Reduction of vascular leakage by imatinib is associated with preserved microcirculatory perfusion and reduced renal injury markers in a rat model of cardiopulmonary bypass.

BACKGROUND: Cardiopulmonary bypass during cardiac surgery leads to impaired microcirculatory perfusion. We hypothesized that vascular leakage is an important contributor to microcirculatory dysfunction. Imatinib, a tyrosine kinase inhibitor, has been shown to reduce vascular leakage in septic mice. We investigated whether prevention of vascular leakage using imatinib preserves microcirculatory perfusion and reduces organ injury markers in a rat model of cardiopulmonary bypass. METHODS: Male Wistar rats underwent cardiopulmonary bypass after treatment with imatinib or vehicle (n=8 per group). Cremaster muscle microcirculatory perfusion and quadriceps microvascular oxygen saturation were measured using intravital microscopy and reflectance spectroscopy. Evans Blue extravasation was determined in separate experiments. Organ injury markers were determined in plasma, intestine, kidney, and lungs. RESULTS: The onset of cardiopulmonary bypass decreased the number of perfused microvessels by 40% in the control group [9.4 (8.6-10.6) to 5.7 (4.8-6.2) per microscope field; P<0.001 vs baseline], whereas this reduction was not seen in the imatinib group. In the control group, the number of perfused capillaries remained low throughout the experiment, whilst perfusion remained normal after imatinib administration. Microvascular oxygen saturation was less impaired after imatinib treatment compared with controls. Imatinib reduced vascular leakage and decreased fluid resuscitation compared with control [3 (3-6) vs 12 ml (7-16); P=0.024]. Plasma neutrophil-gelatinase-associated-lipocalin concentrations were reduced by imatinib. CONCLUSIONS: Prevention of endothelial barrier dysfunction using imatinib preserved microcirculatory perfusion and oxygenation during and after cardiopulmonary bypass. Moreover, imatinib-induced protection of endothelial barrier integrity reduced fluid-resuscitation requirements and attenuated renal and pulmonary injury markers.

Evaluation of Bioenergetic Function in Cerebral Vascular Endothelial Cells.

The integrity of the blood-brain-barrier (BBB) is critical to prevent brain injury. Cerebral vascular endothelial (CVE) cells are one of the cell types that comprise the BBB; these cells have a very high-energy demand, which requires optimal mitochondrial function. In the case of disease or injury, the mitochondrial function in these cells can be altered, resulting in disease or the opening of the BBB. In this manuscript, we introduce a method to measure mitochondrial function in CVE cells by using whole, intact cells and a bioanalyzer. A mito-stress assay is used to challenge the cells that have been perturbed, either physically or chemically, and evaluate their bioenergetic function. Additionally, this method also provides a useful way to screen new therapeutics that have direct effects on mitochondrial function. We have optimized the cell density necessary to yield oxygen consumption rates that allow for the calculation of a variety of mitochondrial parameters, including ATP production, maximal respiration, and spare capacity. We also show the sensitivity of the assay by demonstrating that the introduction of the microRNA, miR-34a, leads to a pronounced and detectable decrease in mitochondrial activity. While the data shown in this paper is optimized for the bEnd.3 cell line, we have also optimized the protocol for primary CVE cells, further suggesting the utility in preclinical and clinical models.

Asiaticoside Inhibits TNF-α-Induced Endothelial Hyperpermeability of Human Aortic Endothelial Cells.

The increase in endothelial permeability often promotes edema formation in various pathological conditions. Tumor necrosis factor-alpha (TNF-α), a pro-atherogenic cytokine, impairs endothelial barrier function and causes endothelial dysfunction in early stage of atherosclerosis. Asiaticoside, one of the triterpenoids derived from Centella asiatica, is known to possess antiinflammatory activity. In order to examine the role of asiaticoside in preserving the endothelial barrier, we assessed its effects on endothelial hyperpermeability and disruption of actin filaments evoked by TNF-α in human aortic endothelial cells (HAEC). TNF-α caused an increase in endothelial permeability to fluorescein isothiocyanate (FITC)-dextran. Asiaticoside pretreatment significantly suppressed TNF-α-induced increased permeability. Asiaticoside also prevented TNF-α-induced actin redistribution by suppressing stress fiber formation. However, the increased F to G actin ratio stimulated by TNF-α was not changed by asiaticoside. Cytochalasin D, an actin depolymerizing agent, was used to correlate the anti-hyperpermeability effect of asiaticoside with actin cytoskeleton. Surprisingly, asiaticoside failed to prevent cytochalasin D-induced increased permeability. These results suggest that asiaticoside protects against the disruption of endothelial barrier and actin rearrangement triggered by TNF-α without a significant change in total actin pool. However, asiaticoside seems to work by other mechanisms to maintain the integrity of endothelial barrier rather than stabilizing the F-actin organization.

Circulatory and Ventilatory Power: Characterization in Patients with Coronary Artery Disease / Potência Circulatória e Ventilatória: Caracterização em Pacientes com Doença Arterial Coronariana

Background: Circulatory power (CP) and ventilatory power (VP) are indices that have been used for the clinical evaluation of patients with heart failure; however, no study has evaluated these indices in patients with coronary artery disease (CAD) without heart failure. Objective: To characterize both indices in patients with CAD compared with healthy controls. Methods: Eighty-seven men [CAD group = 42 subjects and healthy control group (CG) = 45 subjects] aged 40–65 years were included. Cardiopulmonary exercise testing was performed on a treadmill and the following parameters were measured: 1) peak oxygen consumption (VO2), 2) peak heart rate (HR), 3) peak blood pressure (BP), 4) peak rate-pressure product (peak systolic HR x peak BP), 5) peak oxygen pulse (peak VO2/peak HR), 6) oxygen uptake efficiency (OUES), 7) carbon dioxide production efficiency (minute ventilation/carbon dioxide production slope), 8) CP (peak VO2 x peak systolic BP) and 9) VP (peak systolic BP/carbon dioxide production efficiency). Results: The CAD group had significantly lower values for peak VO2 (p < 0.001), peak HR (p < 0.001), peak systolic BP (p < 0.001), peak rate-pressure product (p < 0.001), peak oxygen pulse (p = 0.008), OUES (p < 0.001), CP (p < 0.001), and VP (p < 0.001) and significantly higher values for peak diastolic BP (p = 0.004) and carbon dioxide production efficiency (p < 0.001) compared with CG. Stepwise regression analysis showed that CP was influenced by group (R2 = 0.44, p < 0.001) and VP was influenced by both group and number of vessels with stenosis after treatment (interaction effects: R2 = 0.46, p < 0.001). Conclusion: The indices CP and VP were lower in men with CAD than healthy controls. .

Fundamento: Os índices da Potência Circulatória (PC) e Potência Ventilatória (PV) têm sido utilizados para avaliação clínica de pacientes com insuficiência cardíaca, mas nenhum estudo avaliou esses índices em pacientes com Doença Arterial Coronariana (DAC). Objetivo: Caracterizar ambos os índices em pacientes com DAC comparados a indivíduos saudáveis. Métodos: Oitenta e sete homens [grupo DAC = 42 sujeitos e, grupo controle (GC) = 45 sujeitos] com idade entre 45 e 65 anos foram incluídos. Um Teste de Exercício Cardiopulmonar (TECP) foi realizado em esteira e as seguintes variáveis foram obtidas: 1) consumo de oxigênio (VO2) pico; 2) Frequência Cardíaca (FC) pico; 3) Pressão Arterial (PA) pico; 4) duplo produto pico (PA sistólica pico x FC pico); 5) pulso de oxigênio pico (VO2 pico dividido pela FC pico); 6) eficiência ventilatória para o consumo de oxigênio (OUES); 7) eficiência ventilatória para a produção de dióxido de carbono (VE/VCO2 slope); 8) PC (VO2 pico x PA sistólica pico); e 9) PV (PA sistólica pico dividido pelo VE/VCO2 slope). Resultados: O grupo DAC apresentou valores significativamente menores das seguintes variáveis no pico do exercício: VO2 (p < 0,001), FC (p < 0,001), PA sistólica (p < 0,001), duplo produto (p < 0,001), pulso de oxigênio (p = 0,008), OUES (p < 0,001), PC (p < 0,001) e PV (p < 0,001), e valores significativamente maiores de PA diastólica (p = 0,004) e VE/VCO2 slope (p < 0,001) em relação ao GC. Uma análise de regressão pelo método stepwise mostrou que a PC foi influenciada pelo grupo (R2 = 0,44, p < 0,001) e a PV tanto pelo grupo quanto pelo número de vasos com estenose pós tratamento (efeito de interação: R2 = 0,46, p < 0,001). Conclusion: Os índices da PC e PV foram menores em homens com DAC comparados ao GC, podendo dessa forma ser utilizados na caracterização dessa população. .

Use of medicinal plants by black women: ethnography study in a low-income community / Uso de las plantas medicinales por las mujeres negras: estudio etnográfico en una comunidad de bajos ingresos / Uso de plantas medicinais por mulheres negras: estudo etnográfico em uma comunidade de baixa renda

Objective To explore beliefs, values and practices related to the use of medicinal plants among low-income black families. Method The research method was ethnography and the participant observation process was done in a low-income community in the peripheral area of the City of São Paulo. Twenty black women were interviewed. Results Two cultural sub-themes, I do use medicines that I learned to make with my mother and with religious practitioners to care for diseases and Home medicines are to treat problems that are not serious, and the cultural theme I do use home medicines to treat simple diseases because I always have them at my disposal, they are free and I don’t need a medical prescription represent beliefs, values, and practices related to the use of medicinal plants among low-income black families. Conclusion The development of such practices, which can hide ethnic and social vulnerability, reveals the resilience of low-income black women in the process of confronting problems during the health-illness process. .

Objetivo Explorar las creencias, valores y prácticas sobre el uso de las plantas medicinales entre las familias negras de bajos ingresos. Método El método de investigación fue la etnografía y el proceso de observación participante fue desarrollado en una comunidad de bajos ingresos en las afueras de la Ciudad de São Paulo. Se entrevistó a veinte mujeres negras. Resultados Dos subtemas culturales Uso remedios que aprendí a hacer con mi madre y con los religiosos para cuidar de enfermedades y Remedios caseros se utilizan para resolver problemas que no son graves y el tema cultural Uso remedio casero para resolver enfermedades simples porque tengo todo lo que necesito, es gratuito y no necesita una receta médica simbolizam las prácticas de las mujeres. Conclusión Estas prácticas, que pueden estar enmascarando vulnerabilidades étnicas y sociales, ponen de manifiesto la resiliencia de las mujeres negras de bajos ingresos en el confrontamiento de los problemas del proceso salud-enfermedad. .

Objetivo Explorar crenças, valores e práticas relativas ao uso das plantas medicinais entre famílias negras de baixa renda. Método Pesquisa etnográfica cujo processo de observação participante foi desenvolvido em uma comunidade de baixa renda da periferia da Cidade de São Paulo. Vinte mulheres negras foram entrevistadas. Resultados Dois subtemas culturais, Uso remédios que aprendi a fazer com minha mãe e com os religiosos para cuidar das doenças e Remédios caseiros servem para resolver problemas que não são graves, e o tema cultural Uso remédio caseiro para resolver doenças simples, pois tenho sempre que preciso, é de graça e não precisa de receita médica representam as crenças, valores e práticas relativos ao uso das plantas medicinais entre famílias negras de baixa renda. Conclusão O desenvolvimento dessas práticas, que pode estar mascarando vulnerabilidades étnicas e sociais, revela a resiliência das mulheres negras de baixa renda no enfrentamento dos problemas que encontram no processo saúde-enfermidade. .

Delphinidin-3-glucoside protects human umbilical vein endothelial cells against oxidized low-density lipoprotein-induced injury by autophagy upregulation via the AMPK/SIRT1 signaling pathway.

SCOPE: Oxidized LDL (oxLDL) induced vascular endothelial cell injury is a key event in the pathogenesis of atherosclerosis (AS). In our previous studies, we showed that delphinidin-3-glucoside (Dp), a natural anthocyanin, attenuated oxLDL-induced injury in human umbilical vein endothelial cells (HUVECs), indicating its potential role in preventing AS. However, the involved mechanism is not fully understood. METHODS AND RESULTS: Via methyl thiazolyl tetrazolium and flow cytometry assay, we found that Dp-attenuated oxLDL-induced cell viability decrease and apoptosis in HUVECs. Depending on confocal microscopy, transmission electron microscopy, and Western blot assay, we found that Dp-induced autophagy in HUVECs, whereas suppression of autophagy significantly abolished the protective role of Dp against oxLDL-induced endothelial cell injury. Furthermore, Dp upregulated sirtuin 1 (SIRT1) expression and SIRT1 knockdown notably suppressed Dp-induced autophagy in HUVECs. Dp also increased the expression of phosphorylated adenosine monophosphate-activated protein kinase, while adenosine monophosphate-activated protein kinase (AMPK) knockdown remarkably abolished Dp-induced SIRT1 expression and subsequent autophagy. CONCLUSION: Our data suggested that Dp protected HUVECs against oxLDL-induced injury by inducing autophagy via the adenosine monophosphate-activated protein kinase/SIRT1 signaling pathway. This new finding might shed light to the prevention and therapy of AS.

Injury to the endothelial surface layer induces glomerular hyperfiltration rats with early-stage diabetes.

Glomerular endothelial surface layer (ESL) may play a role in the mechanisms of albuminuria in diabetic nephropathy, which lack evidence in vivo. The effects of high glucose on the passage of albumin across the glomerular ESL were analysed in streptozotocin-induced diabetic Sprague-Dawley rats for 4 weeks. Albuminuria and glomerular mesangial matrix were significantly increased in diabetic rats. The passage of albumin across the ESL, as measured by albumin-colloid gold particle density in the glomerular basement membrane (GBM), was increased significantly in diabetic rats. The thickness of the glomerular ESL, examined indirectly by infusing Intralipid into vessels using an electron microscope, was significantly decreased and the GBM exhibited little change in diabetic rats. In summary, the glomerular ESL may play a role in the pathogenesis of albuminuria in rats with early-stage diabetes.

The effects of hyperbaric air and hyperbaric oxygen on blood-brain barrier integrity in rats.

Hyperbaric oxygen (HBO) treatment yields conflicting results on blood-brain barrier (BBB) integrity under various pathological conditions and the effects of HBO on healthy brain is poorly understood. In this experimental study, the effects of HBO on BBB integrity were investigated in comparison with hyperbaric air (HBA) in intact rats. Four sessions of HBA or HBO were applied to intact rats in 24h. BBB integrity was functionally and structurally evaluated by determining extravasation of Evans blue (EB) dye and horseradish peroxidase (HRP) tracers. In immunohistochemical evaluation, relative staining intensity for occludin, a tight junction (TJ) protein, and aquaporin 4 (AQP4), a water-channel protein, was detected in the barrier type of microvessels of brain by image analysis. BBB permeability to EB dye significantly increased in animals in HBO treatment group compared to those in HBA and control groups (p<0.05). The immunoreactivity of occludin, a tight junction protein, remained essentially unaltered in capillaries of hippocampus in all groups. In animals exposed to HBO, AQP4 immunoreactivity significantly increased in parietal cortex compared to those in HBA and control groups (p<0.01). Ultrastructurally, frequent vesicles containing HRP reaction products were observed in capillary endothelial cells in cerebral cortex and hippocampus of rats subjected to both HBA and HBO. Our results indicate that the HBO administration to intact rats increased BBB permeability to both EB and HRP while HBA increased only HRP extravasation in these animals. The results of this study suggest that HBA also impairs the BBB integrity in intact rats as well as HBO.

Role of Ureaplasma urealyticum in altering the endothelial metal concentration during preeclampsia.

OBJECTIVES: Preeclampsia is a pregnancy specific disorder connected with endothelial cell dysfunction. In vitro stimulation of preeclamptic placental endothelial cell with Ureaplasma urealyticum will help in understanding its relationship with the host cell. Metals and metal-containing compounds are known to play important roles in many biological processes, including metabolic pathway, inflammation and function of proteins. STUDY DESIGN: The variation in expression of various metals was assessed for the first time using FESEM (Field Emission Scanning Electron Microscope) with EDX (Energy Dispersive X-ray spectroscopy) technique in endothelial cells isolated from normotensive and preeclamptic placenta with and without in vitro U. urealyticum stimulation. The results were correlated with the expression of HSP (heat shock protein) 70 in all the 4 endothelial cells. RESULTS: Preeclampsia and U. urealyticum infection alters endothelial cell size, HSP70 expression and metal (sodium, potassium, calcium, iron) concentration. There is a significant increase in the concentration of iron and calcium and decrease in HSP70 expression and endothelial cell size in preeclamptic endothelial cell with U. urealyticum stimulation. CONCLUSION: This work is the first step in the identification of metals pertinent to mollicute infection and lays the foundation for future studies concentrating on characterization of these metal associated or containing molecules. The ionic imbalance observed infers that calcium and iron supplementation should be executed with caution both during preeclampsia and U. urealyticum infection in pregnancy. This study also suggests that the HSP70 mediated protection exhibited in endothelial cell during preeclampsia is lost upon U. urealyticum infection which further contributes to the observed endothelial cell damage.

Efecto del sulodexide sobre la capacidad de relajación y alteraciones estructurales de la arteria aorta en ratas diabéticas por estreptozotocina / Effect of sulodexide on aortic vasodilation capacity and associated morphological changes in rats with streptozotocin-induced diabetes

La disfunción endotelial (DE) se presenta en pacientes con hipercolesterolemia, hipertensión arterial, obesidad y diabetes mellitus. Evidencias sugieren un papel de los glicosaminoglicanos en la DE. Evaluamos el efecto del sulodexide (SLD), un glicosaminoglicano utilizado en el tratamiento de la albuminuria y la enfermedad isquémica en pacientes diabéticos, sobre la relajación arterial y los cambios morfológicos en un modelo experimental de diabetes tipo 1. La diabetes se indujo a ratas Sprague Dawley administrando estreptozotocina (STZ), 60 mg/kg, i.v. Los animales fueron distribuidos en los siguientes grupos: I= control, II= diabéticas, III: control + sulodexide, IV= diabéticas + sulodexide (15 mg/kg/día s.c). A los 3 meses fueron sacrificados, las aortas extraídas para evaluar la relajación vascular inducida por acetilcolina (Ach) y nitroprusiato de sodio en anillos precontraídos con fenilefrina. Fueron evaluadas histológicamente mediante microscopía de luz y coloraciones diversas. El SLD in vitro no modificó la tensión basal de los anillos arteriales en reposo o precontraídos con fenilefrina. La diabetes disminuyó la capacidad de relajación arterial en respuesta a la Ach en un 28,8-35,1% vs control, efecto que fue prevenido por SLD. No se observó diferencia significativa en la relajación inducida por nitroprusiato sódico entre los grupos. El estudio histológico en los animales diabéticos mostró alteraciones estructurales, particularmente en la íntima y la adventicia, cambios que fueron prevenidos por el tratamiento con SLD. Nuestros resultados apoyan la potencial utilidad terapéutica del SLD en el tratamiento de la disfunción endotelial.

Endothelial dysfunction (ED) is observed in patients with hypercholesterolemia, arterial hypertension, obesity and diabetes mellitus. Recent evidences suggest the involvement of glycosaminoglycans(GSG) in ED. We evaluated the effect of sulodexide (SLD), a natural GSG used in albuminuria and ischemic diabetes treatment, on arterial relaxation and vascular morphological changes in a diabetic type I model. Diabetes was induced, in Sprague-Dawley rats by streptozotocine (STZ) administration, 60 mg, iv. Rats were divided into four groups; I: control, II: diabetics, III: control + SLD, IV: diabetics treated with SLD (15 mg/day). After three months, phenylephrine precontracted aortic rings were used to evaluate acetylcholine (ACh) and sodium nitroprusside (NPS) relaxation capacities. Light microscopy of aorta was done with several staining procedures. In vitro, SLD did not change smooth muscle tone in resting or phenylephrine precontracted aortic rings. In diabetic rats, ACh relaxation was 28.8-35.1% lower than in control rats. Diabetic rats treated with SLD showed aortic ACh relaxation similar to control rats. No significative statistical difference was found in endothelium-independent NPS relaxation, between the different groups. Light microscopy histological studies revealed important morphological alterations, particularly in intima and adventitia layers of aortic artery; those changes were dramatically reversed in SLD treated rats. Our experiments support the conclusion that SLD is a potential drug for improving endothelial dysfunction in diabetes.

Sevoflurane preserves the endothelial glycocalyx against ischaemia-reperfusion injury.

BACKGROUND: Healthy vascular endothelium is coated by the glycocalyx, important in multiple endothelial functions, but destroyed by ischaemia-reperfusion. The impact of volatile anaesthetics on this fragile structure has not been investigated. We evaluated the effect of cardiac pre- and post-conditioning with sevoflurane on integrity of the endothelial glycocalyx in conjunction with coronary vascular function. METHODS: Isolated guinea pig hearts perfused with Krebs-Henseleit buffer underwent 20 min stopped-flow ischaemia (37 degrees C), either without or with 1 MAC sevoflurane. This was applied for 15 min before, for 20 min after, or both before and after ischaemia. Transudate was collected for assessing coronary net fluid extravasation and histamine release by mast cells. Coronary release of syndecan-1 and heparan sulphate was measured. In additional experiments with and without continuous sevoflurane, cathepsin B and tryptase beta-like protease activity were measured in effluent. Hearts were perfusion-fixed to visualize the endothelial glycocalyx. RESULTS: Ischaemia led to a significant (P<0.05) increase by 70% in transudate formation during reperfusion only in hearts without sevoflurane. This was accompanied by significant (P<0.05) increases in heparan sulphate (four-fold) and syndecan release (6.5-fold), with electron microscopy revealing massive degradation of glycocalyx. After ischaemia, histamine was released into transudate, and cathepsin B activity increased in effluent (P<0.05). Sevoflurane application attenuated all these changes, except for histamine release. CONCLUSIONS: Sevoflurane protects the endothelial glycocalyx from ischaemia-reperfusion-induced degradation, with both preconditioning and rapid post-conditioning being successful. The mechanism seems to involve attenuation of lysosomal cathepsin B release and to be independent from tissue mast cell degranulation.

Ingestion of native and thermally oxidized polyunsaturated fats acutely increases circulating numbers of endothelial microparticles.

Circulating numbers of endothelial microparticles (EMP) are an index of endothelial injury and dysfunction; and microparticles positive to CD31 antibody increase acutely after cooked, fatty fast-food meals that are rich in saturated fatty acids (SAFA) and lipid oxidation products. The aim of this study was to determine the acute effect of meals rich in SAFA and native and thermally oxidized polyunsaturated vegetable oil on circulating numbers of EMP positive to CD144 antibody, a more specific marker of EMP. Twenty-two apparently healthy subjects received isocaloric meals rich in cream (CR), unheated sunflower oil, or heated sunflower oil in a randomized crossover study design. Circulating numbers of CD144-EMP and plasma lipids and Svedberg unit of flotation (S(f)) greater than 400 triglyceride content were measured before and 1 and 3 hours after the meals. Triglycerides in the plasma S(f) greater than 400 fraction increased significantly (P < .001) after the meals, with a significantly (P < .05) larger increase after the CR meal. Plasma CD144-EMP increased significantly (20%, P < .05) after the unheated sunflower oil and heated sunflower oil meals and did not increase significantly (P = .55) after the CR meal. This response was significantly different among the meals (P = .002) when first-visit fasting plasma glucose was a covariate. In conclusion, these data suggest that ingestion of meals rich in n-6 polyunsaturated vegetable oil irrespective of whether it has been mildly thermally oxidized may acutely alter the state of the vascular endothelium, resulting in increased shedding of CD144-EMP. The physiologic implications of these findings remain to be determined.

[Effect of sulodexide on aortic vasodilation capacity and associated morphological changes in rats with streptozotocin-induced diabetes]. / Efecto del sulodexide sobre la capacidad de relajación y alteraciones estructurales de la arteria aorta en ratas diabéticas por estreptozotocina.

Endothelial dysfunction (ED) is observed in patients with hypercholesterolemia, arterial hypertension, obesity and diabetes mellitus. Recent evidences suggest the involvement of glycosaminoglycans (GSG) in ED. We evaluated the effect of sulodexide (SLD), a natural GSG used in albuminuria and ischemic diabetes treatment, on arterial relaxation and vascular morphological changes in a diabetic type I model. Diabetes was induced, in Sprague-Dawley rats by streptozotocine (STZ) administration, 60 mg, i.v. Rats were divided into four groups; I: control, II: diabetics, III: control + SLD, IV: diabetics treated with SLD (15 mg/day). After three months, phenylephrine precontracted aortic rings were used to evaluate acetylcholine (ACh) and sodium nitroprusside (NPS) relaxation capacities. Light microscopy of aorta was done with several staining procedures. In vitro, SLD did not change smooth muscle tone in resting or phenylephrine precontracted aortic rings. In diabetic rats, ACh relaxation was 28.8-35.1% lower than in control rats. Diabetic rats treated with SLD showed aortic ACh relaxation similar to control rats. No significative statistical difference was found in endothelium-independent NPS relaxation, between the different groups. Light microscopy histological studies revealed important morphological alterations, particularly in intima and adventitia layers of aortic artery; those changes were dramatically reversed in SLD treated rats. Our experiments support the conclusion that SLD is a potential drug for improving endothelial dysfunction in diabetes.

Omega-3 fatty acids and atorvastatin affect connexin 43 expression in the aorta of hereditary hypertriglyceridemic rats.

Statins and omega-3 polyunsaturated fatty acids (n-3 PUFA) reduce cardiovascular disease incidence during hypertriglyceridemia (HTG). To elucidate possible cardioprotective mechanisms, we focused on gap junction protein connexin 43 (Cx43). Its expression is disturbed during atherogenesis, but little information is available on its expression during HTG. Experiments were performed on adult male hereditary HTG (hHTG) rats treated with n-3 PUFA (30 mg/day) and atorvastatin (0.5 mg/100 g body weight per day) for 2 months. Cx43 expression and distribution in the aorta were investigated by using Western blotting and immunolabeling, followed by quantitative analysis. Transmission electronmicroscopy was used to study ultrastructure of endothelial contact sites. In contrast to age-matched Wistar, Cx43 expression in aorta of hHTG rats was significantly higher (p < 0.05), and prominent Cx43 immunospots were seen in tunica media and less in endothelium of hHTG rats. Changes in Cx43 expression were accompanied by local qualitative subcellular alterations of interendothelial connections. Treatment of hHTG rats with n-3 PUFA and atorvastatin markedly lowered Cx43 expression in aorta and modified connexin distribution in endothelium and media (p < 0.05 vs. untreated hHTG). The protective effect of treatment of HTG was observed on the structural integrity of the endothelium and was readily visible at the molecular level. Results indicate the involvement of altered Cx43 expression in vascular pathophysiology during HTG and during HTG treatment.

Effects of focused ultrasound sonodynamic treatment on the rat blood-brain barrier.

BACKGROUND: Ultrasound has recently been applied to the treatment as well as the diagnosis of various pathologies, and its antitumor effects in the treatment of human cancer and experimental models of cancer have been demonstrated. In addition, it is possible that certain photosensitizers will enhance the antitumor effects of ultrasound. However, very few studies have been reported on how the blood-brain barrier is affected by sonodynamic therapy. The purpose of this study was to evaluate disruption of the blood-brain barrier with focused ultrasound with a photosensitizer, for clinical application of sonodynamic therapy to brain tumors. MATERIALS AND METHODS: Rat brains were subjected to focused ultrasound irradiation via a transducer with or without prior intravenous injection of photosensitizer, and lesions were examined histologically by electron microscopy. RESULTS: Electron microscopically, swelling of astroglial processes, denatured cells, protoplasm of endothelial cells, and mitochondria were observed in the center and border of regions of ultrasonic irradiation. There were numerous pinocytotic vesicles in the cytoplasm of the endothelial cells. In addition, disruption of the cytoplasmic membrane of endothelial cells and astroglia was found in these regions. CONCLUSION: These findings suggest that sonodynamic therapy with a photosensitizer affects the blood-brain barrier, and that blood vessel permeability increases not only as a result of destruction of the blood-brain barrier but also by disruption of the cytoplasmic membrane of endothelial cells.

TGF-beta is required for vascular barrier function, endothelial survival and homeostasis of the adult microvasculature.

Pericyte-endothelial cell (EC) interactions are critical to both vascular development and vessel stability. We have previously shown that TGF-beta signaling between EC and mural cells participates in vessel stabilization in vitro. We therefore investigated the role of TGF-beta signaling in maintaining microvessel structure and function in the adult mouse retinal microvasculature. TGF-beta signaling was inhibited by systemic expression of soluble endoglin (sEng) and inhibition was demonstrated by reduced phospho-smad2 in the adult retina. Blockade of TGF-beta signaling led to increased vascular and neural cell apoptosis in the retina, which was associated with decreased retinal function, as measured by electroretinogram (ERG). Perfusion of the inner retinal vasculature was impaired and was accompanied by defective autoregulation and loss of capillary integrity. Fundus angiography and Evans blue permeability assay revealed a breakdown of the blood-retinal-barrier that was characterized by decreased association between the tight junction proteins zo-1 and occludin. Inhibition of TGF-beta signaling in cocultures of EC and 10T1/2 cells corroborated the in vivo findings, with impaired EC barrier function, dissociation of EC from 10T1/2 cells, and endothelial cell death, supporting the role of EC-mesenchymal interactions in TGF-beta signaling. These results implicate constitutive TGF-beta signaling in maintaining the integrity and function of the adult microvasculature and shed light on the potential role of TGF-beta signaling in vasoproliferative and vascular degenerative retinal diseases.

Alumina nanoparticles induce expression of endothelial cell adhesion molecules.

Nanotechnology is a rapidly growing industry that has elicited much concern because of the lack of available toxicity data. Exposure to ultrafine particles may be a risk for the development of vascular diseases due to dysfunction of the vascular endothelium. Increased endothelial adhesiveness is a critical first step in the development of vascular diseases, such as atherosclerosis. The hypothesis that alumina nanoparticles increase inflammatory markers of the endothelium, measured by the induction of adhesion molecules as well as the adhesion of monocytes to the endothelial monolayer, was tested. Following characterization of alumina nanoparticles by transmission electron microscopy (TEM), electron diffraction, and particle size distribution analysis, endothelial cells were exposed to alumina at various concentrations and times. Both porcine pulmonary artery endothelial cells and human umbilical vein endothelial cells showed increased mRNA and protein expression of VCAM-1, ICAM-1, and ELAM-1. Furthermore, human endothelial cells treated with alumina particles showed increased adhesion of activated monocytes. The alumina particles tended to agglomerate at physiological pH in serum-containing media, which led to a range of particle sizes from nano to micron size during treatment conditions. These data show that alumina nanoparticles can elicit a proinflammatory response and thus present a cardiovascular disease risk.

Proteome analysis suggests that mitochondrial dysfunction in stressed endothelial cells is reversed by a soy extract and isolated isoflavones.

Apoptosis is a driving force in atherosclerosis development. A soy extract or a combination of the soy isoflavones genistein and daidzein inhibited apoptosis induced by oxidized LDL in endothelial cells. Proteome analysis revealed that the LDL-induced alterations of numerous proteins were reversed by the extract and the genistein/daidzein mixture but only three protein entities, all functionally linked to mitochondrial dysfunction, were regulated in common by both treatments.

Alterations of cellular bioenergetics in pulmonary artery endothelial cells.

Idiopathic pulmonary arterial hypertension (IPAH) is pathogenetically related to low levels of the vasodilator nitric oxide (NO). Because NO regulates cellular respiration and mitochondrial biogenesis, we hypothesized that abnormalities of bioenergetics may be present in IPAH. Evaluation of pulmonary artery endothelial cells from IPAH and control lungs in vitro revealed that oxygen consumption of IPAH cells was decreased, especially in state 3 respiration with substrates glutamate-malate or succinate, and this decrease paralleled reduction in Complex IV activity and IPAH cellular NO synthesis. IPAH pulmonary artery endothelial cells had decreased mitochondrial dehydrogenase activity and lowered mitochondrial numbers per cell and mitochondrial DNA content, all of which increased after exposure to NO donors. Although IPAH/pulmonary artery endothelial cells' ATP content was similar to control under normoxia, cellular ATP did not change significantly in IPAH cells under hypoxia, whereas ATP decreased 35% in control cells, identifying a greater dependence on cellular respiration for energy in control cells. Evidence that glucose metabolism was subserving the primary role for energy requirements of IPAH cells was provided by the approximately 3-fold greater glycolytic rate of IPAH cells. Positron emission tomography scan with [18F]fluoro-deoxy-D-glucose performed on IPAH patients and healthy controls revealed significantly higher uptake in IPAH lungs as compared with controls, confirming that the glycolytic rate was increased in vivo. Thus, there are substantial changes in bioenergetics of IPAH endothelial cells, which may have consequences for pulmonary hypertensive responses and potentially in development of novel imaging modalities for diagnosis and evaluation of treatment.

Effect of selenium compounds on the damage induced by oxysterol on rat arterial walls.

Wistar rats were fed Se-deficient (0.017 +/- 0.002 mg Se/kg) and Seadequate (0.32 +/- 0.045 Se mg/kg) diets for 12 mo and then were given 5 mg/kg of cholestane-3beta,5alpha,6beta-triol (3-triol), intravenously. Se compounds (Na(2)SeO(3) and ebselen) were supplemented in different doses and times to the Se-deficient rats. Twenty-four hours after 3-triol infusion, the changes in ultrastructures of rat aorta were examined by scanning electron micrography (SEM) and transmission electron micrography (TEM). SEM examinations showed that 3-triol induced diffused injuries on arterial endothelial urfaces of long-term Se-deficient rat, and a large number of holes or craterlike defects were observed. TEM examinations further showed that 3-triol induced swelling, necrosis, and shedding of endothelial cells, which resulted in the destruction of endothelial integrity. Meanwhile, smooth muscle cells proliferated and migrated toward intimae; the breakage of internal elastic lamina benefited the migration of smooth muscle cells. Supplemented with Na(2)SeO(3) (40 microg/kg, 10 d per continuum) and ebselen (20 mg/kg), respectively, exhibited significant protection from damages induced by 3-triol. It seems that protecting mechanisms were different between Na(2)SeO(3) and ebselen. The present investigation gave visual evidence that both injuries induced by cholesterol oxides and the Se nutritional status contributed to the development of atherosclerosis.