bluuebunny/crossref_metadata_2025 huggingface dataset:
Aminoguanidine and ramipril prevent diabetes-induced increases in protein kinase C activity in glomeruli, retina and mesenteric artery
Authors: OSICKA Tanya M. , YU Yunxia , LEE Vincent , PANAGIOTOPOULOS Sianna , KEMP Bruce E. , JERUMS George
Published: August 2015

This study investigated the effects of insulin therapy, inhibition of advanced glycation end-product formation with aminoguanidine and angiotensin-converting enzyme inhibition with ramipril on diabetes-related increases in protein kinase C (PKC) activity in the streptozotocin-diabetic rat. PKC activity in the glomeruli, retina and mesenteric artery was increased by 1.5–2-fold after induction of diabetes, and this increase was maintained over 24 weeks. Treatment with insulin at 2 units or 6 units per day attenuated glomerular PKC in proportion to the level of glycohaemoglobin after 4 weeks of diabetes (r = 0.68, P < 0.0001). The higher dose of insulin prevented the diabetes-related increase in glomerular PKC activity, although blood glucose levels were not normalized. After 8 weeks of diabetes, ramipril completely prevented the diabetes-related increases in PKC activity in the glomeruli, retina and mesenteric artery. By contrast, aminoguanidine treatment resulted in no inhibition of glomerular PKC activity, partial inhibition of retinal PKC activity and complete inhibition of mesenteric artery PKC activity. After 24 weeks of diabetes, both aminoguanidine and ramipril prevented the diabetes-related increases in PKC activity in all three tissues, in parallel with suppression of albuminuria by both agents. Aminoguanidine also prevented diabetes-related increases in retinal permeability at 16 weeks. These results suggest that the organ-protective effects of insulin, aminoguanidine and ramipril in diabetes may be mediated, at least in part, through the differential inhibition of PKC activity in various tissues.
Peroral immunization with Helicobacter pylori adhesin protein genetically linked to cholera toxin A2B subunits
Authors: KIM Byung Oh , SHIN Sung Seup , YOO Young Hyo , PYO Suhkneung
Published: August 2015

Helicobacter pylori is a major cause of gastric-associated diseases. To evaluate the efficacy of a possible vaccine antigen against H. pylori infection, the chimaeric construct adhesin–CTXA2B, derived from H. pylori adhesin genetically coupled to cholera toxin (CTX) subunits A2 and B (CTXA2B), was expressed in Escherichia coli as an insoluble recombinant chimaeric protein. The protein was then purified by denaturation, renaturation and size-exclusion chromatography. The composition of purified adhesin–CTXA2B was verified by SDS/PAGE and Western blotting with antibodies to antigenic components of adhesin and CTXB, and confirmed as a chimaeric protein with GM1-ganglioside binding activity and adhesin epitopes by a GM1-ELISA developed using antibodies to adhesin. Oral immunization of mice with adhesin–CTXA2B induced higher levels of mucosal IgA and serum IgG antibodies to H. pylori adhesin and to CTXB than in mice immunized with adhesin or CTXA2B alone. Adhesin–CTXA2B was also demonstrated to be a potential protective antigen in a mouse model of H. pylori infection. The immunization of mice with adhesin–CTXA2B protected 62.5% of mice infected with H. pylori SS1 strain, whereas adhesin immunization was not able to confer protection to mice. This protection may be correlated with high levels of mucosal IgA and serum IgG antibodies against H. pylori adhesin. Taken together, the results indicate that the genetically linked CTXA2B acts as a useful mucosal adjuvant, and that the adhesin–CTXA2B chimaeric protein could be a potential component in future H. pylori vaccine development.
The bile acid taurocholate impairs rat cardiomyocyte function: a proposed mechanism for intra-uterine fetal death in obstetric cholestasis
Authors: WILLIAMSON Catherine , GORELIK Julia , EATON Bryan M. , LAB Max , SWIET Michael de , KORCHEV Yuri
Published: August 2015

Obstetric cholestasis is a liver disease of pregnancy that can be complicated by sudden, hitherto unexplained, intra-uterine fetal death. Because intra-uterine death occurs suddenly, and because fetal heart rate abnormalities have been reported in obstetric cholestasis, we hypothesized that intra-uterine death is caused by impaired fetal cardiomyocyte function, resulting in fetal cardiac arrest. Obstetric cholestasis is associated with raised levels of maternal and fetal serum bile acids, and we propose that these may alter cardiomyocyte function. It was not possible to investigate the effects of bile acids on the intact human fetal heart at a cellular level. Therefore we used the closest available model of fetal myocardium at term: a primary culture of neonatal rat cardiomyocytes in which cells beat synchronously and develop pacemaker activity. The effect of the primary bile acid taurocholate (0.3 mM and 3 mM) on cultures of single cardiomyocytes, each with its own independent rate of contraction, was a reversible decrease in the rate of contraction and in the proportion of beating cells (P < 0.001). Addition of taurocholate to a network of synchronously beating cells caused a similar decrease in the rate of contraction. Furthermore, the integrity of the network was destroyed, and cells ceased to beat synchronously. Taurocholate also resulted in altered calcium dynamics and loss of synchronous beating. These data suggest that raised levels of the bile acid taurocholate in the fetal serum in obstetric cholestasis may result in the development of a fetal dysrhythmia and in sudden intra-uterine death.
A novel method for the delivery of nitric oxide therapy to the skin of human subjects using a semi-permeable membrane
Authors: HARDWICK J. B. J. , TUCKER A. T. , WILKS M. , JOHNSTON A. , BENJAMIN N.
Published: August 2015

Nitric oxide (NO) is a mediator of essential biological processes, including vasodilatation, anti-microbial activity and wound healing. A chemical system using sodium nitrite and ascorbic acid has been developed which generates significant amounts of NO. The originally described system was messy and impractical, and the high acidity may cause pain and further tissue damage in ulcerated skin. To overcome this, a selectively permeable, hydrophilic polyester co-polymer membrane system (Sympatex™) has been identified that can be placed between the NO-generating chemicals and the skin. The aim of the present study was to determine whether NO derived from this chemical system was able to diffuse through the membrane and have a measurable vasodilatory effect on forearm skin in healthy volunteers. The Sympatex™ 10 μ m membrane was found to be highly permeable to NO, while preventing passage of the constituents of the NO-generation gel to the skin. The transmembrane NO-generation system had a vasodilatory effect comparable with that resulting from direct topical application. Additionally, the NO generated was effective in killing Staphylococcus aureus and Escherichia coli at doses lower than those required to increase skin blood flow. The vasodilatory and anti-microbial effects of this system may be useful as a patch-based topical therapy for skin ulceration, particularly when there is concomitant ischaemia and infection.
β2-Adrenoceptor desensitization in human alveolar macrophages induced by inhaled terbutaline in vivo is not counteracted by budesonide
Authors: ZETTERLUND Anita , HJEMDAHL Paul , LARSSON Kjell
Published: August 2015

In vitro studies suggest that glucocorticoids may counteract β-agonist-induced desensitization of β-adrenoceptors by actions at the transcriptional level, but the clinical relevance of such findings is not clear. Oral terbutaline treatment decreases β-adrenoceptor sensitivity in alveolar macrophages in vivo. This effect is not counteracted by inhaled or orally taken steroids. We therefore examined whether inhaled terbutaline elicited a similar effect on β2-adrenoceptor sensitivity in alveolar macrophages, and if co-treatment with an inhaled steroid, budesonide, would prevent such down-regulation. Bronchoalveolar lavage (BAL) and lung function tests, including bronchodilator responses to inhaled terbutaline, were performed before and after 2 weeks of regular inhalation of terbutaline, 0.5 mg three times daily, and budesonide, 400 µg twice daily, or placebo, in 24 healthy volunteers. Four untreated subjects served as controls. A marked, approx. 90%, decrease in isoprenaline-induced cAMP accumulation in alveolar macrophages was found in both treatment groups after 2 weeks, with no difference between placebo and budesonide (P = 0.45). In the untreated control group, cAMP responses to both isoprenaline and prostaglandin E1 tended to be lower on the second occasion. A limited, non-specific desensitization of adenylate cyclase activity thus contributed to the marked desensitization elicited by terbutaline inhalations. The bronchodilator response to inhaled terbutaline did not change after treatment in any of the three groups (F = 0.9, P = 0.50). In conclusion, inhalation of a β-agonist induced marked down-regulation of β2-adrenoceptor sensitivity in alveolar macrophages in vivo without influencing the bronchodilator response to a β2-agonist in healthy subjects. Co-treatment with an inhaled steroid failed to counteract the desensitization of alveolar macrophage β2-adrenoceptors.
Effects of smoking and abstention from smoking on fibrinogen synthesis in humans
Authors: HUNTER Kirsty A. , GARLICK Peter J. , BROOM Iain , ANDERSON Susan E. , McNURLAN Margaret A.
Published: August 2015

Cigarette smoking and hyperfibrinogenaemia are both significant risk factors for the development of cardiovascular disease. Two studies are described here which aimed to establish the metabolic mechanism responsible for the raised plasma fibrinogen concentration observed in smokers. Chronic smokers had a significantly elevated absolute rate of fibrinogen synthesis (ASR) compared with non-smokers (22.7±1.3 mg/kg per day versus 16.0±1.3 mg/kg per day; means±S.E.M., P < 0.01), with plasma levels of fibrinogen significantly correlated with fibrinogen synthesis (r = 0.65, P = 0.04). Unlike fibrinogen, plasma albumin concentrations were lower in smokers than in non-smokers (45±0.4 versus 47±0.7 g/l, P < 0.05), but there was no difference in rates of albumin synthesis between the two groups. Two weeks cessation from smoking by previously chronic smokers was associated with a rapid and marked fall in plasma fibrinogen concentration (from 3.06±0.11 g/l to 2.49±0.14 g/l, P < 0.001), and a significant reduction in ASR (a 33% reduction, from 24.1±1.7 to 16.1±1.0 mg/kg per day, P < 0.001). These studies suggest a primary role for increased synthesis in producing the hyperfibrinogenaemia associated with smoking. Moreover, abstention from smoking for a period of only 2 weeks induces a significant decrease in the rate of fibrinogen synthesis by the liver, with a concomitant reduction in the plasma fibrinogen concentration.
Gender differences in protein kinase G-mediated vasorelaxation of rat aorta
Authors: TEEDE Helena , VAN DER ZYPP Andrea , MAJEWSKI Henryk
Published: August 2015

Although gender and oestrogen treatment influence production of the vasorelaxant, NO, their influence on factors downstream in the NO signal-transduction pathway, specifically protein kinase G (PKG), remains unknown. We aimed to study the influence of sex hormones on PKG, along with the endothelial modulation of these effects, in rat thoracic aortic rings in two separate groups, control male and female rats and ovariectomized female rats after treatment with oestrogen or vehicle. Vessel preparations were preconstricted with phenylephrine (0.1 μM). Constrictions were greater in male than female aortas. This differential effect was attenuated by endothelium removal, addition of the guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 1μM) and the nitric oxide synthase (NOS) inhibitor NG-monomethyl-l-arginine (l-NMMA, 100 μM), supporting the role of NO in maintenance of basal relaxation and vascular tone in females. We have examined the relative activity of the specific PKG subtypes 1α and 1β in vascular smooth muscle, based on relaxation of rat aortas by two cGMP analogues with different selectivity, β-phenyl-l-N2-ethano-8-bromo-cGMP (8-Br-PET-cGMP) and 8-(2-aminophenylthio)cGMP (8-APT-cGMP). 8-Br-PET-cGMP was more potent than 8-APT-cGMP in both sexes, suggesting that PKG 1α is the primary subtype involved in vasorelaxation. The gender differences in PKG activity were examined based on relaxation responses in male and female rat aortas. Both 8-Br-PET-cGMP and 8-APT-cGMP were more potent in aortas from male than female rats. In further studies on the endothelial modulation of relaxation with 8-APT-cGMP, the differential gender-vasorelaxation response was negated by endothelium removal and addition of the guanylate cyclase inhibitor ODQ (1 μM), but not by the NOS inhibitor l-NMMA (100 μM), suggesting that an endothelial-dependent factor other than NO may be responsible for the observed differential PKG-mediated vasorelaxation between the sexes. To further investigate oestrogen influence on PKG, treated female rats were studied. Contrary to our hypothesis, in the presence of 1 μM ODQ, there were no differences in either the phenylephrine constriction, or the relaxation with 8-APT-cGMP from either sham-operated, vehicle-treated or oestrogen-treated ovariectomized rats. In conclusion, female rat aortas have greater basal NO production compared with males. Relaxant responses to PKG activation are greater in aortas from male compared with female rats. These findings suggest hormonal regulation of PKG; however, oestrogen treatment of ovariectomized rats did not affect PKG activity, suggesting factors other than oestrogen may be responsible for the gender differences noted in this study.
Increased levels of typically fetal bile acid species in patients with hepatocellular carcinoma
Authors: EL-MIR Mohamad Y. , BADIA Maria D. , LUENGO Nazaret , MONTE Maria J. , MARIN Jose J. G.
Published: August 2015

The aim of this work was to investigate the reappearance during liver neoplasia of bile acids (BAs) species, which are unusual in healthy adults, but common in fetuses. Serum and urine samples were collected from patients with hepatocellular carcinoma (HCC; n = 27), and for comparative purposes, with liver cirrhosis (n = 49), liver metastasis (n = 19), chronic viral hepatitis (n = 11) and healthy volunteer (control group; n = 26) groups. BAs were identified and measured by GC–MS. Hypercholanaemia was found in all groups of patients. In HCC, this was characterized by a marked increase in the chenodeoxycholate/cholate ratio in both serum and urine. Although increased levels of BAs, with hydroxylations at unusual positions, and oxo-BAs were found in HCC, these were not significantly different from those observed in other groups. However, BAs with a flat structure, i.e. Δ4-unsaturated- and 5α- or allo-BAs, which were almost absent in healthy subjects, were markedly increased in the serum and urine of HCC patients. They were also detected, although in much lower amounts, in liver metastasis and liver cirrhosis, but not in viral hepatitis. Flat-BAs were better detected in urine than in serum. Urinary Δ4-unsaturated-BA output was significantly lower in patients with small tumours (< 3 cm) compared with those with higher size tumours. No correlation between flat-BA output into urine and serum alpha-fetoprotein or total BAs was found. These results suggest that Δ4- and/or allo-BAs are particularly elevated in patients with HCC, which may be a potentially useful complementary, rather than alternative, marker for early detection of liver neoplasia.
Effects of prior moderate exercise on exogenous and endogenous lipid metabolism and plasma factor VII activity
Authors: GILL Jason M. R. , FRAYN Keith N. , WOOTTON Stephen A. , MILLER George J. , HARDMAN Adrianne E.
Published: August 2015

Moderate exercise reduces postprandial triacylglycerol concentrations, which are a risk marker for coronary heart disease. The present study sought to determine the qualitative nature of exercise-induced changes in lipid metabolism and their association (if any) with changes in factor VII activation. Eleven normotriglyceridaemic men, aged 51.7±6.1 years (mean±S.D.), participated in two oral fat tolerance tests after different pre-conditions: control (no exercise), and exercise (90 min of brisk walking the day before). Venous blood samples were obtained in the fasted state and for 8 h after ingestion of a high-fat meal (1.32 g of fat, 1.36 g of carbohydrate, 0.30 g of protein and 10 mg of [1,1,1-13C] tripalmitin·kg-1 body mass). Prior exercise reduced postprandial plasma triacylglycerol concentrations by 25±3% (mean±S.E.M.), with lower concentrations in the Svedberg flotation rate (Sf) 20–400 (very-low-density lipoprotein) fraction accounting for 79±10% of this reduction. There was no effect on plasma factor VII coagulant activity or on the concentration of the active form of factor VIIa. Prior exercise increased postprandial serum 3-hydroxybutyrate and plasma fatty acid concentrations, decreased serum postprandial insulin concentrations and increased exogenous (8 h 13C breath excretion of 15.1±0.9% of ingested dose compared with 11.9±0.8%; P = 0.00001) and endogenous postprandial fat oxidation. These data raise the possibility that reduced hepatic secretion of very-low-density lipoprotein plays a role in the attenuation of plasma triacylglycerol concentrations seen after exercise, although it is possible that increased triacylglycerol clearance also contributes to this effect.
Influence of an inducible nitric oxide synthase promoter variant on clinical variables in patients with coronary artery disease
Authors: MORRIS Brian J. , GLENN Cheryl L. , WILCKEN David E. L. , WANG Xing Li
Published: August 2015

Pathophysiological processes in coronary artery disease (CAD) are influenced by genetic factors. Since (i) inducible nitric oxide synthase (iNOS) has important cardiovascular effects, and (ii) the promoter of the iNOS gene (NOS2A) is genetically modulated by a 4 bp insertion/deletion (+/-) polymorphism located 0.7 kb upstream, we decided to examine the influence of this variant on clinical variables in 856 CAD patients of Anglo-Celtic/Northern European extraction. We found that 2% of CAD patients were homozygous for the + allele, and 19% were heterozygous. Males made up 74% of the patient group, and in these the + allele was associated with 38% higher plasma glucose levels (P = 0.005), a 4.8% elevation in the waist/hip ratio (P = 0.009) and a 48% greater frequency of unstable angina (P = 0.014). The + allele, by influencing iNOS expression, could thus contribute to indices of insulin resistance and angina severity in male CAD patients.
Measurement of free and complexed soluble vascular endothelial growth factor receptor, Flt-1, in fluid samples: development and application of two new immunoassays
Authors: BELGORE Funmi M. , BLANN Andrew D. , LIP Gregory Y.
Published: August 2015

Vascular endothelial growth factor (VEGF) mediates endothelial cell mitogenesis and enhances vascular permeability. VEGF interacts with the endothelium via two membrane-spanning receptors, fms-like tyrosine kinase (Flt)-1 and kinase domain receptor. A soluble form of Flt-1 (sFlt-1) was isolated from endothelial cell media; however, its biological significance is still unknown, with limited data on plasma sFlt-1 levels in disease states. We have developed two new ELISAs for detecting free and VEGF-complexed sFlt-1, which were tested in accordance with standard validation and assessment methodologies employed in commercial settings. The intra-and inter-assay coefficients of variation are < 5% and 10% respectively, and results are highly reproducible. Applying these ELISAs in a clinical setting, we measured levels of VEGF, free and complexed sFlt-1 in citrated plasma from 40 patients with cardiovascular disease and 40 healthy controls. Median (interquartile range) plasma levels of VEGF in patients were significantly greater than controls [403 pg/ml (158–925 pg/ml) versus 113 pg/ml (33–231 pg/ml), P ⩽ 0.05]. Free sFlt-1 was significantly lower in patients compared with controls [8 ng/ml (2–22 ng/ml) versus 21 ng/ml (10–73 ng/ml), P ⩽ 0.05]. There was no significant difference in the levels of complexed sFlt-1 between the two groups. Plasma levels of VEGF-complexed sFlt-1 are minimal, despite the presence of excess free sFlt-1. Thus unbound plasma VEGF detected by ELISA may represent the majority of circulating VEGF, and justifies the measurement of plasma VEGF as an indicator of circulating VEGF levels. Furthermore, these results suggest that circulating sFlt-1 may serve as a selective inhibitor of VEGF activity, and that this regulatory mechanism may be altered by pathological conditions.
Diet-induced endothelial dysfunction in the rat is independent of the degree of increase in total body weight
Authors: NADERALI Ebrahim K. , PICKAVANCE Lucy C. , WILDING John P. H. , WILLIAMS Gareth
Published: August 2015

A growing number of studies indicate an association between obesity, insulin resistance, dyslipidaemia and cardiovascular disorders, collectively known as Syndrome X. In this study we have aimed to produce a model of Syndrome X by voluntary feeding of Wistar rats with a highly palatable cafeteria diet, and examined its effects on metabolic changes and vascular reactivity of Wistar rats. At the end of the experiment, the cafeteria-diet fed group was divided into two groups of low weight gain (LWG) and high weight gain (HWG). Both LWG and HWG groups had significantly (P < 0.01) higher fat-pad mass than their chow-fed counterparts, while gastrocnemius muscle mass were comparable. All cafeteria-diet fed rats had significantly (P < 0.01) raised plasma triacylglycerol (TG) levels whereas plasma non-esterified fatty acids, glucose and insulin levels were similar between chow-fed and cafeteria-diet fed rats. Vasorelaxation responses to acteylcholine, insulin and sodium nitroprusside were significantly (P < 0.01) attenuated in cafeteria-diet fed animals; however, there were no differences in contractile responses of the mesenteric arteries to noradrenaline or KCl between the groups. Multiple regression analysis showed a significant (P < 0.05) negative association between plasma TG levels and reduction in acetylcholine-induced vasorelaxation. Acetylcholine-induced vasorelaxation was also significantly (P < 0.05) associated with the amount of fat-pad mass. These data suggest that diet-induced vascular dysfunction can occur in the absence of insulin resistance, and that plasma TGs may have a detrimental effect on vascular reactivity.
Ethnic differences in erythrocyte membrane fluidity and the association with serum triacylglycerols
Authors: MILLER Michelle A. , SAGNELLA Giuseppe A. , MARKANDU Nirmala D. , MacGREGOR Graham A.
Published: August 2015

The objectives of this study were to determine whether there are differences between black and white individuals with regard to the membrane fluidity of isolated erythrocytes, and/or in the relationships between membrane fluidity, gender and circulating lipids. Fluorescent polarization anisotropy, as an index of membrane fluidity, was determined using the fluorescent probe 1-(4-trimethylammoniumphenyl)-6-phenyl-1,3,5-hexatriene (TMA-DPH) in 52 black and 52 white individuals, of whom 39 pairs were matched for age, sex and blood pressure. In the 39 matched pairs, the TMA-DPH anisotropy was significantly higher in the black (0.262±0.007) compared with the white (0.258±0.005) subjects (P < 0.005). There was also a significant difference in serum lipids. Gender differences in TMA-DPH anisotropy were observed in the white but not in the black individuals. The associations between membrane fluidity and serum lipids were examined in the total group, separated according to ethnic group. Although the associations were in the same direction in both groups, the association was only significant in the white subjects (r = - 0.42; P < 0.02). The ethnic difference in membrane fluidity was abolished when adjusting for serum triacylglycerols. In conclusion, ethnic differences in erythrocyte membrane fluidity, as determined by the use of TMA-DPH anisotropy, appear to be the result of ethnic differences in the level of serum triacylglycerols.
Effect of cigarette smoking on levels of bioavailable testosterone in healthy men
Authors: ENGLISH Katherine M. , PUGH Peter J. , PARRY Helen , SCUTT Nanette E. , CHANNER Kevin S. , JONES T. Hugh
Published: August 2015

The effect of smoking on androgen levels is important given the recent interest in the link between low levels of androgens and the development of cardiovascular disease. Numerous studies examining the effects of cigarette smoking on the levels of total and free testosterone have reported conflicting findings, but there has been no accurate assessment of the effects of cigarette smoking on the levels of bioavailable testosterone [not bound to sex hormone-binding globulin (SHBG)]. We attempted to determine whether smoking affects the level of bioavailable testosterone. We undertook a case-control study of 25 healthy male smokers and 25 healthy never-smokers, matched by age and body mass index. Early morning levels of total, free and bioavailable testosterone, 17β-oestradiol, SHBG and cotinine were determined and compared between the two groups. Levels of total (18.5 ± 4.6 ± nM versus 15.1 ± 4.9 ± nM, P = 0.01) and free testosterone (462 ± 91 ± pM versus 402 ± 93 ± pM, P = 0.03) were found to be higher in smokers compared with non-smokers respectively, as was SHBG (34.1 ± 12.8 versus 28.1 ± 9.0 ± nM, P = 0.06). There were no significant differences in the levels of bioavailable testosterone (3.78 ± 1.59 versus 3.51 ± 1.26 ± nM, P = 0.49) or 17β-oestradiol (44.5 ± 11.4 versus 42.3 ± 11.5 ± pM, P = 0.50) between smokers and non-smokers respectively. These data suggest that cigarette smoking has no significant effect on the biologically active fraction of testosterone, but may influence the levels of total and free testosterone through changes in the levels of SHBG.
Effect of adrenomedullin on the production of endothelin-1 and on its vasoconstrictor action in resistance arteries: evidence for a receptor-specific functional interaction in patients with heart failure
Authors: HILLIER Chris , PETRIE Mark C. , LOVE Michael P. , JOHNSTON Fiona , MACLEAN Margaret R. , MCMURRAY John J. V.
Published: August 2015

Endothelin-1 (ET-1) and adrenomedullin (ADM) are both produced in the arterial wall, but have opposing biological actions. Evidence from experimental animals suggests a functional interaction between ET-1 and ADM. We have tested this in humans. Small resistance arteries were obtained from gluteal biopsies taken from patients with chronic heart failure (CHF) due to coronary heart disease (CHD), or with CHD and preserved ventricular function. The contractile responses to big ET-1 and to ET-1 in both sets of vessels were studied in the absence (control) and presence of ADM at 20 pmol/l (low ADM) or 200 pmol/l (high ADM), using wire myography. ADM did not affect the conversion of big ET-1 into ET-1 in vessels from patients with either CHD or CHF. Low ADM did not alter the contractile response to ET-1 in vessels from patients with CHF. Low ADM was not tested in vessels from patients with CHD, but high ADM did not affect this response in arteries from these patients. High ADM did, however, significantly reduce the vasoconstrictor effect of ET-1 in vessels from patients with CHF. The maximum response, as a percentage of the response to high potassium, was 199% (S.E.M. 25%) in the control experiments (n = 14), 205% (27%) in the low-ADM (n = 7) studies and 150% (17%) in the high-ADM (n = 6) experiments (P < 0.001). Furthermore, the Hill coefficient increased from 0.57±0.05 in the absence of ADM to 1.16±0.15 in the high-ADM experiments, indicating that ADM at 200 pmol/l specifically antagonized one receptor type in vessels from patients with CHF. We conclude that there is a one-site receptor interaction between ADM and ET-1 that is specific for vessels from patients with CHF. This functional interaction between ADM and ET-1 in resistance arteries may be of pathophysiological importance in CHF.
Binding and uptake of chylomicron remnants by primary and THP-1 human monocyte-derived macrophages: determination of binding proteins
Authors: ELSEGOOD Caryn L. , PAL Sebely , ROACH Paul D. , MAMO John C. L.
Published: August 2015

The binding and uptake of chylomicron remnants by human macrophages was studied in order to resolve paradoxical observations that have described the putative mechanisms by which postprandial lipoproteins induce foam cell formation. Chylomicron remnants bound to human monocyte-derived macrophages (HMMs) and to the transformed monocytic cell line THP-1 with high affinity (Kd of approx. 5.5 µg of chylomicron remnant protein/ml). Binding was found to be saturable for both cell types, and was strongly inhibited in the presence of unlabelled chylomicron remnants. Ligand blot studies with colloidal-gold-labelled chylomicron remnants identified two cell surface binding sites on both HMMs and THP-1 cells, with molecular masses of approx. 128 kDa and 43 kDa. The high-molecular-mass binding site was found to be the low-density lipoprotein (LDL) receptor, based on the strong inhibition of chylomicron remnant binding in the presence of unlabelled LDL, Fab2 antibody fragments to the LDL receptor or calcium chelators. Competition studies suggested that, in HMMs, the LDL receptor appeared to facilitate approximately half of the total chylomicron remnant uptake. In contrast, the LDL receptor was not significantly involved in macrophage uptake of chylomicron remnants by THP-1 cells. The identity of the 43 kDa binding site is presently unknown, but, importantly, expression was not inhibited as a consequence of sterol loading, which was induced by incubating HMMs and THP-1 cells with 25-hydroxycholesterol. In contrast, the expression of the LDL receptor was substantially attenuated following lipid loading. Collectively, our data suggest that, while the macrophage LDL receptor can bind chylomicron remnants and facilitate uptake in non-lipid-loaded HMMs, other sterol-insensitive sites are responsible for the unabated uptake of chylomicron remnants by macrophages. We propose that the 43 kDa macrophage chylomicron remnant binding protein may be a candidate for the sterol loading of macrophages.
Differences in lymph drainage between swollen and non-swollen regions in arms with breast-cancer-related lymphoedema
Authors: STANTON A. W. B. , SVENSSON W. E. , MELLOR R. H. , PETERS A. M. , LEVICK J. R. , MORTIMER P. S.
Published: August 2015

Recent research indicates that the pathophysiology of breast-cancer-related lymphoedema (BCRL) is more complex than simple axillary lymphatic obstruction as a result of the cancer treatment. Uneven distribution of swelling (involvement of the mid-arm region is common, but the hand is often spared) is puzzling. Our aim was to test the hypothesis that local differences in lymphatic drainage contribute to the regionality of the oedema. Using lymphoscintigraphy, we measured the removal rate constant, k (representing local lymph flow per unit distribution volume, VD), for 99mTc-labelled human immunoglobulin G in the oedematous proximal forearm, and in the hand (finger web) in women in whom the hand was unaffected. Tracer was injected subcutaneously, and the depot plus the rest of the arm was monitored with a γ-radiation camera for up to 6 h. VD was assessed from image width. Contralateral arms served as controls. k was 25% lower in oedematous forearm tissue than in the control arm (BCRL, -0.070±0.026%·min-1; control, -0.093±0.028%·min-1; mean±S.D.; P = 0.012) and VD was greater. In the non-oedematous hand of the BCRL arm, k was 18% higher than in the control hand (BCRL, -0.110±0.027%·min-1; control, -0.095±0.028%·min-1; P = 0.057) and 59% higher than forearm k on the BCRL side (P = 0.0014). VD did not differ between the hands. Images of the BCRL arm following hand injection showed diffuse activity in the superficial tissues, sometimes extending almost to the shoulder. A possible interpretation is that the hand is spared in some patients because local lymph flow is increased and diverted along collateral dermal routes. The results support the hypothesis that regional differences in surviving lymphatic function contribute to the distribution of swelling.
Acute mental stress elicits delayed increases in circulating inflammatory cytokine levels
Authors: STEPTOE Andrew , WILLEMSEN Gonneke , OWEN Natalie , FLOWER Louise , MOHAMED-ALI Vidya
Published: August 2015

The influence of acute mental stress on cardiovascular responses andconcentrations of inflammatory cytokines up to 2 h later was assessed in 12 subjects exposed to stress and in eight control subjects. Beat-by-beat recordings of finger blood pressure and heart rate were made at rest and during two behavioural tasks (colour–word interference and mirror tracing). Blood was drawn after adaptation and at 45 min and 2 h after the tasks, and assayed for interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α), interleukin-1 receptor antagonist (IL-1Ra), C-reactive protein (CRP) and haematocrit. Saliva was sampled periodically and assayed for free cortisol. The tasks were rated as stressful by the participants. The stress group showed significant increases in systolic and diastolic blood pressure (mean rises of 16.4±12.3 and 12.6±6.9 mmHg respectively) and heart rate (5.39±5.3 beats/min); these values returned to baseline during the recovery period. The IL-6 concentration was increased by 56% at 2 h after the tasks (P< 0.05), while IL-1Ra was increased by 12.3% (P< 0.01). No changes in cardiovascular variables or cytokine concentrations were observed in the control subjects, and haematocrit did not change. The magnitude of blood pressure responses during tasks was correlated positively with the IL-6 concentration after 45 min (r = 0.70, P< 0.05), and with the IL-1Ra concentration after 2 h (r = 0.63, P< 0.05). Increases in TNF-α after 2 h were correlated with heart rate responses to tasks (r = 0.66, P< 0.05). Associations between IL-6 and IL-1Ra concentrations were also recorded. This study indicates that inflammatory cytokines respond to acute mental stress in humans with delayed increases, and suggest that individual differences in cytokine responses are associated with sympathetic reactivity.
RAPID COMMUNICATIONIn vivo inhibition of renal 11β-hydroxysteroid dehydrogenase in the rat stimulates collecting duct sodium reabsorption
Authors: BAILEY M. A. , UNWIN R. J. , SHIRLEY D. G.
Published: August 2015

In order to test the proposal that the aldosterone specificity of mineralocorticoid receptors in the collecting duct depends on inactivation of glucocorticoids by the enzyme 11β-hydroxysteroid dehydrogenase (11β-HSD), we have assessed the effect of pharmacological inhibition of 11β-HSD on collecting duct Na+ reabsorption in vivo. Adrenalectomized rats (n = 14) were infused intravenously with high-dose corticosterone, and late-distal tubules were perfused orthogradely with artificial tubular fluid containing [14C]inulin and 22Na; urinary recoveries of the radioisotopes were monitored. Half of the rats received intravenous carbenoxolone to inhibit renal 11β-HSD activity. The urinary recovery of [14C]inulin was complete in both groups of animals (101ŷ2% versus 101ŷ3%), but the recovery of 22Na was lower in carbenoxolone-treated rats (34ŷ5%) than in the corticosterone-alone group (54ŷ4%, P < 0.01). These data, which provide the first demonstration of enhanced Na+ reabsorption in the distal nephron during inhibition of renal 11β-HSD in vivo, strongly support the proposal that 11β-HSD normally prevents endogenous glucocorticoid from exerting mineralocorticoid-like effects.
Influence of β1-adrenoceptor blockade on the gene expression of adenylate cyclase subtypes and β-adrenoceptor kinase in human atrium
Authors: WANG Tao , BROWN Morris J.
Published: August 2015

There is accumulating evidence of cross-regulation between stimulatory G-protein (Gs)-coupled receptors in human atrium. Chronic β1-adrenoceptor antagonist treatment can sensitize β2-adrenoreceptors, 5-HT4 receptors, histamine H2 receptors and possibly β4-adrenoreceptors. To investigate the mechanism of such cross-talk between Gs-coupled receptors, we have measured the mRNA expression of the cardiac adenylate cyclases (types IV, V, VI and VII) and of β-adrenoceptor kinase 1 in human atria using TaqMan quantitative PCR, and compared expression between patients chronically receiving a β1-adrenoceptor antagonist and non-treated patients. The results showed no difference in gene expression between the two groups of patients; however, significant positive correlations of gene expression between adenylate cyclase subtypes were found. We conclude that β1-adrenoceptor antagonists do not change the absolute levels of gene expression of adenylate cyclase subtypes in human atrium. The co-ordinate regulation among adenylate cyclase subtypes and the influence of β1-adrenoceptor antagonists need to be investigated further.
Increased DNA oxidative susceptibility without increased plasma LDL oxidizability in Type II diabetes: effects of α-tocopherol supplementation
Authors: SAMPSON Michael J. , ASTLEY Sian , RICHARDSON Tristan , WILLIS Gavin , DAVIES Isabel R. , HUGHES David A. , SOUTHON Sue
Published: August 2015

In vivo supplementation studies of the antioxidant α-tocopherol in human Type II diabetes have used surrogate, rather than direct, markers of oxidative damage/antioxidant protection and have used higher doses of α-tocopherol than used in coronary secondary prevention trials. We tested the hypothesis that oral α-tocopherol in a dosage regimen used in secondary prevention trials would reduce directly observed oxidatively induced single-strand breaks in lymphocyte DNA in Type II diabetes. We studied 40 people with Type II diabetes and 30 controls in a randomized, double-blind, placebo-controlled trial of 400 i.u. of oral α-tocopherol daily for 8 weeks. Lymphocyte DNA single-strand breaks and low-density lipoprotein (LDL) particle size and oxidizability were measured at baseline, after 8 weeks, and after 4 weeks washout. Polymorphisms in the gene for the antioxidant enzyme paraoxonase-1 gene (position 192) were measured. The diabetics had increased DNA oxidative susceptibility (P = 0.008), without increased LDL oxidative susceptibility. There was a direct relationship between DNA oxidative susceptibility and baseline plasma α-tocopherol in the diabetes group alone (r = 0.421, r2 = 0.177 and P = 0.023), but DNA and LDL oxidative susceptibility were not influenced by α-tocopherol supplementation in either group in this regimen. Paraoxonase-1 gene polymorphisms did not contribute to LDL or DNA oxidative susceptibility or response to α-tocopherol. Increased DNA oxidative susceptibility, therefore, can occur in Type II diabetes without increased LDL oxidative susceptibility, but α-tocopherol supplementation in this regimen has no influence on DNA or LDL oxidative susceptibility in Type II diabetes or controls. Polymorphisms in the paraoxonase gene (position 192) are not associated with differences in oxidative susceptibility or responses to α-tocopherol.
Haemodynamics of the pressor effect of oral water in human sympathetic denervation due to autonomic failure
Authors: CARIGA P. , MATHIAS C. J.
Published: August 2015

Oral water ingestion increases blood pressure in normal elderly subjects and in patients suffering from autonomic failure, but the time course of the haemodynamic changes is not known. We therefore studied 14 subjects with documented sympathetic denervation due to pure autonomic failure, with continuous haemodynamic recordings obtained before and after ingestion of 500 ml of distilled water at room temperature. The time course of changes in values of systolic and diastolic beat-by-beat finger blood pressure, heart rate, stroke volume, cardiac output, ejection fraction and total peripheral resistance were analysed. Systolic blood pressure rose from 115±8 mmHg (mean±S.E.M.) to 133±8 mmHg (P < 0.001), and diastolic blood pressure from 64±4 to 73±4 mmHg (P < 0.001), with the pressor response beginning a few minutes after water ingestion, plateauing between 10 and 35 min (peak at 14 min), and returning to baseline at 50 min. Heart rate fell from 71±2.5 to 67±2 beats/min (P < 0.001), and total peripheral resistance increased from 1.31±0.19 to 1.61±0.24 m-units (P < 0.001). There were no significant changes in ejection fraction, stroke volume or cardiac output. This study confirmed a pressor response to oral water in subjects with sympathetic denervation. The temporal profile of the response did not favour reflexly mediated sympathetic activation. As subjects with autonomic failure are prone to salt and water depletion, and since blood pressure is exquisitely sensitive to such changes, it may be that the observed response is due to repletion or restoration of intravascular and extravascular fluid volume.
Carbon dioxide, critical closing pressure and cerebral haemodynamics prior to vasovagal syncope in humans
Authors: CAREY Brian J. , EAMES Penelope J. , PANERAI Ronney B. , POTTER John F.
Published: August 2015

The cerebrovascular changes that occur prior to vasovagal syncope (VVS) are unclear, with both increases and decreases in cerebrovascular resistance being reported during pre-syncope. This study assessed the cerebrovascular responses, and their potential underlying mechanisms, that occurred before VVS induced by head-up tilt (HUT). Groups of 65 normal subjects with no previous history of syncope and of 16 patients with recurrent VVS were subjected to 70° HUT for up to 30min. Bilateral middle cerebral artery (MCA) cerebral blood flow velocities (CBFVs) were measured using transcranial Doppler ultrasound, along with simultaneous measures of MCA blood pressure, heart rate, and end-tidal and transcutaneous carbon dioxide concentrations. All 16 patients and 14 of the control subjects developed VVS during HUT. During pre-syncope, mean CBFV declined, due predominantly to a decrease in diastolic rather than systolic CBFV (decreases of 44.5±;19.8% and 6.3±;12.9% respectively; P < 0.0001). CO2 levels and indices of cerebrovascular resistance decreased during pre-syncope, while critical closing pressure (CrCP) increased to levels approaching MCA diastolic blood pressure before decreasing precipitously on syncope. Pre-syncopal changes were similar in syncopal patients and syncopal controls. CrCP, therefore, rises during pre-syncope, possibly related to progressive hypocapnia, and may account for the relatively greater fall in diastolic CBFV. Falls in cerebrovascular resistance, therefore, may be offset by rises in CrCP due to hypocapnia, leading to diminished cerebral blood flow during pre-syncope.
Electrophysiological and biochemical changes in rabbit hearts stored at 4 °C for 6 or 24 h
Authors: FAGBEMI Oluwole S. , BRACK Kieran , GOLAR Surjit , CRISP David , ECONOMIDES Apollo
Published: August 2015

This study examines the electrophysiological and metabolic changes that occur in rabbit hearts during hypothermic storage in vitro. Hearts were microperfused at 4°C for 6 or 24h with either normal Krebs-Henseleit buffer (KHB) or KHB containing 2,3-butanedione monoxime (BDM). After hypothermic storage, hearts were rewarmed to 37°C with KHB. Cardiac function was then assessed in Langendorff perfusion mode. Electrophysiological changes were also assessed from the ventricular paced-evoked responses. After storage, mitochondria were isolated from the hearts and their respiratory control ratio, rate of ATP synthesis and outer membrane intactness were assessed. Compared with values from fresh non-stored hearts, hearts stored hypothermically for 24h showed significant decreases in both left ventricular developed pressure and coronary flow when reperfused in Langendorff mode. On the other hand, the decrease in left ventricular developed pressure in hearts that were stored for only 6h (with or without BDM) was not significant. Compared with values obtained from fresh non-stored hearts, hypothermic storage significantly decreased the R-wave amplitude, and both the R-E and ST-E intervals of paced-evoked responses. This was true for hearts microperfused for 6h (with or without BDM) and for hearts microperfused with buffer containing BDM for 24h. The ST-R intervals in hearts microperfused hypothermically for 6h were prolonged, but this change was not statistically significant compared with those obtained from unstored hearts. In hearts microperfused with KHB containing BDM for 24h, the ST-R interval was significantly prolonged. Hypothermic microperfusion for 24h significantly decreased both the mitochondrial coupling ratio and the rate of ATP synthesis. In hearts microperfused with BDM for 6h, mitochondrial coupling ratios and the rate of ATP synthesis were not significantly different from those in fresh hearts. In conclusion, the present study has shown that long-term hypothermic storage significantly impaired both paced-evoked responses and mitochondrial function. Inclusion of BDM in the perfusion buffer during storage significantly ameliorated some of these changes.
Spontaneously hypertensive rat resistance artery structure related to myogenic and mechanical properties
Authors: BUND Stuart J.
Published: August 2015

This investigation related arterial structure to myogenic (pressure-dependent) contractile responses in resistance arteries from spontaneously hypertensive rats (SHRs) and Wistar-Kyoto (WKY) normotensive control rats under pressurized conditions in vitro. Femoral and mesenteric resistance arteries from either strain were cannulated and pressurized in an arteriograph for the determination of pressure-diameter relationships under passive and active conditions in the range 5-200mmHg transmural pressure. Arterial geometrical measurements were made under relaxed conditions at 100mmHg. Media thickness/lumen diameter (M/L) ratios were significantly increased in SHR femoral (5.00±0.44% compared with 3.63±0.34%; P<0.05) and mesenteric (4.40±0.29% compared with 2.62±0.23%; P<0.001) arteries compared with those from WKY rats. Maximum myogenic contractions, assessed as minimum normalized diameters, were not significantly different in SHR and WKY rat femoral (0.41±0.03 and 0.40±0.02 respectively) or mesenteric (0.56±0.02 and 0.63±0.03 respectively) arteries. Arterial mechanical analyses demonstrated that incremental elastic modulus is reduced in SHR mesenteric arteries, but is not significantly different in SHR femoral arteries, compared with those from WKY rats. Additionally, wall stress at estimated in vivo pressures under passive and active conditions are similar in SHR and WKY rat arteries. These data demonstrate that increased M/L ratios in resistance arteries from SHRs are not associated with increased maximum pressure-dependent contractile responses. Increased M/L ratios in resistance arteries from SHRs are not accounted for by increased vessel wall stiffness, but the hypertension-associated arterial geometrical abnormalities act to normalize wall stress in the face of increased arterial pressure.