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Background and Purpose: Leaves of Jacaranda decurrens are used in traditional Brazilian medicine to treat metabolic diseases related to increased reactive oxygen species. The present study evaluated the antioxidant and cytotoxic potential of hydroethanolic extract from the leaves of Jacaranda decurrens subsp. symmetrifoliolata. Experimental Approach: Phenolic compounds, flavonoids and saponins were evaluated in an ethanol∶water (80∶20, v/v) extract from the leaves of Jacaranda decurrens subsp. symmetrifoliolata (E-Jds). The antioxidant activity of E-Jds was investigated by assessing the following: 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging activity; protection against 2,2′-azobis (2-amidinopropane) dihydrochloride (AAPH)-induced hemolysis of erythrocytes; in vitro and in vivo malondialdehyde dosage; and the ability to activate antioxidant enzymes. K562 leukemia cells were used for the cytotoxic evaluation of E-Jds and for the assessment of the cell death profile through flow cytometry. Key Results: Phenolic and flavonoid compounds were quantified as 14.38% and 2.15%, respectively, of E-Jds. These phenolic and flavonoid compounds proved to be able to scavenge DPPH free radicals with an IC50 of 9.3±3.3 µg/mL, to protect up to 50% of erythrocytes against AAPH-induced hemolysis and to reduce in vitro and in vivo malondialdehyde levels up to 84% and 22%, respectively. E-Jds also increased glutathione peroxidase enzyme activity, with a concomitant decrease in superoxide dismutase and catalase activity, and exhibited dose-dependent cytotoxic activity on K562 erythroleukemia cells with cell death occurring via both late apoptosis and necrosis. Conclusions: E-Jds exhibits in vitro and in vivo antioxidant potential, which may be the mechanism mediating the metabolic activities reported in folk medicine. Furthermore, the cytotoxic activity identified in this study contributes with the knowledge of antiproliferative activities that have been described in the literature for the genus Jacaranda. [ABSTRACT FROM AUTHOR]

Maternal obesity affects the incidence of cardiovascular disease and diabetes in offspring. Also the use of assisted reproductive technologies (ART) has been associated with cardiovascular deficiencies in offspring. Obese women often suffer from infertility and use ART to achieve a pregnancy, but the combined effects of maternal obesity and ART on cardiovascular health and incidence of diabetes in the offspring is not known. Here, we report the effects of the use of ART within an obesogenic environment, consisting of feeding a western diet (WD) to dams and offspring, on resistance artery function and presence of diabetes biomarkers in juvenile mice offspring. Our results indicate that WD and ART interacted to induce endothelial dysfunction in mesenteric resistance arteries isolated from 7-week-old mice offspring. This was determined by presence of a reduced acetylcholine-induced dilation compared to controls. The arteries from these WD-ART mice also had greater wall cross-sectional areas and wall to lumen ratios indicative of vascular hypertrophic remodeling. Of the diabetes biomarkers measured, only resistin was affected by a WD×ART interaction. Serum resistin was significantly greater in WD-ART offspring compared to controls. Diet and sex effects were observed in other diabetes biomarkers. Our conclusion is that in mice the use of ART within an obesogenic environment interacts to favor the development of endothelial dysfunction in the resistance arteries of juvenile offspring, while having marginal effects on diabetes biomarkers. [ABSTRACT FROM AUTHOR]

Objective: In diabetes, vascular dysfunction is characterized by impaired endothelial function due to increased oxidative stress. Empagliflozin, as a selective sodium-glucose co-transporter 2 inhibitor (SGLT2i), offers a novel approach for the treatment of type 2 diabetes by enhancing urinary glucose excretion. The aim of the present study was to test whether treatment with empagliflozin improves endothelial dysfunction in type I diabetic rats via reduction of glucotoxicity and associated vascular oxidative stress. Methods: Type I diabetes in Wistar rats was induced by an intravenous injection of streptozotocin (60 mg/kg). One week after injection empagliflozin (10 and 30 mg/kg/d) was administered via drinking water for 7 weeks. Vascular function was assessed by isometric tension recording, oxidative stress parameters by chemiluminescence and fluorescence techniques, protein expression by Western blot, mRNA expression by RT-PCR, and islet function by insulin ELISA in serum and immunohistochemical staining of pancreatic tissue. Advanced glycation end products (AGE) signaling was assessed by dot blot analysis and mRNA expression of the AGE-receptor (RAGE). Results: Treatment with empagliflozin reduced blood glucose levels, normalized endothelial function (aortic rings) and reduced oxidative stress in aortic vessels (dihydroethidium staining) and in blood (phorbol ester/zymosan A-stimulated chemiluminescence) of diabetic rats. Additionally, the pro-inflammatory phenotype and glucotoxicity (AGE/RAGE signaling) in diabetic animals was reversed by SGLT2i therapy. Conclusions: Empagliflozin improves hyperglycemia and prevents the development of endothelial dysfunction, reduces oxidative stress and improves the metabolic situation in type 1 diabetic rats. These preclinical observations illustrate the therapeutic potential of this new class of antidiabetic drugs. [ABSTRACT FROM AUTHOR]

Neuropathological symptoms of Alzheimer's disease appear in advances stages, once neuronal damage arises. Nevertheless, recent studies demonstrate that in early asymptomatic stages, ß-amyloid peptide damages the cerebral microvasculature through mechanisms that involve an increase in reactive oxygen species and calcium, which induces necrosis and apoptosis of endothelial cells, leading to cerebrovascular dysfunction. The goal of our work is to study the potential preventive effect of the lipophilic antioxidant coenzyme Q (CoQ) against ß-amyloid-induced damage on human endothelial cells. We analyzed the protective effect of CoQ against Aβ-induced injury in human umbilical vein endothelial cells (HUVECs) using fluorescence and confocal microscopy, biochemical techniques and RMN-based metabolomics. Our results show that CoQ pretreatment of HUVECs delayed Aβ incorporation into the plasma membrane and mitochondria. Moreover, CoQ reduced the influx of extracellular Ca2+, and Ca2+ release from mitochondria due to opening the mitochondrial transition pore after β-amyloid administration, in addition to decreasing O2.− and H2O2 levels. Pretreatment with CoQ also prevented ß-amyloid-induced HUVECs necrosis and apoptosis, restored their ability to proliferate, migrate and form tube-like structures in vitro, which is mirrored by a restoration of the cell metabolic profile to control levels. CoQ protected endothelial cells from Aβ-induced injury at physiological concentrations in human plasma after oral CoQ supplementation and thus could be a promising molecule to protect endothelial cells against amyloid angiopathy. [ABSTRACT FROM AUTHOR]

Background: Angiotensin II (AII) plays a central role in vascular remodeling via oxidative stress. However, the interaction between AII and reduced glutathione (GSH) redox status in cardiovascular remodeling remains unknown. Methods: In vivo: The cuff-induced vascular injury model was applied to Sprague Dawley rats. Then we administered saline or a GSH inhibitor, buthionine sulfoximine (BSO, 30 mmol/L in drinking water) for a week, subsequently administered 4 more weeks by osmotic pump with saline or AII (200 ng/kg/minute) to the rats. In vitro: Incorporation of bromodeoxyuridine (BrdU) was measured to determine DNA synthesis in cultured rat vascular smooth muscle cells (VSMCs). Results: BSO reduced whole blood GSH levels. Systolic blood pressure was increased up to 215±4 mmHg by AII at 4 weeks (p<0.01), which was not affected by BSO. Superoxide production in vascular wall was increased by AII and BSO alone, and was markedly enhanced by AII+BSO. The left ventricular weight to body weight ratio was significantly increased in AII and AII+BSO as compared to controls (2.52±0.08, 2.50±0.09 and 2.10±0.07 mg/g respectively, p<0.05). Surprisingly, the co-treatment of BSO totally abolished these morphological changes. Although the vascular circumferential wall stress was well compensated in AII, significantly increased in AII+BSO. The anti-single-stranded DNA staining revealed increasing apoptotic cells in the neointima of injured arteries in BSO groups. BrdU incorporation in cultured VSMCs with AII was increased dose-dependently. Furthermore it was totally abolished by BSO and was reversed by GSH monoethyl ester. Conclusions: We demonstrated that a vast oxidative stress in impaired GSH redox system totally abolished AII-induced vascular, not cardiac remodeling via enhancement of apoptosis in the neointima and suppression of cell growth in the media. The drastic suppression of remodeling may result in fragile vasculature intolerable to mechanical stress by AII. [ABSTRACT FROM AUTHOR]

Identification of the regulators of vascular inflammation is important if we are to understand the molecular mechanisms leading to atherosclerosis and consequent ischemic heart disease, including acute myocardial infarction. Gene polymorphisms in family with sequence similarity 5, member C (FAM5C) are associated with an increased risk of acute myocardial infarction, but little is known about the function of this gene product in blood vessels. Here, we report that the regulation of the expression and function of FAM5C in endothelial cells. We show here that FAM5C is expressed in endothelial cells in vitro and in vivo. Immunofluorescence microcopy showed localization of FAM5C in the Golgi in cultured human endothelial cells. Immunohistochemistry on serial sections of human coronary artery showed that FAM5C-positive endothelium expressed intercellular adhesion molecule-1 (ICAM-1) or vascular cell adhesion molecule-1 (VCAM-1). In cultured human endothelial cells, the overexpression of FAM5C increased the reactive oxygen species (ROS) production, nuclear factor-κB (NF-κB) activity and the expression of ICAM-1, VCAM-1 and E-selectin mRNAs, resulting in enhanced monocyte adhesion. FAM5C was upregulated in response to inflammatory stimuli, such as TNF-α, in an NF-κB- and JNK-dependent manner. Knockdown of FAM5C by small interfering RNA inhibited the increase in the TNF-α-induced production of ROS, NF-κB activity and expression of these leukocyte adhesion molecule mRNAs, resulting in reduced monocyte adhesion. These results suggest that in endothelial cells, when FAM5C is upregulated in response to inflammatory stimuli, it increases the expression of leukocyte adhesion molecules by increasing ROS production and NF-κB activity. [ABSTRACT FROM AUTHOR]

Exposure to nanoparticles (NPs) may cause vascular effects including endothelial dysfunction and foam cell formation, with oxidative stress and inflammation as supposed central mechanisms. We investigated oxidative stress, endothelial dysfunction and lipid accumulation caused by nano-sized carbon black (CB) exposure in cultured human umbilical vein endothelial cells (HUVECs), THP-1 (monocytes) and THP-1 derived macrophages (THP-1a). The proliferation of HUVECs or co-cultures of HUVECs and THP-1 cells were unaffected by CB exposure, whereas there was increased cytotoxicity, assessed by the LDH and WST-1 assays, especially in THP-1 and THP-1a cells. The CB exposure decreased the glutathione (GSH) content in THP-1 and THP-1a cells, whereas GSH was increased in HUVECs. The reactive oxygen species (ROS) production was increased in all cell types after CB exposure. A reduction of the intracellular GSH concentration by buthionine sulfoximine (BSO) pre-treatment further increased the CB-induced ROS production in THP-1 cells and HUVECs. The expression of adhesion molecules ICAM-1 and VCAM-1, but not adhesion of THP-1 to HUVECs or culture dishes, was elevated by CB exposure, whereas these effects were unaffected by BSO pre-treatment. qRT-PCR showed increased VCAM1 expression, but no change in GCLM and HMOX1 expression in CB-exposed HUVECs. Pre-exposure to CB induced lipid accumulation in THP-1a cells, which was not affected by the presence of the antioxidant N-acetylcysteine. In addition, the concentrations of CB to induce lipid accumulation were lower than the concentrations to promote intracellular ROS production in THP-1a cells. In conclusion, exposure to nano-sized CB induced endothelial dysfunction and foam cell formation, which was not dependent on intracellular ROS production. [ABSTRACT FROM AUTHOR]

Background: Exposure to subclinical levels of lipopolysaccharide (LPS) occurs commonly and is seemingly well tolerated. However, recurrent LPS exposure induces cardiac fibrosis over 2 to 3 months in a murine model, not mediated by the renin-angiotensin system. Subclinical LPS induces cardiac fibrosis by unique mechanisms. Methods: In C57/Bl6 mice, LPS (10 mg/kg) or saline (control) were injected intraperitoneally once a week for 1–4 weeks. Mice showed no signs of distress, change in activity, appetite, or weight loss. Mice were euthanized after 3 days, 1, 2, or 4 weeks to measure cardiac expression of fibrosis-related genes and potential mediators (measured by QRT-PCR), including micro-RNA (miR) and NADPH oxidase (NOX). Collagen fraction area of the left ventricle was measured with picrosirius red staining. Cardiac fibroblasts isolated from adult mouse hearts were incubated with 0, 0.1, 1.0 or 10 ng/ml LPS for 48 hours. Results: Cardiac miR expression profiling demonstrated decreased miR-29c after 3 and 7 days following LPS, which were confirmed by QRT-PCR. The earliest changes in fibrosis-related genes and mediators that occurred 3 days after LPS were increased cardiac expression of TIMP-1 and NOX-2 (but not of NOX-4). This persisted at 1 and 2 weeks, with additional increases in collagen Iα1, collagen IIIα1, MMP2, MMP9, TIMP1, TIMP2, and periostin. There was no change in TGF-β or connective tissue growth factor. Collagen fraction area of the left ventricle increased after 2 and 4 weeks of LPS. LPS decreased miR-29c and increased NOX-2 in isolated cardiac fibroblasts. Conclusions: Recurrent exposure to subclinical LPS induces cardiac fibrosis after 2–4 weeks. Early changes 3 days after LPS were decreased miR-29c and increased NOX2 and TIMP1, which persisted at 1 and 2 weeks, along with widespread activation of fibrosis-related genes. Decreased miR-29c and increased NOX2, which induce cardiac fibrosis in other conditions, may uniquely mediate LPS-induced cardiac fibrosis. [ABSTRACT FROM AUTHOR]

Background: During 2007 and 2008 it is likely that millions of patients in the US received heparin contaminated (CH) with oversulfated chondroitin sulfate, which was associated with anaphylactoid reactions. We tested the hypothesis that CH was associated with serious morbidity, mortality, intensive care unit (ICU) stay and heparin-induced thrombocytopenia following adult cardiac surgery. Methods and Findings: We conducted a single center, retrospective, propensity-matched cohort study during the period of CH and the equivalent time frame in the three preceding or the two following years. Perioperative data were obtained from the institutional record of the Society of Thoracic Surgeons National Database, for which the data collection is prospective, standardized and performed by independent investigators. After matching, logistic regression was performed to evaluate the independent effect of CH on the composite adverse outcome (myocardial infarction, stroke, pneumonia, dialysis, cardiac arrest) and on mortality. Cox regression was used to determine the association between CH and ICU length of stay. The 1∶5 matched groups included 220 patients potentially exposed to CH and 918 controls. There were more adverse outcomes in the exposed cohort (20.9% versus 12.0%; difference = 8.9%; 95% CI 3.6% to 15.1%, P<0.001) with an odds ratio for CH of 2.0 (95% CI, 1.4 to 3.0, P<0.001). In the exposed group there was a non-significant increase in mortality (5.9% versus 3.5%, difference = 2.4%; 95% CI, −0.4 to 3.5%, P = 0.1), the median ICU stay was longer by 14.1 hours (interquartile range −26.6 to 79.8, S = 3299, P = 0.0004) with an estimated hazard ratio for CH of 1.2 (95% CI, 1.0 to 1.4, P = 0.04). There was no difference in nadir platelet counts between cohorts. Conclusions: The results from this single center study suggest the possibility that contaminated heparin might have contributed to serious morbidity following cardiac surgery. [ABSTRACT FROM AUTHOR]

It has been clearly established that mitogen-activated protein kinases (MAPKS) are important mediators of angiotensin II (Ang II) signaling via AT1 receptors in the vasculature. However, evidence for a role of these kinases in changes of Ang II-induced vasoconstriction in obesity is still lacking. Here we sought to determine whether vascular MAPKs are differentially activated by Ang II in obese animals. The role of AT2 receptors was also evaluated. Male monosodium glutamate-induced obese (obese) and non-obese Wistar rats (control) were used. The circulating concentrations of Ang I and Ang II, determined by HPLC, were increased in obese rats. Ang II-induced isometric contraction was decreased in endothelium-intact resistance mesenteric arteries from obese compared with control rats and exhibited a retarded AT1 receptor antagonist response. Blocking of AT2 receptors and inhibition of either endothelial nitric oxide synthase (eNOS) or extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) restored Ang II-induced contraction in obese rats. Western blot analysis revealed increased protein expression of AT2 receptors in arteries from obese rats. Basal and Ang II-induced ERK1/2 phosphorylation was also increased in obese rats. Blockade of either AT1 or AT2 receptors corrected the increased ERK1/2 phosphorylation in arteries from obese rats to levels observed in control preparations. Phosphorylation of eNOS was increased in obese rats. Incubation with the ERK1/2 inhibitor before Ang II stimulation did not affect eNOS phosphorylation in control rats; however, it corrected the increased phosphorylation of eNOS in obese rats. These results clearly demonstrate that enhanced AT2 receptor and ERK1/2-induced, NO-mediated vasodilation reduces Ang II-induced contraction in an endothelium-dependent manner in obese rats. [ABSTRACT FROM AUTHOR]

Heart failure provokes alterations in the expression of nucleocytoplasmic transport-related genes. To elucidate the nucleocytoplasmic transport-linked functional network underlying the two major causes of heart failure, ischemic cardiomyopathy (ICM) and dilated cardiomyopathy (DCM), we examined global transcriptome profiles of left ventricular myocardium tissue samples from 31 patients (ICM, n = 10; DCM, n = 13) undergoing heart transplantation and control donors (CNT, n = 8) using RNA-Sequencing and GeneMANIA. Comparative profiling of ICM versus control and DCM versus control showed 1081 and 2440 differentially expressed genes, respectively (>1.29-fold; P<0.05). GeneMANIA revealed differentially regulated functional networks specific to ICM and DCM. In comparison with CNT, differential expression was seen in 9 and 12 nucleocytoplasmic transport-related genes in ICM and DCM groups, respectively. DDX3X, KPNA2, and PTK2B were related to ICM, while SMURF2, NUP153, IPO5, RANBP3, NOXA1, and RHOJ were involved in DCM pathogenesis. Furthermore, the two pathologies shared 6 altered genes: XPO1, ARL4, NFKB2, FHL3, RANBP2, and RHOU showing an identical trend in expression in both ICM and DCM. Notably, the core of the derived functional networks composed of nucleocytoplasmic transport-related genes (XPO1, RANBP2, NUP153, IPO5, KPNA2, and RANBP3) branched into several pathways with downregulated genes. Moreover, we identified genes whose expression levels correlated with left ventricular mass index and left ventricular function parameters in HF patients. Collectively, our study provides a clear distinction between the two pathologies at the transcriptome level and opens up new possibilities to search for appropriate therapeutic targets for ICM and DCM. [ABSTRACT FROM AUTHOR]

Catechol (benzenediol) is present in plant-derived products, such as vegetables, fruits, coffee, tea, wine, areca nut and cigarette smoke. Because platelet dysfunction is a risk factor of cardiovascular diseases, including stroke, atherosclerosis and myocardial infarction, the purpose of this study was to evaluate the anti-platelet and anti-inflammatory effect of catechol and its mechanisms. The effects of catechol on cyclooxygenase (COX) activity, arachidonic acid (AA)-induced aggregation, thromboxane B2 (TXB2) production, lactate dehydrogenase (LDH) release, reactive oxygen species (ROS) production and extracellular signal-regulated kinase (ERK)/p38 phosphorylation were determined in rabbit platelets. In addition, its effect on IL-1β-induced prostaglandin E2 (PGE2) production by fibroblasts was determined. The ex vivo effect of catechol on platelet aggregation was also measured. Catechol (5-25 µM) suppressed AA-induced platelet aggregation and inhibited TXB2 production at concentrations of 0.5–5 µM; however, it showed little cytotoxicity and did not alter U46619-induced platelet aggregation. Catechol (10–50 µM) suppressed COX-1 activity by 29–44% and COX-2 activity by 29–50%. It also inhibited IL-1β-induced PGE2 production, but not COX-2 expression of fibroblasts. Moreover, catechol (1–10 µM) attenuated AA-induced ROS production in platelets and phorbol myristate acetate (PMA)-induced ROS production in human polymorphonuclear leukocytes. Exposure of platelets to catechol decreased AA-induced ERK and p38 phosphorylation. Finally, intravenous administration of catechol (2.5–5 µmole/mouse) attenuated ex vivo AA-induced platelet aggregation. These results suggest that catechol exhibited anti-platelet and anti-inflammatory effects, which were mediated by inhibition of COX, ROS and TXA2 production as well as ERK/p38 phosphorylation. The anti-platelet effect of catechol was confirmed by ex vivo analysis. Exposure to catechol may affect platelet function and thus cardiovascular health. [ABSTRACT FROM AUTHOR]

Aims: The molecular mechanisms of the vasoconstrictor responses evoked by hydrogen peroxide (H2O2) have not been clearly elucidated in skeletal muscle arterioles. Methods and Results: Changes in diameter of isolated, cannulated and pressurized gracilis muscle arterioles (GAs) of Wistar-Kyoto rats were determined under various test conditions. H2O2 (10–100 µM) evoked concentration-dependent constrictions in the GAs, which were inhibited by endothelium removal, or by antagonists of phospholipase A (PLA; 100 µM 7,7-dimethyl-(5Z,8Z)-eicosadienoic acid), protein kinase C (PKC; 10 µM chelerythrine), phospholipase C (PLC; 10 µM U-73122), or Src family tyrosine kinase (Src kinase; 1 µM Src Inhibitor-1). Antagonists of thromboxane A2 (TXA2; 1 µM SQ-29548) or the non-specific cyclooxygenase (COX) inhibitor indomethacin (10 µM) converted constrictions to dilations. The COX-1 inhibitor (SC-560, 1 µM) demonstrated a greater reduction in constriction and conversion to dilation than that of COX-2 (celecoxib, 3 µM). H2O2 did not elicit significant changes in arteriolar Ca2+ levels measured with Fura-2. Conclusions: These data suggest that H2O2 activates the endothelial Src kinase/PLC/PKC/PLA pathway, ultimately leading to the synthesis and release of TXA2 by COX-1, thereby increasing the Ca2+ sensitivity of the vascular smooth muscle cells and eliciting constriction in rat skeletal muscle arterioles. [ABSTRACT FROM AUTHOR]

Atrial hypertrophy and fibrosis are essential pathological features of atrial fibrillation. Recently, adiponectin has become a protein of interest due to its beneficial effects on cardiovascular diseases. However, the molecular mechanism of atrial structural remodeling and signaling pathways evoked by adiponectin remain unclear. In the present study, we investigated the cardioprotective effect of globular adiponectin (gAcrp) on angiotensin II-induced atrial hypertrophy and fibrosis in neonatal Sprague-Dawley rat. To further investigate the molecular mechanisms underlying the preventive effect of gAcrp, transfection of cells with siRNA was used to suppress the mRNA expression of adiponectin receptor 1 (AdipoR1) and its downstream adaptor protein APPL1. Non-silencing-Cy-3 labelled siRNA was used to determine transfection efficiency using fluorescence microscopy. The expression of atrial natriuretic peptide and procollagen type1 α-1, hypertrophy marker and fibrosis one, respectively, was detected by real-time PCR. Furthermore, the expression of adenosine monophosphate-activated protein kinase (AMPK), phosphatidylinositol 3-kinase (PI3K) and Akt was detected by western blotting. In addition, nuclear p65 translocation activity was analyzed by EMSA supershift assay. Our results showed that AdipoR1 and the adaptor protein APPL1 mediated the protective effects of gAcrp. In addition, the function of adiponectin and phosphorylation of AMPK were prominently diminished by inhibition of PI3K. Furthermore, nuclear factor-κB (NF-κB) transcription was diminished by the specific inhibition of AMPK. Taken together, AMPK pivotally interacts with NF-κB and PI3K, mediating the cardioprotective effect of adiponectin, and may serve as a therapeutic target for preventing atrial hypertrophy and fibrosis. Our present study suggests that gAcrp could ameliorate AngII-induced cardiac hypertrophy and fibrosis in rat atrial cells, which is mediated by the activation of AMPK signaling pathways. APPL1 and AdipoR1 are the key factors involved in the downstream of gAcrp approach. [ABSTRACT FROM AUTHOR]

Skin is in direct contact with the environment and therefore undergoes aging as a consequence of environmentally induce damage. Wrinkle formation is a striking feature of intrinsic and photo-induced skin aging, which are both associated with oxidative stress and inflammatory response. The present study was undertaken to identify the mechanisms responsible for the anti-wrinkle effects of MLB, and thus, we investigated whether magnesium lithospermate B (MLB) from Salvia miltiorrhiza BUNGE associated with wrinkle formation caused by intrinsic and extrinsic skin aging using Sprague-Dawley rats aged 5 and 20 months and ultraviolet B (UVB)-irradiated human skin fibroblasts cells, respectively. The results obtained showed that the oral administration of MLB significantly upregulated the level of type I procollagen and downregulated the activities and expressions of matrix-metalloproteinases (MMPs) in rat skin. In fibroblasts, MLB suppressed the transactivation of nuclear factor-kB (NF-kB) and activator protein 1(AP-1), which are the two transcription factors responsible for MMP expression, by suppressing oxidative stress and the mitogen activated protein kinase (MAPK) pathway. Our results show that the antioxidant effect of MLB is due to the direct scavenging of reactive oxygen species (ROS) and its inhibitory effects on NF-kB-dependent inflammation genes, such as, cyclooxygenase-2 and inducible nitric oxide synthase. MLB was found to reverse both age- and UVB-related reductions in skin procollagen levels by suppressing the expressions and activities of NF-kB and AP-1-dependent MMPs by modulating ROS generation and the MAPK signaling pathway. We suggest that MLB potentially has anti-wrinkle and anti-skin aging effects. [ABSTRACT FROM AUTHOR]

The study investigated the effect of oxytetracycline (OTC) on the anti-oxidative defense system, the structure (hemolysis rate and morphology) and function (ATP enzyme activity) of human red blood cells (hRBCs) to investigate the possible toxic mechanism of OTC to hRBCs. The experimental results indicate that OTC can cause a decline in the function of the antioxidant defense system of hRBCs, resulting in oxidative stress. OTC can bring about morphological changes to hRBCs, and further leads to hemolysis, when the concentration of OTC is over 8×10−5 M (about 164 µg/ml). At a low OTC concentration, below 4×10−5 M (82 µg/ml), OTC can enhance the activity of ATP enzyme of hRBCs, known as hormesis. However, at a high concentration, above 4×10−5 M (about 82 µg/ml), the ATP enzymatic activity was inhibited, affecting the function of hRBCs. The estalished mechanism of toxicity of OTC to hRBCs can facilitate a deeper understanding of the toxicity of OTC in vivo. [ABSTRACT FROM AUTHOR]

Diabetes is associated with endothelial dysfunction and platelet activation, both of which may contribute to increased cardiovascular risk. The purpose of this study was to characterize circulating platelets in diabetes and clarify their effects on endothelial function. Male Wistar rats were injected with streptozotocin (STZ) to induce diabetes. Each experiment was performed by incubating carotid arterial rings with platelets (1.65×107 cells/mL; 30 min) isolated from STZ or control rats. Thereafter, the vascular function was characterized in isolated carotid arterial rings in organ bath chambers, and each expression and activation of enzymes involved in nitric oxide and oxidative stress levels were analyzed. Endothelium-dependent relaxation induced by acetylcholine was significantly attenuated in carotid arteries treated with platelets isolated from STZ rats. Similarly, treatment with platelets isolated from STZ rats significantly reduced ACh-induced Akt/endothelial NO synthase signaling/NO production and enhanced TXB2 (metabolite of TXA2), while CD61 (platelet marker) and CD62P (activated platelet marker) were increased in carotid arteries treated with platelets isolated from STZ rats. Furthermore, the platelets isolated from STZ rats decreased total eNOS protein and eNOS dimerization, and increased oxidative stress. These data provide direct evidence that circulating platelets isolated from diabetic rats cause dysfunction of the endothelium by decreasing NO production (via Akt/endothelial NO synthase signaling pathway) and increasing TXA2. Moreover, activated platelets disrupt the carotid artery by increasing oxidative stress. [ABSTRACT FROM AUTHOR]

Background and Objectives: Blood-brain barrier (BBB) dysfunction is an integral feature of neurological disorders and involves the action of multiple proinflammatory cytokines on the microvascular endothelial cells lining cerebral capillaries. There is still however, considerable ambiguity throughout the scientific literature regarding the mechanistic role(s) of cytokines in this context, thereby warranting a comprehensive in vitro investigation into how different cytokines may cause dysregulation of adherens and tight junctions leading to BBB permeabilization. Methods: The present study employs human brain microvascular endothelial cells (HBMvECs) to compare/contrast the effects of TNF-α and IL-6 on BBB characteristics ranging from the expression of interendothelial junction proteins (VE-cadherin, occludin and claudin-5) to endothelial monolayer permeability. The contribution of cytokine-induced NADPH oxidase activation to altered barrier phenotype was also investigated. Results: In response to treatment with either TNF-α or IL-6 (0–100 ng/ml, 0–24 hrs), our studies consistently demonstrated significant dose- and time-dependent decreases in the expression of all interendothelial junction proteins examined, in parallel with dose- and time-dependent increases in ROS generation and HBMvEC permeability. Increased expression and co-association of gp91 and p47, pivotal NADPH oxidase subunits, was also observed in response to either cytokine. Finally, cytokine-dependent effects on junctional protein expression, ROS generation and endothelial permeability could all be attenuated to a comparable extent using a range of antioxidant strategies, which included ROS depleting agents (superoxide dismutase, catalase, N-acetylcysteine, apocynin) and targeted NADPH oxidase blockade (gp91 and p47 siRNA, NSC23766). Conclusion: A timely and wide-ranging investigation comparing the permeabilizing actions of TNF-α and IL-6 in HBMvECs is presented, in which we demonstrate how either cytokine can similarly downregulate the expression of interendothelial adherens and tight junction proteins leading to elevation of paracellular permeability. The cytokine-dependent activation of NADPH oxidase leading to ROS generation was also confirmed to be responsible in-part for these events. [ABSTRACT FROM AUTHOR]

Background: Hemodynamic insults at arterial bifurcations are believed to play a critical role in initiating intracranial aneurysms. Recent studies in a rabbit model indicate that aneurysmal damage initiates under specific wall shear stress conditions when smooth muscle cells (SMCs) become pro-inflammatory and produce matrix metalloproteinases (MMPs). The mechanisms leading to SMC activation and MMP production during hemodynamic aneurysm initiation are unknown. The goal is to determine if nitric oxide and/or superoxide induce SMC changes, MMP production and aneurysmal remodeling following hemodynamic insult. Methods: Bilateral common carotid artery ligation was performed on rabbits (n = 19, plus 5 sham operations) to induce aneurysmal damage at the basilar terminus. Ligated animals were treated with the nitric oxide synthase (NOS) inhibitor LNAME (n = 7) or the superoxide scavenger TEMPOL (n = 5) and compared to untreated animals (n = 7). Aneurysm development was assessed histologically 5 days after ligation. Changes in NOS isoforms, peroxynitrite, reactive oxygen species (ROS), MMP-2, MMP-9, and smooth muscle α-actin were analyzed by immunohistochemistry. Results: LNAME attenuated ligation-induced IEL loss, media thinning and bulge formation. In untreated animals, immunofluorescence showed increased endothelial NOS (eNOS) after ligation, but no change in inducible or neuronal NOS. Furthermore, during aneurysm initiation ROS increased in the media, but not the intima, and there was no change in peroxynitrite. In LNAME-treated animals, ROS production did not change. Together, this suggests that eNOS is important for aneurysm initiation but not by producing superoxide. TEMPOL treatment reduced aneurysm development, indicating that the increased medial superoxide is also necessary for aneurysm initiation. LNAME and TEMPOL treatment in ligated animals restored α-actin and decreased MMPs, suggesting that eNOS and superoxide both lead to SMC de-differentiation and MMP production. Conclusion: Aneurysm-inducing hemodynamics lead to increased eNOS and superoxide, which both affect SMC phenotype, increasing MMP production and aneurysmal damage. [ABSTRACT FROM AUTHOR]

Although TAK1 has been implicated in inflammation and oxidative stress, its roles in vascular smooth muscle cells (VSMCs) and in response to vascular injury have not been investigated. The present study aimed to investigate the role of TAK1 in modulating oxidative stress in VSMCs and its involvement in neointima formation after vascular injury. Double immunostaining reveals that vascular injury induces a robust phosphorylation of TAK1 (Thr187) in the medial VSMCs of injured arteries in wildtype mice, but this effect is blocked in CD40-deficient mice. Upregulation of TAK1 in VSMCs is functionally important, as it is critically involved in pro-oxidative and pro-inflammatory effects on VSMCs and eventual neointima formation. In vivo, pharmacological inhibition of TAK1 with 5Z-7-oxozeaenol blocked the injury-induced phosphorylation of both TAK1 (Thr187) and NF-kB/p65 (Ser536), associated with marked inhibition of superoxide production, 3-nitrotyrosine, and MCP-1 in the injured arteries. Cell culture experiments demonstrated that either siRNA knockdown or 5Z-7-oxozeaenol inhibition of TAK1 significantly attenuated NADPH oxidase activation and superoxide production induced by CD40L/CD40 stimulation. Co-immunoprecipitation experiments indicate that blockade of TAK1 disrupted the CD40L-induced complex formation of p22phox with p47phox, p67phox, or Nox4. Blockade of TAK1 also inhibited CD40L-induced NF-kB activation by modulating IKKα/β and NF-kB p65 phosphorylation and this was related to reduced expression of proinflammatory genes (IL-6, MCP-1 and ICAM-1) in VSMCs. Lastly, treatment with 5Z-7-oxozeaenol attenuated neointimal formation in wire-injured femoral arteries. Our findings demonstrate previously uncharacterized roles of TAK1 in vascular oxidative stress and the contribution to neointima formation after vascular injury. [ABSTRACT FROM AUTHOR]

Objectives: Hypertension is one of the major cardiovascular diseases. It affects nearly 1.56 billion people worldwide. The present study is about a particular genetic polymorphism (A1166C), gene expression and protein expression of the angiotensin II type I receptor (AT1R) (SNP ID: rs5186) and its association with essential hypertension in a Northern Indian population. Methods: We analyzed the A1166C polymorphism and expression of AT1R gene in 250 patients with essential hypertension and 250 normal healthy controls. Results: A significant association was found in the AT1R genotypes (AC+CC) with essential hypertension (χ2 = 22.48, p = 0.0001). Individuals with CC genotypes were at 2.4 times higher odds (p = 0.0001) to develop essential hypertension than individuals with AC and AA genotypes. The statistically significant intergenotypic variation in the systolic blood pressure was found higher in the patients with CC (169.4±36.3 mmHg) as compared to that of AA (143.5±28.1 mmHg) and AC (153.9±30.5 mmHg) genotypes (p = 0.0001). We found a significant difference in the average delta-CT value (p = 0.0001) wherein an upregulated gene expression (approximately 16 fold) was observed in case of patients as compared to controls. Furthermore, higher expression of AT1R gene was observed in patients with CC genotype than with AC and AA genotypes. A significant difference (p = 0.0001) in the protein expression of angiotensin II Type 1 receptor was also observed in the plasma of patients (1.49±0.27) as compared to controls (0.80±0.24). Conclusion: Our findings suggest that C allele of A1166C polymorphism in the angiotensin II type 1 receptor gene is associated with essential hypertension and its upregulation could play an important role in essential hypertension. [ABSTRACT FROM AUTHOR]

Recent studies have demonstrated that the anti-diabetic drug, metformin, can exhibit direct antitumoral effects, or can indirectly decrease tumor proliferation by improving insulin sensitivity. Despite these recent advances, the underlying molecular mechanisms involved in decreasing tumor formation are not well understood. In this study, we examined the antiproliferative role and mechanism of action of metformin in MCF-7 cancer cells treated with 10 mM of metformin for 24, 48, and 72 hours. Using BrdU and the MTT assay, it was found that metformin demonstrated an antiproliferative effect in MCF-7 cells that occurred in a time- and concentration- dependent manner. Flow cytometry was used to analyze markers of cell cycle, apoptosis, necrosis and oxidative stress. Exposure to metformin induced cell cycle arrest in G0-G1 phase and increased cell apoptosis and necrosis, which were associated with increased oxidative stress. Gene and protein expression were determined in MCF-7 cells by real time RT-PCR and western blotting, respectively. In MCF-7 cells metformin decreased the activation of IRβ, Akt and ERK1/2, increased p-AMPK, FOXO3a, p27, Bax and cleaved caspase-3, and decreased phosphorylation of p70S6K and Bcl-2 protein expression. Co-treatment with metformin and H2O2 increased oxidative stress which was associated with reduced cell number. In the presence of metformin, treating with SOD and catalase improved cell viability. Treatment with metformin resulted in an increase in p-p38 MAPK, catalase, MnSOD and Cu/Zn SOD protein expression. These results show that metformin has an antiproliferative effect associated with cell cycle arrest and apoptosis, which is mediated by oxidative stress, as well as AMPK and FOXO3a activation. Our study further reinforces the potential benefit of metformin in cancer treatment and provides novel mechanistic insight into its antiproliferative role. [ABSTRACT FROM AUTHOR]

Ionizing radiation is responsible for oxidative stress by generating reactive oxygen species (ROS), which alters the cellular redox potential. This change activates several redox sensitive enzymes which are crucial in activating signaling pathways at molecular level and can lead to oxidative stress induced inflammation. Therefore, the present study was intended to assess the anti-inflammatory role of ferulic acid (FA), a plant flavonoid, against radiation-induced oxidative stress with a novel mechanistic viewpoint. FA was administered (50 mg/kg body wt) to Swiss albino mice for five consecutive days prior to exposing them to a single dose of 10 Gy 60Co γ-irradiation. The dose of FA was optimized from the survival experiment and 50 mg/kg body wt dose showed optimum effect. FA significantly ameliorated the radiation induced inflammatory response such as phosphorylation of IKKα/β and IκBα and consequent nuclear translocation of nuclear factor kappa B (NF-κB). FA also prevented the increase of cycloxygenase-2 (Cox-2) protein, inducible nitric oxide synthase-2 (iNOS-2) gene expression, lipid peroxidation in liver and the increase of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in serum. It was observed that exposure to radiation results in decreased activity of superoxide dismutase (SOD), catalase (CAT) and the pool of reduced glutathione (GSH) content. However, FA treatment prior to irradiation increased the activities of the same endogenous antioxidants. Thus, pretreatment with FA offers protection against gamma radiation induced inflammation. [ABSTRACT FROM AUTHOR]

Major facts about the development of restenosis include vascular smooth muscle cells (VSMCs) proliferation and migration. A previous study showed that in vitro treatment with magnesium chloride has the potential to affect the proliferation and migration of VSMCs. Magnesium is the major element in deep sea water (DSW) and is a biologically active mineral. It is unclear whether DSW intake can prevent abnormal proliferation and migration of VSMCs as well as balloon angioplasty-induced neointimal hyperplasia. Thus, we attempted to evaluate the anti-restenotic effects of DSW and its possible molecular mechanisms. Several concentrations of DSW, based on the dietary recommendations (RDA) for magnesium, were applied to a model of balloon angioplasty in SD rats. The results showed that DSW intake markedly increased magnesium content within the vascular wall and reduced the development of neointimal hyperplasia. The immunohistochemical analysis also showed that the expression of proteins associated with cell proliferation and migration were decreased in the balloon angioplasty groups with DSW supplement. Furthermore, in vitro treatment with DSW has a dose-dependent inhibitory effect on serum-stimulated proliferation and migration of VSMCs, whose effects might be mediated by modulation of mitogen-activated protein kinase (MAPK) signaling and of the activity of matrix metalloproteinase-2 (MMP-2). Our study suggested that DSW intake can help prevent neointimal hyperplasia (or restenosis), whose effects may be partially regulated by magnesium and other minerals. [ABSTRACT FROM AUTHOR]

Background: Cardiovascular complication due to diabetes has remained a major cause of death. There is an urgent need to intervene the cardiac complications in diabetes by nutritional or pharmacological agents. Thus the present study was designed to find out the effectiveness of garlic on cardiac complications in insulin-resistant diabetic rats. Methods and Results: SD rats were fed high fructose (65%) diet alone or along with raw garlic homogenate (250 mg/kg/day) or nutrient-matched (65% corn starch) control diet for 8 weeks. Fructose-fed diabetic rats showed cardiac hypertrophy, increased NFkB activity and increased oxidative stress. Administration of garlic significantly decreased (p<0.05) cardiac hypertrophy, NFkB activity and oxidative stress. Although we did not observe any changes in myocardial catalase, GSH and GPx in diabetic heart, garlic administration showed significant (p<0.05) increase in all three antioxidant/enzymes levels. Increased endogenous antioxidant enzymes and gene expression in garlic treated diabetic heart are associated with higher protein expression of Nrf2. Increased myocardial H2S levels, activation of PI3K/Akt pathway and decreased Keap levels in fructose-fed heart after garlic administration might be responsible for higher Nrf2 levels. Conclusion: Our study demonstrates that raw garlic homogenate is effective in reducing cardiac hypertrophy and fructose-induced myocardial oxidative stress through PI3K/AKT/Nrf2-Keap1 dependent pathway. [ABSTRACT FROM AUTHOR]

Exposure to microgravity results in post-flight cardiovascular deconditioning and orthostatic intolerance in astronauts. Vascular oxidative stress injury and mitochondrial dysfunction have been indicated in this process. To elucidate the mechanism for this condition, we investigated whether mitochondria regulated NADPH oxidase in hindlimb unweighting (HU) rat cerebral and mesenteric arteries. Four-week HU was used to simulate microgravity in rats. Vascular superoxide generation, protein and mRNA levels of Nox2/Nox4, and the activity of NADPH oxidase were examined in the present study. Compared with control rats, the levels of superoxide increased in cerebral (P<0.001) but not in mesenteric vascular smooth muscle cells. The protein and mRNA levels of Nox2 and Nox4 were upregulated significantly (P<0.001 and P<0.001 for Nox2, respectively; P<0.001 and P<0.001 for Nox4, respectively) in HU rat cerebral arteries but not in mesenteric arteries. NADPH oxidases were activated significantly by HU (P<0.001) in cerebral arteries but not in mesenteric arteries. Chronic treatment with mitochondria-targeted antioxidant mitoTEMPO attenuated superoxide levels (P<0.001), decreased the protein and mRNA expression levels of Nox2/Nox4 (P<0.01 and P<0.05 for Nox2, respectively; P<0.001 and P<0.001 for Nox4, respectively) and the activity of NADPH oxidase (P<0.001) in HU rat cerebral arteries, but exerted no effects on HU rat mesenteric arteries. Therefore, mitochondria regulated the expression and activity of NADPH oxidases during simulated microgravity. Both mitochondria and NADPH oxidase participated in vascular redox status regulation. [ABSTRACT FROM AUTHOR]

Angiotensin II (AngII) type 1 receptor (AT1-R) can be activated by mechanical stress (MS) without the involvement of AngII during the development of cardiomyocyte hypertrophy, in which G protein-independent pathways are critically involved. Although β-arrestin2-biased signaling has been speculated, little is known about how AT1-R/β-arrestin2 leads to ERK1/2 activation. Here, we present a novel mechanism by which Src kinase mediates AT1-R/β-arrestin2-dependent ERK1/2 phosphorylation in response to MS. Differing from stimulation by AngII, MS-triggered ERK1/2 phosphorylation is neither suppressed by overexpression of RGS4 (the negative regulator of the G-protein coupling signal) nor by inhibition of Gαq downstream protein kinase C (PKC) with GF109203X. The release of inositol 1,4,5-triphosphate (IP3) is increased by AngII but not by MS. These results collectively suggest that MS-induced ERK1/2 activation through AT1-R might be independent of G-protein coupling. Moreover, either knockdown of β-arrestin2 or overexpression of a dominant negative mutant of β-arrestin2 prevents MS-induced activation of ERK1/2. We further identifies a relationship between Src, a non-receptor tyrosine kinase and β-arrestin2 using analyses of co-immunoprecipitation and immunofluorescence after MS stimulation. Furthermore, MS-, but not AngII-induced ERK1/2 phosphorylation is attenuated by Src inhibition, which also significantly improves pressure overload-induced cardiac hypertrophy and dysfunction in mice lacking AngII. Finally, MS-induced Src activation and hypertrophic response are abolished by candesartan but not by valsartan whereas AngII-induced responses can be abrogated by both blockers. Our results suggest that Src plays a critical role in MS-induced cardiomyocyte hypertrophy through β-arrestin2-associated angiotensin II type 1 receptor signaling. [ABSTRACT FROM AUTHOR]

Neonatal high-oxygen exposure leads to elevated blood pressure, microvascular rarefaction, vascular dysfunction and arterial (aorta) rigidity in adult rats. Whether structural changes are present in the matrix of aorta wall is unknown. Considering that elastin synthesis peaks in late fetal life in humans, and early postnatal life in rodents, we postulated that transient neonatal high-oxygen exposure can trigger premature vascular remodelling. Sprague Dawley rat pups were exposed from days 3 to 10 after birth to 80% oxygen (vs. room air control) and were studied at 4 weeks. Blood pressure and vasomotor response of the aorta to angiotensin II and to the acetylcholine analogue carbachol were not different between groups. Vascular superoxide anion production was similar between groups. There was no difference between groups in aortic cross sectional area, smooth muscle cell number or media/lumen ratio. In oxygen-exposed rats, aorta elastin/collagen content ratio was significantly decreased, the expression of elastinolytic cathepsin S was increased whereas collagenolytic cathepsin K was decreased. By immunofluorescence we observed an increase in MMP-2 and TIMP-1 staining in aortas of oxygen-exposed rats whereas TIMP-2 staining was reduced, indicating a shift in the balance towards degradation of the extra-cellular matrix and increased deposition of collagen. There was no significant difference in MMP-2 activity between groups as determined by gelatin zymography. Overall, these findings indicate that transient neonatal high oxygen exposure leads to vascular wall alterations (decreased elastin/collagen ratio and a shift in the balance towards increased deposition of collagen) which are associated with increased rigidity. Importantly, these changes are present prior to the elevation of blood pressure and vascular dysfunction in this model, and may therefore be contributory. [ABSTRACT FROM AUTHOR]

The effects of weak magnetic fields on the biological production of reactive oxygen species (ROS) from intracellular superoxide (O2•−) and extracellular hydrogen peroxide (H2O2) were investigated in vitro with rat pulmonary arterial smooth muscle cells (rPASMC). A decrease in O2•− and an increase in H2O2 concentrations were observed in the presence of a 7 MHz radio frequency (RF) at 10 μTRMS and static 45 μT magnetic fields. We propose that O2•− and H2O2 production in some metabolic processes occur through singlet-triplet modulation of semiquinone flavin (FADH•) enzymes and O2•− spin-correlated radical pairs. Spin-radical pair products are modulated by the 7 MHz RF magnetic fields that presumably decouple flavin hyperfine interactions during spin coherence. RF flavin hyperfine decoupling results in an increase of H2O2 singlet state products, which creates cellular oxidative stress and acts as a secondary messenger that affects cellular proliferation. This study demonstrates the interplay between O2•− and H2O2 production when influenced by RF magnetic fields and underscores the subtle effects of low-frequency magnetic fields on oxidative metabolism, ROS signaling, and cellular growth. [ABSTRACT FROM AUTHOR]

Objective: An electron paramagnetic resonance (EPR) technique using the spin probe cyclic hydroxylamine 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CMH) was introduced as a versatile method for high precision quantification of reactive oxygen species, including the superoxide radical in frozen biological samples such as cell suspensions, blood or biopsies. Materials and Methods: Loss of measurement precision and accuracy due to variations in sample size and shape were minimized by assembling the sample in a well-defined volume. Measurement was carried out at low temperature (150 K) using a nitrogen flow Dewar. The signal intensity was measured from the EPR 1st derivative amplitude, and related to a sample, 3-carboxy-proxyl (CP•) with known spin concentration. Results: The absolute spin concentration could be quantified with a precision and accuracy better than ±10 µM (k = 1). The spin concentration of samples stored at −80°C could be reproduced after 6 months of storage well within the same error estimate. Conclusion: The absolute spin concentration in wet biological samples such as biopsies, water solutions and cell cultures could be quantified with higher precision and accuracy than normally achievable using common techniques such as flat cells, tissue cells and various capillary tubes. In addition; biological samples could be collected and stored for future incubation with spin probe, and also further stored up to at least six months before EPR analysis, without loss of signal intensity. This opens for the possibility to store and transport incubated biological samples with known accuracy of the spin concentration over time. [ABSTRACT FROM AUTHOR]

Homeostasis of the intracellular pH (pHi) in mammalian cells plays a pivotal role in maintaining cell function. Thus far, the housekeeping Na+-H+ exchanger (NHE) and the Na+-HCO3− co-transporter (NBC) have been confirmed in many mammalian cells as major acid extruders. However, the role of acid-extruding regulators in human renal artery smooth muscle cells (HRASMCs) remains unclear. It has been demonstrated that lipopolysaccharide (LPS)-induced vascular occlusion is associated with the apoptosis, activating calpain and increased [Ca2+]i that are related to NHE1 activity in endothelia cells. This study determines the acid-extruding mechanisms and the effect of LPS on the resting pHi and active acid extruders in cultured HRASMCs. The mechanism of pHi recovery from intracellular acidosis (induced by NH4Cl-prepulse) is determined using BCECF-fluorescence in cultured HRASMCs. It is seen that (a) the resting pHi is 7.19±0.03 and 7.10±0.02 for HEPES- and CO2/HCO3−- buffered solution, respectively; (b) apart from the housekeeping NHE1, another Na+-coupled HCO3− transporter i.e. NBC, functionally co-exists to achieve acid-equivalent extrusion; (c) three different isoforms of NBC: NBCn1 (SLC4A7; electroneutral), NBCe1 (SLC4A4; electrogenic) and NBCe2 (SLC4A5), are detected in protein/mRNA level; and (d) pHi and NHE protein expression/activity are significantly increased by LPS, in both a dose- and time- dependent manner, but NBCs protein expression is not. In conclusion, it is demonstrated, for the first time, that four pHi acid-extruding regulators: NHE1, NBCn1, NBCe1 and NBCe2, co-exist in cultured HRASMCs. LPS also increases cellular growth, pHi and NHE in a dose- and time-dependent manner. [ABSTRACT FROM AUTHOR]

Objective: We examined the role of syndecan-1 in modulating the phenotype of vascular smooth muscle cells in the context of endogenous inflammatory factors and altered microenvironments that occur in disease or injury-induced vascular remodeling. Methods and Results: Vascular smooth muscle cells (vSMCs) display a continuum of phenotypes that can be altered during vascular remodeling. While the syndecans have emerged as powerful and complex regulators of cell function, their role in controlling vSMC phenotype is unknown. Here, we isolated vSMCs from wild type (WT) and syndecan-1 knockout (S1KO) mice. Gene expression and western blotting studies indicated decreased levels of α-smooth muscle actin (α-SMA), calponin, and other vSMC-specific differentiation markers in S1KO relative to WT cells. The spread area of the S1KO cells was found to be greater than WT cells, with a corresponding increase in focal adhesion formation, Src phosphorylation, and alterations in actin cytoskeletal arrangement. In addition, S1KO led to increased S6RP phosphorylation and decreased AKT and PKC-α phosphorylation. To examine whether these changes were present in vivo, isolated aortae from aged WT and S1KO mice were stained for calponin. Consistent with our in-vitro findings, the WT mice aortae stained higher for calponin relative to S1KO. When exposed to the inflammatory cytokine TNF-α, WT vSMCs had an 80% reduction in syndecan-1 expression. Further, with TNF-α, S1KO vSMCs produced increased pro-inflammatory cytokines relative to WT. Finally, inhibition of interactions between syndecan-1 and integrins αvβ3 and αvβ5 using the inhibitory peptide synstatin appeared to have similar effects on vSMCs as knocking out syndecan-1, with decreased expression of vSMC differentiation markers and increased expression of inflammatory cytokines, receptors, and osteopontin. Conclusions: Taken together, our results support that syndecan-1 promotes vSMC differentiation and quiescence. Thus, the presence of syndecan-1 would have a protective effect against vSMC dedifferentiation and this activity is linked to interactions with integrins αvβ3 and αvβ5. [ABSTRACT FROM AUTHOR]

Angiotensin II (Ang II) type 1 (AT1) receptor is known to mediate a variety of physiological actions of Ang II including autophagy. However, the role of AT1 receptor in cardiomyocyte autophagy triggered by mechanical stress still remains elusive. The aim of this study was therefore to examine whether and how AT1 receptor participates in cardiomyocyte autophagy induced by mechanical stresses. A 48-hour mechanical stretch and a 4-week transverse aorta constriction (TAC) were imposed to cultured cardiomyocytes of neonatal rats and adult male C57B/L6 mice, respectively, to induce cardiomyocyte hypertrophy prior to the assessment of cardiomyocyte autophagy using LC3b-II. Losartan, an AT1 receptor blocker, but not PD123319, the AT2 inhibitor, was found to significantly reduce mechanical stretch-induced LC3b-II upregulation. Moreover, inhibition of p38MAP kinase attenuated not only mechanical stretch-induced cardiomyocyte hypertrophy but also autophagy. To the contrary, inhibition of ERK and JNK suppressed cardiac hypertrophy but not autophagy. Intriguingly, mechanical stretch-induced autophagy was significantly inhibited by Losartan in the absence of Ang II. Taken together, our results indicate that mechanical stress triggers cardiomyocyte autophagy through AT1 receptor-mediated activation of p38MAP kinase independently of Ang II. [ABSTRACT FROM AUTHOR]

Pentachlorophenol (PCP) has been used extensively as a biocide and a wood preservative and has been reported to be immunosuppressive in rodents and humans. Tetrachlorohydroquinone (TCHQ) is a major metabolite of PCP. TCHQ has been identified as the main cause of PCP-induced genotoxicity due to reactive oxidant stress (ROS). However, the precise mechanisms associated with the immunotoxic effects of PCP and TCHQ remain unclear. The aim of this study was to examine the effects of PCP and TCHQ on the induction of ROS and injury to primary mouse splenocytes. Our results shown that TCHQ was more toxic than PCP and that a high dose of TCHQ led to necrotic cell death of the splenocytes through induction of massive and sudden ROS and prolonged ROS-triggered ERK activation. Inhibition of ROS production by N-acetyl-cysteine (NAC) partially restored the mitochondrial membrane potential, inhibited ERK activity, elevated caspase-3 activity and PARP cleavage, and, eventually, switched the TCHQ-induced necrosis to apoptosis. We suggest that prolonged ERK activation is essential for TCHQ-induced necrosis, and that ROS play a pivotal role in the different TCHQ-induced cell death mechanisms. [ABSTRACT FROM AUTHOR]

NADPH oxidase5 (Nox5) is a novel Nox isoform which has recently been recognized as having important roles in the pathogenesis of coronary artery disease, acute myocardial infarction, fetal ventricular septal defect and cancer. The activity of Nox5 and production of reactive oxygen species is regulated by intracellular calcium levels and phosphorylation. However, the kinases that phosphorylate Nox5 remain poorly understood. Previous studies have shown that the phosphorylation of Nox5 is PKC dependent, but this contention was based on the use of pharmacological inhibitors and the isoforms of PKC involved remain unknown. Thus, the major goals of this study were to determine whether PKC can directly regulate Nox5 phosphorylation and activity, to identify which isoforms are involved in the process, and to understand the functional significance of this pathway in disease. We found that a relatively specific PKCα inhibitor, Ro-32-0432, dose-dependently inhibited PMA-induced superoxide production from Nox5. PMA-stimulated Nox5 activity was significantly reduced in cells with genetic silencing of PKCα and PKCε, enhanced by loss of PKCδ and the silencing of PKCθ expression was without effect. A constitutively active form of PKCα robustly increased basal and PMA-stimulated Nox5 activity and promoted the phosphorylation of Nox5 on Ser490, Thr494, and Ser498. In contrast, constitutively active PKCε potently inhibited both basal and PMA-dependent Nox5 activity. Co-IP and in vitro kinase assay experiments demonstrated that PKCα directly binds to Nox5 and modifies Nox5 phosphorylation and activity. Exposure of endothelial cells to high glucose significantly increased PKCα activation, and enhanced Nox5 derived superoxide in a manner that was in prevented by a PKCα inhibitor, Go 6976. In summary, our study reveals that PKCα is the primary isoform mediating the activation of Nox5 and this maybe of significance in our understanding of the vascular complications of diabetes and other diseases with increased ROS production. [ABSTRACT FROM AUTHOR]

Senescence of vascular endothelial cells is an important contributor to the pathogenesis of age-associated vascular disorders such as atherosclerosis. We investigated the effects of antihypertensive agents on high glucose-induced cellular senescence in human umbilical venous endothelial cells (HUVECs). Exposure of HUVECs to high glucose (22 mM) for 3 days increased senescence-associated- β-galactosidase (SA-β-gal) activity, a senescence marker, and decreased telomerase activity, a replicative senescence marker. The calcium channel blocker nifedipine, but not the β1-adrenergic blocking agent atenolol or the angiotensin-converting enzyme inhibitor perindopril, reduced SA-β-gal positive cells and prevented a decrease in telomerase activity in a high-glucose environment. This beneficial effect of nifedipine was associated with reduced reactive oxygen species (ROS) and increased endothelial nitric oxide synthase (eNOS) activity. Thus, nifedipine prevented high glucose-induced ROS generation and increased basal eNOS phosphorylation level at Ser-1177. Treatment with NG-nitro-L-arginine (L-NAME) and transfection of small interfering RNA (siRNA) targeting eNOS eliminated the anti-senscence effect of nifedipine. These results demonstrate that nifedipine can prevent endothelial cell senescence in an eNOS-dependent manner. The anti-senescence action of nifedipine may represent a novel mechanism by which it protects against atherosclerosis. [ABSTRACT FROM AUTHOR]

OASIS is a basic leucine zipper (bZIP) transmembrane transcription factor that is activated in response to endoplasmic reticulum (ER) stress. Previously, we showed that OASIS regulates final maturation of goblet cells in the large intestine. In the present study, to elucidate the roles of OASIS under pathophysiological conditions, we examined the stress response and inflammatory responses in Oasis deficient (Oasis−/−) mice exposed to dextran sulfate sodium (DSS) to induce colitis. A significant loss of body weight and an increase of mortality were observed in Oasis−/− mice with DSS-induced colitis compared with those in WT mice. The mucosa of the large intestine in Oasis−/− mice exhibited severe damage involving inflammatory cell infiltration. The expression levels of ER stress and apoptosis markers in intestinal epithelial cells were upregulated in Oasis−/− mice. These abnormalities were improved by treatment with tauroursodeoxycholic acid, a chemical chaperone that facilitates protein folding. Taken together, our findings demonstrate that OASIS plays important roles in protection of the large intestinal mucosa in DSS-induced colitis through attenuation of ER stress and inflammation. [ABSTRACT FROM AUTHOR]

Background/Aim: Neointimal formation after vessel injury is a complex process involving multiple cellular and molecular processes. Inhibition of intimal hyperplasia plays an important role in preventing proliferative vascular diseases, such as restenosis. In this study, we intended to identify whether sodium ferulate could inhibit neointimal formation and further explore potential mechanisms involved. Methods: Cultured vascular smooth muscle cells (VSMCs) isolated from rat thoracic aorta were pre-treated with 200 µmol/L sodium ferulate for 1 hour and then stimulated with 1 µmol/L angiotensin II (Ang II) for 1 hour or 10% serum for 48 hours. Male Sprague-Dawley rats subjected to balloon catheter insertion were administrated with 200 mg/kg sodium ferulate (or saline) for 7 days before sacrificed. Results: In presence of sodium ferulate, VSMCs exhibited decreased proliferation and migration, suppressed intracellular reactive oxidative species production and NADPH oxidase activity, increased SOD activation and down-regulated p38 phosphorylation compared to Ang II-stimulated alone. Meanwhile, VSMCs treated with sodium ferulate showed significantly increased protein expression of smooth muscle α-actin and smooth muscle myosin heavy chain protein. The components of Notch pathway, including nuclear Notch-1 protein, Jagged-1, Hey-1 and Hey-2 mRNA, as well as total β-catenin protein and Cyclin D1 mRNA of Wnt signaling, were all significantly decreased by sodium ferulate in cells under serum stimulation. The levels of serum 8-iso-PGF2α and arterial collagen formation in vessel wall were decreased, while the expression of contractile markers was increased in sodium ferulate treated rats. A decline of neointimal area, as well as lower ratio of intimal to medial area was observed in sodium ferulate group. Conclusion: Sodium ferulate attenuated neointimal hyperplasia through suppressing oxidative stress and phenotypic switching of VSMCs. [ABSTRACT FROM AUTHOR]

Objective: Understanding variations in size and pattern of development of angiotensin II (Ang II)-induced abdominal aortic aneurysms (AAA) may inform translational research strategies. Thus, we sought insight into the temporal evolution of AAA in apolipoprotein (apo)E−/− mice. Approach: A cohort of mice underwent a 4-week pump-mediated infusion of saline (n = 23) or 1500 ng/kg/min of Ang II (n = 85) and AAA development was tracked via in vivo ultrasound imaging. We adjusted for hemodynamic covariates in the regression models for AAA occurrence in relation to time. Results: The overall effect of time was statistically significant (p<0.001). Compared to day 7 of AngII infusion, there was no decrease in the log odds of AAA occurrence by day 14 (−0.234, p = 0.65), but compared to day 21 and 28, the log odds decreased by 9.07 (p<0.001) and 2.35 (p = 0.04), respectively. Hemodynamic parameters were not predictive of change in aortic diameter (Δ) (SBP, p = 0.66; DBP, p = 0.66). Mean total cholesterol (TC) was higher among mice with large versus small AAA (601 vs. 422 mg/ml, p<0.0001), and the difference was due to LDL. AngII exposure was associated with 0.43 mm (95% CI, 0.27 to 0.61, p<0.0001) increase in aortic diameter; and a 100 mg/dl increase in mean final cholesterol level was associated with a 12% (95% CI, 5.68 to 18.23, p<0.0001) increase in aortic diameter. Baseline cholesterol was not associated with change in aortic diameter (p = 0.86). Conclusions: These are the first formal estimates of a consistent pattern of Ang II-induced AAA development. The odds of AAA occurrence diminish after the second week of Ang II infusion, and TC is independently associated with AAA size. [ABSTRACT FROM AUTHOR]

Background: Coronary artery disease (CAD) is a leading cause of mortality in many countries. Considerable studies have been carried out to investigate the relationship between the C242T and A640G polymorphisms of CYBA gene and CAD, but the results were still inconsistent. Hence we conducted a meta-analysis to clarify the association. Methods and Results: A total of 21 eligible literatures were included in the meta-analysis. We observed a significant decreased risk of CAD for C242T polymorphism in Asian population under an allelic model (OR 0.75; 95% CI 0.67–0.84) and a dominant model (OR 0.69; 95% CI 0.61–0.79), however, in overall population and other population no significant association was revealed. We also found A640G polymorphism may contribute to reducing CAD risk under an allelic model (OR 0.84; 95% CI 0.75–0.93), dominant model (OR0.77; 95% CI 0.64–0.92) and recessive model (OR0.82; 95% CI 0.69–0.97). No publication bias was found. Conclusion: Our meta-analysis confirmed a protective effect of C242Tpolymorphism on CAD in Asian population and indicated that A640G polymorphism was significantly associated with decreased risk of CAD. [ABSTRACT FROM AUTHOR]

Background: Adipose afferent reflex (AAR) is a sympatho-excitatory reflex induced by chemical stimulation of white adipose tissue (WAT). Ionotropic glutamate receptors including NMDA receptors (NMDAR) and non-NMDA receptors (non-NMDAR) in paraventricular nucleus (PVN) mediate the AAR. Enhanced AAR contributes to sympathetic activation and hypertension in obesity rats. This study was designed to investigate the role and mechanism of superoxide anions in PVN in modulating the AAR. Methodology/Principal Findings: Renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) were recorded in anesthetized rats. AAR was evaluated by the RSNA and MAP responses to injections of capsaicin into four sites of right inguinal WAT (8.0 nmol in 8.0 µl for each site). Microinjection of polyethylene glycol-superoxide dismutase (PEG-SOD), the superoxide anion scavenger tempol or the NAD(P)H oxidase inhibitor apocynin into the PVN decreased the baseline RSNA and MAP, and attenuated the AAR. Unilateral WAT injection of capsaicin increased superoxide anions in bilateral PVN, which was prevented by the WAT denervation. WAT injection of capsaicin increased superoxide anion level and NAD(P)H oxidase activity in the PVN, which was abolished by the PVN pretreatment with the combined NMDAR antagonist AP5 and non-NMDAR antagonist CNQX. Microinjection of the NMDAR agonist NMDA or the non-NMDAR agonist AMPA increased superoxide anion level and NAD(P)H oxidase activity in the PVN. Conclusions: NAD(P)H oxidase-derived superoxide anions in the PVN contributes to the tonic modulation of AAR. Activation of ionotropic glutamate receptors in the PVN is involved in the AAR-induced production of superoxide anions in the PVN. [ABSTRACT FROM AUTHOR]

Background: Reactive oxygen species are implicated in the physiopathogenesis of salt-induced hypertension and the C242T polymorphism of the p22-phox gene has been associated with higher superoxide production. This study investigated the impact of this polymorphism on the relationship between urinary sodium excretion (USE) and blood pressure levels in an urban Brazilian population. Methods: We cross-sectionally evaluated 1,298 subjects from the city of Vitoria-ES, located in the Southeast region of Brazil, by clinical history, physical examination, anthropometry, analysis of laboratory parameters, USE measurement and p22-phox C242T polymorphism genotyping. Results: No significant differences in studied parameters were detected between the studied genotype groups (CC vs. CT+TT). Systolic blood pressure exhibited significant correlation with USE only in T allele carriers (r = 0.166; p<0.001), while diastolic blood pressure and hypertension status correlated with USE in both genotypes albeit more weakly in subjects with CC genotype (r = 0.098; p = 0.021 and r = 0.105; p = 0.013, respectively) than in T carriers (r = 0.236; p<0.001 and r = 0.213; p<0.001, respectively). Regression analyses adjusted for confounding factors showed that USE remained independently associated with systolic (p<0.001) and diastolic blood pressure (p<0.001) and hypertension status (p = 0.004) only in T allele carriers. Finally, higher diastolic and systolic blood pressure levels were detected in T allele carriers than in CC genotype individuals in the highest tertile of USE. Conclusions: The p22-phox 242T allele is associated with higher blood pressure levels among subjects with higher USE in an urban Brazilian population. [ABSTRACT FROM AUTHOR]

Nuclear factor kappa B (NF-κB) and the Ets like gene-1 (Elk-1) are two transcription factors that have been previously established to contribute to the Angiotensin II mediated upregulation of Angiotensin II type 1 receptor (AT1R) in neurons. The cAMP response element binding protein (CREB) is another transcription factor that has also been implicated in AT1R gene transcription. The goal of the current study was to determine if NF-κB and CREB association was required for AT1R upregulation. We hypothesized that the transcription of the AT1R gene occurs via an orchestration of transcription factor interactions including NF-κB, CREB, and Elk-1. The synergistic role of CREB and NFκB in promoting AT1R gene expression was determined using siRNA-mediated silencing of CREB. Electrophorectic Mobility Shift Assay studies employing CREB and NF-κB demonstrated increased protein – DNA binding as a result of Ang II stimulation which was blunted by siRNA silencing of CREB. Upstream inhibition of p38 mitogen activated protein kinase (p38 MAPK) with SB203580 or inhibition of the calmodulin kinase (CAMK) pathway using KN-62 blunted changes in CREB and NF-κB expression. These findings suggest that Ang II may activate multiple signaling pathways involving p38 MAPK leading to the activation of NF-κB and CREB, which feed back to upregulate the AT1R gene. This study provides insight into the molecular mechanisms involving multiple transcription factor activation in a coordinated fashion which may be partially responsible for sympathoexcitation in clinical conditions associated with increased activation of the renin angiotensin system. [ABSTRACT FROM AUTHOR]

Background: Hypertension is a highly prevalent disorder and a major risk factor for cardiovascular diseases. Hypertensive vascular remodeling is the pathological mal-adaption of blood vessels to the hypertensive condition that contributes to further development of high blood pressure and end-organ damage. Hypertensive remodeling involves, at least in part, changes in protein turnover. The ubiquitin proteasome system (UPS) is a major protein quality and quantity control system. This study tested the hypothesis that the proteasome inhibitor, bortezomib, would attenuate AngII-induced hypertension and its sequelae such as aortic remodeling in rats. Methodology/Principal Findings: Male Sprague Dawley rats were subjected to AngII infusion for two weeks in the absence or presence of bortezomib. Mean arterial pressure was measured in conscious rats. Aortic tissue was collected for estimation of wall area, collagen deposition and expression of tissue inhibitors of matrix metalloproteases (TIMP), Ki67 (a marker of proliferation), reactive oxygen species (ROS) and VCAM-1 (a marker of inflammation). AngII infusion increased arterial pressure significantly (160±4 mmHg vs. vehicle treatment 133±2 mmHg). This hypertensive response was attenuated by bortezomib (138±5 mmHg). AngII hypertension was associated with significant increases in aortic wall to lumen ratio (∼29%), collagen deposition (∼14%) and expression of TIMP1 and TIMP2. AngII also increased MMP2 activity, proteasomal chymotrypsin-like activity, Ki67 staining, ROS generation and VCAM-1 immunoreactivity. Co-treatment of AngII-infused rats with bortezomib attenuated these AngII-induced responses. Conclusions: Collectively, these data support the idea that proteasome activity contributes to AngII-induced hypertension and hypertensive aortic vascular remodeling at least in part by modulating TIMP1/2 and MMP2 function. Preliminary observations are consistent with a role for ROS, inflammatory and proliferative mechanisms in this effect. Further understanding of the mechanisms by which the proteasome is involved in hypertension and vascular structural remodeling may reveal novel targets for pharmacological treatment of hypertension, hypertensive remodeling or both. [ABSTRACT FROM AUTHOR]

Background: We sought to quantify plasma high sensitivity cardiac troponin (hs-cTnT) levels, their determinants, and their associations with left ventricular (LV) myocardial deformation in adult survivors of childhood acute leukaemias. Methods and Results: One hundred adult survivors (57 males) of childhood acute leukaemias, aged 24.1±4.2 years, and 42 age-matched controls (26 males) were studied. Plasma cTnT was determined using a highly sensitive assay. Genotyping of NAD(P)H oxidase and multidrug resistance protein polymorphisms was performed. Left ventricular function was assessed by conventional, three-dimensional, and speckle tracking echocardiography. The medians (interquartile range) of hs-cTnT in male and female survivors were 4.9 (4.2 to 7.2) ng/L and 1.0 (1.0 to 3.5) ng/L, respectively. Nineteen survivors (13 males, 6 females) (19%) had elevated hs-cTnT (>95th centile of controls). Compared to those without elevated hs-TnT levels, these subjects had received larger cumulative anthracycline dose and were more likely to have leukaemic relapse, stem cell transplant, and cardiac irradiation. Their LV systolic and early diastolic myocardial velocities, isovolumic acceleration, and systolic longitudinal strain rate were significantly lower. Survivors having CT/TT at CYBA rs4673 had higher hs-cTnT levels than those with CC genotype. Functionally, increased hs-cTnT levels were associated with worse LV longitudinal systolic strain and systolic and diastolic strain rates. Conclusions: Increased hs-cTnT levels occur in a significant proportion of adult survivors of childhood acute leukaemias and are associated with larger cumulative anthracycline dose received, history of leukaemic relapse, stem cell transplant, and cardiac irradiation, genetic variants in free radical metabolism, and worse LV myocardial deformation. [ABSTRACT FROM AUTHOR]

Some studies have reported that angiotensin converting enzyme (ACE) and angiotensinogen (AGT) genes have been associated with hypertrophic cardiomyopathy (HCM). However, there have been inconsonant results among different studies. To clarify the influence of ACE and AGT on HCM, a systemic review and meta-analysis of case-control studies were performed. The following databases were searched to indentify related studies: PubMed database, the Embase database, the Cochrane Central Register of Controlled Trials database, China National Knowledge Information database, and Chinese Scientific and Technological Journal database. Search terms included “hypertrophic cardiomyopathy”, “angiotensin converting enzyme” (ACE) or “ACE” and “polymorphism or mutation”. For the association of AGT M235T polymorphism and HCM, “angiotensin converting enzyme” or “ACE” was replaced with “angiotensinogen”. A total of seventeen studies were included in our review. For the association of ACE I/D polymorphism and HCM, eleven literatures were included in the meta-analysis on association of penetrance and genotype. Similarly, six case-control studies were included in the meta-analysis for AGT M235T. For ACE I/D polymorphism, the comparison of DI/II genotype vs DD genotype was performed in the present meta-analysis. The OR was 0.73 (95% CI: 0.527, 0.998, P = 0.049, power = 94%, alpha = 0.05) after the study which deviated from Hardy-Weinberg Equilibrium was excluded, indicating that the ACE I/D gene polymorphism might be associated with HCM. The AGT M235T polymorphism did not significantly affect the risk of HCM. In addition, ACE I/D gene polymorphism did not significantly influence the interventricular septal thickness in HCM patients. In conclusion, the ACE I/D polymorphism might be associated with the risk of HCM. [ABSTRACT FROM AUTHOR]

Differentiation of cardiac fibroblasts into myofibroblasts is a critical event in the progression of cardiac fibrosis that leads to pathological cardiac remodeling. Metformin, an antidiabetic agent, exhibits a number of cardioprotective properties. However, much less is known regarding the effect of metformin on cardiac fibroblast differentiation. Thus, in the present study, we examined the effect of metformin on angiotensin (Ang) II-induced differentiation of cardiac fibroblasts into myofibroblasts and its underlying mechanism. Adult rat cardiac fibroblasts were stimulated with Ang II (100 nM) in the presence or absence of metformin (10–200 µM). Ang II stimulation induced the differentiation of cardiac fibroblasts into myofibroblasts, as indicated by increased expression of α-smooth muscle actin (α-SMA) and collagen types I and III, and this effect of Ang II was inhibited by pretreatment of cardiac fibroblasts with metformin. Metformin also decreased Ang II-induced reactive oxygen species (ROS) generation in cardiac fibroblasts via inhibiting the activation of the PKC-NADPH oxidase pathway. Further experiments using PKC inhibitor calphostin C and NADPH oxidase inhibitor apocynin confirmed that inhibition of the PKC-NADPH oxidase pathway markedly attenuated Ang II-induced ROS generation and myofibroblast differentiation. These data indicate that metformin inhibits Ang II-induced myofibroblast differentiation by suppressing ROS generation via the inhibition of the PKC-NADPH oxidase pathway in adult rat cardiac fibroblasts. Our results provide new mechanistic insights regarding the cardioprotective effects of metformin and provide an efficient therapeutic strategy to attenuate cardiac fibrosis. [ABSTRACT FROM AUTHOR]

Background: Observational and experimental studies have thus far been unable to resolve whether the CYBA C242T polymorphism is associated with coronary artery disease (CAD). Therefore, we undertook a comprehensive meta-analysis to more precisely evaluate the influence of this polymorphism on CAD and potential biases. Methods: We screened MEDLINE, Embase, CNKI, Wanfang and CBM up to January 2013 and extracted data from 22 studies with 9,279 CAD patients and 9,349 controls. A random-effects model was exploited to synthesize the inconsistent outcomes of the individual studies, while addressing between-study heterogeneity and publication bias. Results: The CYBA C242T polymorphism conformed to Hard-Weinberg Equilibrium for all studies (P>0.05). Overall comparison of the T allele with the C allele produced a non-significant risk estimate for CAD but with striking heterogeneity (T versus C: P = 0.87, OR = 0.99, 95%CI 0.89–1.11, Pheterogeneity<0.0001, I2 = 67.8%). However, subgroup analysis by ethnicity documented that the T allele carriers had a marginal risk increase (21%) of CAD among Caucasians (recessive genetic model: P = 0.05, 95%CI 1.00–1.46, Pheterogeneity = 0.15, I2 = 29.1%). Then data were divided into study design, the significance of CAD risk increase was substantially strengthened in matched case-control studies (allele comparison: P = 0.02, OR = 1.13, 95%CI 1.02–1.26, Pheterogeneity = 0.24, I2 = 21.6%).Further meta-regression analysis identified that a large proportion of heterogeneity was explained by body mass index (BMI) (P = 0.03, OR = 1.07, 95%CI 1.01–1.15) and study design (P = 0.03, OR = 1.30, 95%CI 1.02–1.64).There was no obvious publication bias as verified by funnel plot and Egger's linear regression test (t = −0.25, P = 0.81 for allele comparison). Conclusion: Taken together, our results suggested the CYBA C242T polymorphism might be a risk-conferring factor on developing CAD and BMI and study design were probable sources of between-study heterogeneity. [ABSTRACT FROM AUTHOR]

Objective: The risk of stroke in patients with Parkinson’s disease (PD) remains controversial. The purpose of this population-based propensity score-matched longitudinal follow-up study was to determine whether there is an increased risk of ischemic stroke after PD. Methods: We used a logistic regression model that includes age, sex, pre-existing comorbidities and socioeconomic status as covariates to compute the propensity score. A total of 2204 patients with at least two ambulatory visits with the principal diagnosis of PD in 2001 was enrolled in the PD group. The non- PD group consisted of 2204, propensity score-matched subjects without PD. The ischemic stroke-free survival rates of the two groups were estimated using the Kaplan-Meier method. Stratified Cox proportional hazard regression with patients matched on propensity score was used to estimate the effect of PD on the occurrence of ischemic stroke. Results: During the three-year follow-up period, 328 subjects in the PD group and 156 subjects in the non-PD group developed ischemic stroke. The ischemic stroke-free survival rate of the PD group was significantly lower than that of the non-PD group (P<0.0001). The hazard ratio (HR) of stroke for the PD group was 2.37 (95% confidence interval [CI], 1.92 to 2.93, P<0.0001) compared to the non- PD group. Conclusions: This study shows a significantly increased risk of ischemic stroke in PD patients. Further studies are required to investigate the underlying mechanism. [ABSTRACT FROM AUTHOR]

The CD38-ADP-ribosylcyclase-mediated Ca2+ signaling pathway importantly contributes to the vasomotor response in different arteries. Although there is evidence indicating that the activation of CD38-ADP-ribosylcyclase is associated with CD38 internalization, the molecular mechanism mediating CD38 internalization and consequent activation in response to a variety of physiological and pathological stimuli remains poorly understood. Recent studies have shown that CD38 may sense redox signals and is thereby activated to produce cellular response and that the NADPH oxidase isoform, NOX1, is a major resource to produce superoxide (O2·−) in coronary arterial myocytes (CAMs) in response to muscarinic receptor agonist, which uses CD38-ADP-ribosylcyclase signaling pathway to exert its action in these CAMs. These findings led us hypothesize that NOX1-derived O2·− serves in an autocrine fashion to enhance CD38 internalization, leading to redox activation of CD38-ADP-ribosylcyclase activity in mouse CAMs. To test this hypothesis, confocal microscopy, flow cytometry and a membrane protein biotinylation assay were used in the present study. We first demonstrated that CD38 internalization induced by endothelin-1 (ET-1) was inhibited by silencing of NOX1 gene, but not NOX4 gene. Correspondingly, NOX1 gene silencing abolished ET-1-induced O2·− production and increased CD38-ADP-ribosylcyclase activity in CAMs, while activation of NOX1 by overexpression of Rac1 or Vav2 or administration of exogenous O2·− significantly increased CD38 internalization in CAMs. Lastly, ET-1 was found to markedly increase membrane raft clustering as shown by increased colocalization of cholera toxin-B with CD38 and NOX1. Taken together, these results provide direct evidence that Rac1-NOX1-dependent O2·− production mediates CD38 internalization in CAMs, which may represent an important mechanism linking receptor activation with CD38 activity in these cells. [ABSTRACT FROM AUTHOR]