins.
RESUMEN
We evaluated the cardioprotective effect of Amalaki Rasayana (AR), a rejuvenating Ayurvedic drug prepared from Phyllanthus emblica fruits in the reversal of remodeling changes in pressure overload left ventricular cardiac hypertrophy (LVH) and age-associated cardiac dysfunction in male Wistar rats. Six groups (aging groups) of 3 months old animals were given either AR or ghee and honey (GH) orally; seventh group was untreated. Ascending aorta was constricted using titanium clips in 3 months old rats (N = 24; AC groups) and after 6 months, AR or GH was given for further 12 months to two groups; one group was untreated. Histology, gene and protein expression analysis were done in heart tissues. Chemical composition of AR was analyzed by HPLC, HPTLC and LC-MS. AR intake improved (P < 0.05) cardiac function in aging rats and decreased LVH (P < 0.05) in AC rats as well as increased (P < 0.05) fatigue time in treadmill exercise in both groups. In heart tissues of AR administered rats of both the groups, SERCA2, CaM, Myh11, antioxidant, autophagy, oxidative phosphorylation and TCA cycle proteins were up regulated. ADRB1/2 and pCREB expression were increased; pAMPK, NF-kB were decreased. AR has thus a beneficial effect on myocardial energetics, muscle contractile function and exercise tolerance capacity.
Asunto(s)
Cardiomegalia/tratamiento farmacológico , Cardiomegalia/fisiopatología , Medicina Tradicional , Mitocondrias Cardíacas/metabolismo , Contracción Miocárdica , Extractos Vegetales/uso terapéutico , Envejecimiento/metabolismo , Animales , Aorta/patología , Aorta/fisiopatología , Cardiomegalia/genética , Muerte Celular/efectos de los fármacos , Constricción Patológica , Metabolismo Energético/efectos de los fármacos , Fibrosis , Regulación de la Expresión Génica/efectos de los fármacos , Masculino , Mitocondrias Cardíacas/efectos de los fármacos , Modelos Biológicos , Contracción Miocárdica/efectos de los fármacos , Extractos Vegetales/farmacología , Presión , Ratas WistarRESUMEN
Insulin resistance is associated with endothelial dysfunction and ensuing cardiovascular diseases in type 2 diabetes mellitus (T2DM) patients. ENPP1 is a key modulator of insulin signaling and its polymorphism, K121Q, increases the potency to competitively inhibit insulin receptor binding. We investigated the association of ENPP1 121Q variant with coronary artery disease (CAD) in patients with and without T2DM in South Indian population. Our study was conducted in 913 subjects: 198 patients with CAD, 284 patients in whom T2DM and CAD co-exists, 160 patients with T2DM and no CAD history, and 271 healthy volunteers. Genotyping was performed using PCR-RFLP and PCR-DNA sequencing. Genotype frequency of ENPP1 121Q was higher in disease groups compared to healthy subjects (p < 0.05). T2DM patients who carried polymorphic AC/CC genotypes were at 12.8-fold enhanced risk to CAD (95% CI 4.97-37.18, p < 0.01). Moreover we observed that 121Q, both in heterozygous and homozygous polymorphic states, was a risk factor for CAD without diabetes (OR 4.15, p < 0.01). 121Q variant was associated with T2DM patients with no CAD history too, but the risk was statistically insignificant after multivariate logistic regression analysis (OR 2.32, p > 0.05). We conclude that ENPP1 121Q variant is associated with increased risk for CAD in patients with T2DM in South Indian population. We also report that 121Q variant of ENPP1 was an independent risk factor for CAD irrespective of diabetic milieu. Factors which enhance insulin resistance increase the risk for onset and progression of coronary atherosclerosis irrespective of a diabetic background.
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Enfermedad de la Arteria Coronaria/genética , Diabetes Mellitus Tipo 2/genética , Angiopatías Diabéticas/genética , Predisposición Genética a la Enfermedad , Mutación Missense , Hidrolasas Diéster Fosfóricas/genética , Pirofosfatasas/genética , Adulto , Anciano , Anciano de 80 o más Años , Enfermedad de la Arteria Coronaria/epidemiología , Diabetes Mellitus Tipo 2/epidemiología , Angiopatías Diabéticas/epidemiología , Femenino , Humanos , India/epidemiología , Masculino , Persona de Mediana EdadRESUMEN
Endocardial endothelial cells (EECs), when compared with endothelial cells of arteries and veins, possess higher resistance to apoptosis-inducing anticancer agents. The mechanism of this resistance property is unknown. We have investigated the molecular mechanism, which contributes to increased cell survival capacity in EECs. We explored whether the resistance to apoptosis is associated with the cellular expression of ATP-binding cassette transporters such as P-glycoprotein, MRP-1, and ABCG2. We used primary and immortalized porcine endocardial endothelial cells (PEECs and hTERT PEECs) and compared the results with that in porcine aortic endothelial cells (PAECs), left atrioventricular valve endothelial cells (PVECs), and human umbilical vein endothelial cell line (EA.hy926). FACS and immunoblot analysis revealed a significantly higher expression of ABCG2 in PEECs and hTERT PEECs compared to PAECs, PVECs, and EA.hy926. Using apoptosis-inducing anticancer agents such as doxorubicin and camptothecin, through chromatin condensation assay and immunoblot analysis, we demonstrated a higher resistance to apoptosis in EECs compared to PAECs, PVECs, and EA.hy926. Interestingly, resistance in EECs reversed in presence of ABCG2 specific inhibitor, fumitremorgin C. Our observations suggest that an inherently high expression of ABCG2 in EECs protects them against apoptosis in presence of anticancer agents.
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Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Antineoplásicos/toxicidad , Apoptosis/efectos de los fármacos , Resistencia a Antineoplásicos , Células Endoteliales/efectos de los fármacos , Proteínas de Neoplasias/metabolismo , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/antagonistas & inhibidores , Animales , Línea Celular , Supervivencia Celular/efectos de los fármacos , Endocardio , Células Endoteliales/metabolismo , Células Endoteliales/patología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/patología , Humanos , Indoles/farmacología , Proteínas de Neoplasias/antagonistas & inhibidores , Porcinos , Regulación hacia ArribaRESUMEN
Plasma level of cyclophilin A is a promising marker of vascular disease in patients with type 2 diabetes. Genetic variants in the peptidylprolyl isomerase A gene, encoding human cyclophilin may alter protein synthesis thus affecting its activity, function, and circulating plasma levels. We examined the effect of single-nucleotide polymorphisms (SNPs) within the PPIA gene on plasma levels of cyclophilin A and coupled this with status of vascular disease in patients with and without type 2 diabetes in 212 South Indian subjects. The regulatory region of PPIA gene was sequenced for SNPs. The association of SNPs with known blood markers of type 2 diabetes and coronary artery disease such as HbA1c, low- and high-density lipoproteins, triglycerides, fasting and postprandial blood sugar levels, and cyclophilin A were probed. We identified three SNPs namely, rs6850: A > G; (AG/-) c.*227_*228delAG and (-/T) c.*318_*319insT. Welchs two-sample t test indicated an association of SNP rs6850: A > G, located at the 5' UTR region with increased plasma levels of cyclophilin A in patients with coronary artery disease and with coronary artery disease associated with diabetes. The presence of rs6850: A > G variant was significantly associated with coronary artery disease irrespective of whether the patients had diabetes or not. In silico analysis of the sequence using different tools and matrix libraries did not predict any significant differential binding sites for rs6850: A > G, c.*227_*228delAG and c.*318_*319insT. Our results indicate that the SNP rs6850: A > G is associated with increased risk for elevated plasma levels of cyclophilin A and coronary artery disease in patients with and without type 2 diabetes.
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Enfermedad de la Arteria Coronaria/genética , Ciclofilina A/sangre , Isomerasa de Peptidilprolil/genética , Polimorfismo de Nucleótido Simple , Regiones no Traducidas 5' , Adulto , Enfermedad de la Arteria Coronaria/sangre , Enfermedad de la Arteria Coronaria/complicaciones , Diabetes Mellitus Tipo 2/complicaciones , Femenino , Frecuencia de los Genes , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Insulin-like growth factor-1 (IGF-1) is known to promote proliferation in many cell types including c-kit(pos) cardiac stem cells (CSCs). Downstream signaling pathways of IGF-1 induced CSC proliferation have not been investigated. An important downstream target of IGF-1/Akt-1 signaling is FoxO3a, a key negative regulator of cell-cycle progression. We studied the effect of IGF-1 on proliferation of c-kit(pos) murine CSCs and found that IGF-1-mediated cell proliferation is associated with FoxO3a phosphorylation and inactivation of its transcriptional activity. PI3 inhibitors LY294002 and Wortmannin abolished the effect of IGF-1 on FoxO3a phosphorylation indicating that FoxO3a phosphorylation is mediated by PI3/Akt-1 pathway. In cells with FoxO3a translocation to the cytoplasm, there is decreased expression of cell-cycle inhibitors such as p27(kip1) and p57(kip2) and increased expression of CyclinD1. Our study provides evidence that IGF-1 induced CSC proliferation could be the result of FoxO3a inactivation and its downstream effect on cell-cycle regulators.
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Factores de Transcripción Forkhead/metabolismo , Factor I del Crecimiento Similar a la Insulina/farmacología , Miocardio/citología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Células Madre/metabolismo , Transporte Activo de Núcleo Celular , Androstadienos/farmacología , Animales , Ciclo Celular , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Cromonas/farmacología , Ciclina D1/biosíntesis , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/biosíntesis , Inhibidor p57 de las Quinasas Dependientes de la Ciclina/biosíntesis , Proteína Forkhead Box O3 , Factores de Transcripción Forkhead/biosíntesis , Factores de Transcripción Forkhead/genética , Ratones , Ratones Endogámicos BALB C , Morfolinas/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de las Quinasa Fosfoinosítidos-3 , Fosforilación , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-kit/biosíntesis , Transducción de Señal/efectos de los fármacos , Células Madre/citología , WortmaninaRESUMEN
AIMS/HYPOTHESIS: Cyclophilin A, an immunophilin is secreted from human monocytes activated by high glucose. Given its role as an inflammatory mediator of vascular tissue damage associated with inflammation and oxidative stress, we examined plasma levels of cyclophilin A in normal healthy volunteers and patients with type 2 diabetes (DM), with or without coronary artery disease (CAD). METHODS: Study subjects comprised of 212 patients with DM and CAD,101 patients with diabetes, 122 patients with CAD and 121 normal healthy volunteers. Diabetes was assessed by HbA1c levels while coronary artery disease was established by a positive treadmill test and/or coronary angiography. Plasma cyclophilin A was measured using a cyclophilin A ELISA Kit. Relationship of plasma cyclophilin A levels with blood markers of type 2 diabetes, blood lipid levels and medication for diabetes and coronary artery disease were also explored. RESULTS: Plasma Cyclophilin levels were higher in diabetes patients with or without CAD compared to normal subjects (P < 0.001). Age, fasting blood sugar levels and HbA1C levels were positively associated with increased plasma cyclophilin. Patients using metformin had reduced levels of plasma cyclophilin (p < 0.001).Serum levels of total cholesterol, LDL cholesterol and triglycerides had no significant association with plasma cyclophilin levels. In patients with increased serum CRP levels, plasma cyclophilin A was also elevated (p = 0.016). Prevalence odds for DM, DM + CAD and CAD are higher in those with high cyclophilin values, compared to those with lower values, after adjusting for age and sex, indicating strong association of high cyclophilin values with diabetes and vascular disease. CONCLUSIONS/INTERPRETATIONS: Our study demonstrates that patients with type 2 diabetes have higher circulating levels of cyclophilin A than the normal population. Plasma cyclophilin levels were increased in patients with diabetes and coronary artery disease suggesting a role of this protein in accelerating vascular disease in type 2 diabetes. Considering the evidence that Cyclophilin A is an inflammatory mediator in atherogenesis, the mechanistic role of cyclophilin A in diabetic vascular disease progression deserves detailed investigation.
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Ciclofilina A/sangre , Diabetes Mellitus Tipo 2/sangre , Angiopatías Diabéticas/sangre , Adulto , Biomarcadores/sangre , Diabetes Mellitus Tipo 2/diagnóstico , Diabetes Mellitus Tipo 2/epidemiología , Angiopatías Diabéticas/diagnóstico , Angiopatías Diabéticas/epidemiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Factores de RiesgoRESUMEN
Hyperglycemia is widely recognized to be a potent stimulator of monocyte activity, which is a crucial event in the pathogenesis of atherosclerosis. We analyzed the monocyte proteome for potential markers that would enhance the ability to screen for early inflammatory status in Type 2 diabetes mellitus (T2DM), using proteomic technologies. Monocytic cells (THP-1) were primed with high glucose (HG), their protein profiles were analyzed using 2DE and the downregulated differentially expressed spots were identified using MALDI TOF/MS. We selected five proteins that were secretory in function with the help of bioinformatic programs. A predominantly downregulated protein identified as cyclophilin A (sequence coverage 98%) was further validated by immunoblotting experiments. The cellular mRNA levels of cyclophilin A in various HG-primed cells were studied using qRT-PCR assays and it was observed to decrease in a dose-dependent manner. LC-ESI-MS was used to identify this protein in the conditioned media of HG-primed cells and confirmed by Western blotting as well as ELISA. Cyclophilin A was also detected in the plasma of patients with diabetes. We conclude that cyclophilin A is secreted by monocytes in response to HG. Given the paracrine and autocrine actions of cyclophilin A, the secreted immunophilin could be significant for progression of atherosclerosis in type 2 diabetes. Our study also provides evidence that analysis of monocyte secretome is a viable strategy for identifying candidate plasma markers in diabetes.
Asunto(s)
Ciclofilina A/inmunología , Diabetes Mellitus Tipo 2/inmunología , Glucosa/inmunología , Monocitos/inmunología , Proteoma/inmunología , Línea Celular , Células Cultivadas , Ciclofilina A/genética , Diabetes Mellitus Tipo 2/complicaciones , Diabetes Mellitus Tipo 2/genética , Femenino , Regulación de la Expresión Génica , Humanos , Inflamación/diagnóstico , Inflamación/inmunología , Masculino , Persona de Mediana Edad , Monocitos/metabolismo , Proteoma/genética , Proteómica , ARN Mensajero/genética , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
The pathophysiology of vascular disease in diabetes involves abnormalities in endothelial cells, vascular smooth muscle cells, and monocytes. The metabolic abnormalities that characterize diabetes, such as hyperglycemia, increased free fatty acids, and insulin resistance, each provoke molecular mechanisms that contribute to vascular dysfunction. Several molecules have been identified as risk markers, and have been studied to prevent progression of disease and long-term complications. Markers such as C-reactive protein and monocyte chemoattractant protein-1 are used to assess risk for adverse cardiac events, but elevated levels are possible due to the presence of other risk factors as part of the natural physiological defense mechanism. In this review we discuss potential of cyclophilin-A, a secreted oxidative-stress-induced immunophilin with diverse functions. We present evidence for a significant role of cyclophilin-A in the pathogenesis of atherosclerosis in diabetes, and its potential as a marker for vascular disease in type-2 diabetes.
Asunto(s)
Enfermedades Cardiovasculares/fisiopatología , Ciclofilina A/metabolismo , Diabetes Mellitus Tipo 2/fisiopatología , Biomarcadores/metabolismo , Proteína C-Reactiva/metabolismo , Enfermedades Cardiovasculares/sangre , Enfermedades Cardiovasculares/complicaciones , Ciclofilina A/fisiología , Diabetes Mellitus Tipo 2/sangre , Diabetes Mellitus Tipo 2/complicaciones , Humanos , Inflamación/fisiopatología , Mediadores de Inflamación/sangre , Modelos Cardiovasculares , Monocitos/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factores de RiesgoRESUMEN
Chemotherapy is one of the common treatment modalities for cancer. Some of the antineoplastic drugs have, however, been found to be toxic for vascular endothelium, resulting in complications such as endothelial dysfunction, thromboembolism, heart failure, and cardiomyopathy. In this study, we investigated the cytotoxic effect of widely used antitumor agents doxorubicin, camptothecin, and thapsigargin on primary and immortalized porcine endocardial endothelial cells and compared with the effects of these agents on human umbilical vein endothelial cells, human aortic endothelial cells, and EA.hy926 cells. Our study revealed that endocardial endothelial cells are relatively resistant to apoptosis induced by these drugs. Interestingly, our study indicates that response to antitumor agents greatly differs depending on the site of origin of endothelial cells. Doxorubicin, camptothecin, and thapsigargin induce mitochondrial-dependent cell death following loss of mitochondrial membrane potential (MMP) in vascular endothelial cells, with subsequent increase in sub-G0 population. In endocardial endothelial cells, there was no MMP loss; and only cell cycle arrest either at G1 or S phases was observed when the cells were treated with doxorubicin, camptothecin, and thapsigargin.
Asunto(s)
Antineoplásicos/toxicidad , Apoptosis/efectos de los fármacos , Endocardio/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Animales , Camptotecina/toxicidad , Línea Celular , Relación Dosis-Respuesta a Droga , Doxorrubicina/toxicidad , Endocardio/patología , Células Endoteliales/patología , Puntos de Control de la Fase G1 del Ciclo Celular/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/patología , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Puntos de Control de la Fase S del Ciclo Celular/efectos de los fármacos , Porcinos , Tapsigargina/toxicidadRESUMEN
BACKGROUND: Laboratory measurements are an integral part of epidemiological studies in cardiovascular disease. Standardization and quality assurance is of utmost importance in the context of multicentre studies. METHODS: We evaluated a simple and cost-effective method of quality assurance for measurement of total cholesterol, high density lipoprotein (HDL) cholesterol and triglycerides in a study involving 10 centres. Three methods for quality assessment were used for the study that involved measurement of cholesterol, triglycerides and HDL cholesterol and included internal quality control, external quality control and 10% repeat analysis in addition to a uniform standardized protocol developed for the 10 centres. External quality control material was prepared and circulated by the coordinating laboratory. RESULTS: External quality control material was distributed 20 times during the study. The mean variance index suggested a substantial improvement in the performance of participating laboratories over a period of time for cholesterol and triglycerides. This was also evident in the improvement in per cent technical error as a measure of bias and a higher correlation between replicates of samples analysed in the coordinating laboratory and the participating centres for cholesterol, triglycerides and HDL cholesterol. CONCLUSION: A cost-effective quality assurance model for laboratory measurement using local capacities was developed and implemented in a multicentre epidemiology study. Such a programme would be useful for developing countries where cost-cutting is important.
Asunto(s)
Benchmarking/economía , Pruebas de Química Clínica/economía , Lípidos/sangre , Benchmarking/normas , Pruebas de Química Clínica/normas , Análisis Costo-Beneficio , Estudios Epidemiológicos , Humanos , India , Modelos Teóricos , Desarrollo de Programa , Evaluación de Programas y Proyectos de SaludRESUMEN
Several studies have demonstrated that excess of vitamin D3 is toxic particularly to vascular tissues. A notable pathological feature is arterial calcification. The nature of the toxic metabolite in hypervitaminosis D and the pathogenesis of arterial calcification are not clearly understood. The present study was undertaken to explore whether arterial calcification is a sequel of increased calcium uptake by arterial smooth muscle mediated by up regulation of vitamin D receptor in the cells in response to elevated circulating levels of vitamin D3 in serum. The experimental study was performed in 20 New Zealand white female rabbits aged 6 months. Animals in the test group were injected 10,000 IU of cholecalciferol intramuscularly twice a week for one month. Six control animals were given intra-muscular injections of plain cottonseed oil. Animals were sacrificed and aortas were examined for pathological lesions, 1,25-dihyroxyvitamin D3 (1,25(OH)2 D3) receptor levels and 45Ca uptake in smooth muscle cells. Serum samples collected at intervals were assayed for levels of 25-OH-D3 and calcium. The results showed that in animals given injections of cholecalciferol, serum levels of 25-OH-D3 were elevated. In four of these animals calcification and aneurysmal changes were seen in the aorta. Histological lesions comprised of fragmentation of elastic fibers as well as extensive loss of elastic layers. 1,25(OH)2 D3 receptor levels were up regulated and 45Ca uptake enhanced in aortas of animals which were given excessive vitamin D3. The evidences gathered suggest that excess vitamin D is arteriotoxic and that the vitamin induces arterial calcification through up regulation of 1,25(OH)2D3 receptor and increased calcium uptake in smooth muscle cells of the arteries.