The evidence for histamine H3 receptor-mediated endothelium-dependent relaxation in isolated rat aorta

The presence of histamine H3 receptors was evaluated on the rat aorta endothelium. In the presence of pyrilamine (1 nM, 7 nM, 10 nM) or thioperamide (1 nM, 10 nM, 30 nM) the concentration–response curve for histamine-induced (0.1 nM − 0.01 mM) endothelium-dependent rat aorta relaxation was shifted to the right without significant change of the Emax indicating competitive antagonism by pyrilamine (pA2 = 9.33 ± 0.34, slope = 1.09 ± 0.36) or thioperamide (pA2 =9.31 ± 0.16, slope=0.94 ± 0.10). Cimetidine (1 μM) did not influence histamine-induced endothelium-dependent rat aorta relaxation. In the presence of thioperamide (1 nM, 10 nM, 30 nM) the concentration–response curve for (R)α-MeHA-induced (0.1 nM − 0.01 mM) endothelium-dependent relaxation was shifted to the right without significant change of Emax indicated competitive antagonism by thioperamide (pA2 = 9.21 ± 0.4, slope = 1.03 ± 0.35). Pyrilamine (100 nM) or cimetidine (1 μM) did not influence (R)α-MeHA-induced endothelium-dependent rat aorta relaxation. These results suggest the presence of a heterogenous population of histamine receptors, H1 and H3, on rat aorta endothelium

Endothelium-dependent relaxation of rat aorta to a histamine H3 agonist is reduced by inhibitors of nitric oxide synthase, guanylate cyclase and Na+,K+-ATPase

The possible involvement of different effector systems (nitric oxide synthase, guanylate cyclase, β-adrenergic and muscarinic cholinergic receptors, cyclooxygenase and lipoxygenase, and Na+,K+-ATPase) was evaluated in a histamine H3 receptor agonist-induced ((R)α-methylhistamine, (R)α-MeHA) endothelium-dependent rat aorta relaxation assay. (R)α-MeHA (0.1 nM – 0.01 mM) relaxed endothelium-dependent rat aorta, with a pD2 value of 8.22 ± 0.06, compared with a pD2 value of 7.98 ± 0.02 caused by histamine (50% and 70% relaxation, respectively). The effect of (R)α-MeHA (0.1 nM – 0.01 mM) was competitively antagonized by thioperamide (1, 10 and 30 nM) (pA2 = 9.21 ± 0.40; slope = 1.03 ± 0.35) but it was unaffected by pyrilamine (100 nM), cimetidine (1 μM), atropine (10 μM), propranolol (1 μM), indomethacin (10 μM) or nordthydroguaiaretic acid (0.1 mM). Inhibitors of nitric oxide synthase, L-NG-monomethylarginine (L-NMMA, 10 μM) and NG-nitro-L-arginine methylester (L-NOARG, 10 μM) inhibited the relaxation effect of (R)α-MeHA, by approximately 52% and 70%, respectively). This inhibitory effect of L-NMMA was partially reversed by L-arginine (10 μM). Methylene blue (10 μM) and ouabain (10 μM) inhibited relaxation (R)α-MeHA-induced by approximately 50% and 90%, respectively. The products of cyclooxygenase and lipoxygenase are not involved in (R)α-MeHA-induced endothelium-dependent rat aorta relaxation nor are the muscarinic cholinergic and β-adrenergic receptors. The results also suggest the involvement of NO synthase, guanylate cyclase and Na+,K+-ATPase in (R)α-MeHA-induced endothelium-dependent rat aorta relaxation

Bacterial iron-oxide nanowires from biofilm waste as a new adsorbent for the removal of arsenic from water

Biofilm, generated by the bacteria in the groundwater pumping system pipelines of the Salt Interception Scheme on the River Murray in South Australia is discarded as a waste material accumulated after periodic cleaning of the pipes. Structural and chemical composition characterizations confirm that this waste material is composed of amorphous twisted iron-oxide nanowires (ION), generated by bacteria, and they have a unique structure and properties. The adsorption performance of these iron-oxide nanowires for arsenic removal from water was evaluated to define their adsorption capacity for As(III) and As(V) and kinetics. Obtained results demonstrate considerable adsorption properties of this waste biological material and suggest its promising application as a new and low-cost adsorbent for water treatment.Ivan Andjelkovic, Sara Azari, Mason Erkelens, Peter Forward, Martin F. Lambert and Dusan Losi

Influence of Higenamine on Exercise Performance of Recreational Female Athletes: A Randomized Double-Blinded Placebo-Controlled Trial

The aim of this study was to determine the ergogenic effects and the safety profile of a one-component higenamine supplement in female recreational athletes. Twelve recreational female basketball players (age 29–41 years, oxygen consumption (VO2max) > 30 ml⋅kg–1⋅min–1, with training > 5 h wk–1) were randomized either to the higenamine group, or to the placebo group for 3 weeks. In order to determine ergogenic effects and safety profile of higenamine administration, we assessed the following variables before and after 3 weeks of supplementation: anthropometric parameters, resting metabolic rate (RMR), exercise testing variables, serum free fatty acids (FFAs), blood pressure, enzyme activity, urea, lipid profile, and complete blood count. There were no differences between groups in anthropometric parameters, including basal metabolic rate (BMR), RMR and body fat [p = 0.706 (Cohen’s d 0.223), p = 0.169 (Cohen’s d 0.857), and p = 0.223 (Cohen’s d 0.750), respectively], FFAs [0.43 ± 0.03 vs. 0.54 ± 0.23, p = 0.206 (Cohen’s d 0.540)], neither significant differences in cardiopulmonary parameters after the intervention period. Furthermore, all measured outcome variables in the safety assessment were not significant, with values remaining stable during the intervention period for participants in both groups. This is the first study to document the effects and the safety profile of higenamine-based dietary supplements at a specified dose in female recreational athletes. Our data indicate that 21-day of supplementation with 75 mg higenamine would not result in improving cardiopulmonary exercise fitness and weight loss in female recreational athletes. Moreover, supplementation with 75 mg higenamine is safe and well-tolerated in younger recreational female athletes

IKKβ regulates essential functions of the vascular endothelium through kinase-dependent and -independent pathways

Vascular endothelium provides a selective barrier between the blood and tissues, participates in wound healing and angiogenesis, and regulates tissue recruitment of inflammatory cells. Nuclear factor (NF)-κB transcription factors are pivotal regulators of survival and inflammation, and have been suggested as potential therapeutic targets in cancer and inflammatory diseases. Here we show that mice lacking IKKβ, the primary kinase mediating NF-κB activation, are smaller than littermates and born at less than the expected Mendelian frequency in association with hypotrophic and hypovascular placentae. IKKβ-deleted endothelium manifests increased vascular permeability and reduced migration. Surprisingly, we find that these defects result from loss of kinase-independent effects of IKKβ on activation of the serine-threonine kinase, Akt. Together, these data demonstrate essential roles for IKKβ in regulating endothelial permeability and migration, as well as an unanticipated connection between IKKβ and Akt signalling

Perifosine as a Potential Novel Anti-Cancer Agent Inhibits EGFR/MET-AKT Axis in Malignant Pleural Mesothelioma

PI3K/AKT signalling pathway is aberrantly active and plays a critical role for cell cycle progression of human malignant pleural mesothelioma (MMe) cells. AKT is one of the important cellular targets of perifosine, a novel bio-available alkylphospholipid that has displayed significant anti-proliferative activity in vitro and in vivo in several human tumour model systems and is currently being tested in clinical trials.We tested Perifosine activity on human mesothelial cells and different mesothelioma cell lines, in order to provide evidence of its efficacy as single agent and combined therapy.We demonstrate here that perifosine, currently being evaluated as an anti-cancer agent in phase 1 and 2 clinical trials, caused a dose-dependent reduction of AKT activation, at concentrations causing MMe cell growth arrest. In this study we firstly describe that MMe cells express aside from AKT1 also AKT3 and that either the myristoylated, constitutively active, forms of the two proteins, abrogated perifosine-mediated cell growth inhibition. Moreover, we describe here a novel mechanism of perifosine that interferes, upstream of AKT, affecting EGFR and MET phosphorylation. Finally, we demonstrate a significant increase in cell toxicity when MMe cells were treated with perifosine in combination with cisplatin.This study provides a novel mechanism of action of perifosine, directly inhibiting EGFR/MET-AKT1/3 axis, providing a rationale for a novel translational approach to the treatment of MMe

An AP-MS- and BioID-compatible MAC-tag enables comprehensive mapping of protein interactions and subcellular localizations

Protein-protein interactions govern almost all cellular functions. These complex networks of stable and transient associations can be mapped by affinity purification mass spectrometry (AP-MS) and complementary proximity-based labeling methods such as BioID. To exploit the advantages of both strategies, we here design and optimize an integrated approach combining AP-MS and BioID in a single construct, which we term MAC-tag. We systematically apply the MAC-tag approach to 18 subcellular and 3 sub-organelle localization markers, generating a molecular context database, which can be used to define a protein's molecular location. In addition, we show that combining the AP-MS and BioID results makes it possible to obtain interaction distances within a protein complex. Taken together, our integrated strategy enables the comprehensive mapping of the physical and functional interactions of proteins, defining their molecular context and improving our understanding of the cellular interactome.Peer reviewe

Activation of Akt by the Bacterial Inositol Phosphatase, SopB, is Wortmannin Insensitive

Salmonella enterica uses effector proteins translocated by a Type III Secretion System to invade epithelial cells. One of the invasion-associated effectors, SopB, is an inositol phosphatase that mediates sustained activation of the pro-survival kinase Akt in infected cells. Canonical activation of Akt involves membrane translocation and phosphorylation and is dependent on phosphatidyl inositide 3 kinase (PI3K). Here we have investigated these two distinct processes in Salmonella infected HeLa cells. Firstly, we found that SopB-dependent membrane translocation and phosphorylation of Akt are insensitive to the PI3K inhibitor wortmannin. Similarly, depletion of the PI3K regulatory subunits p85α and p85ß by RNAi had no inhibitory effect on SopB-dependent Akt phosphorylation. Nevertheless, SopB-dependent phosphorylation does depend on the Akt kinases, PDK1 and rictor-mTOR. Membrane translocation assays revealed a dependence on SopB for Akt recruitment to Salmonella ruffles and suggest that this is mediated by phosphoinositide (3,4) P2 rather than phosphoinositide (3,4,5) P3. Altogether these data demonstrate that Salmonella activates Akt via a wortmannin insensitive mechanism that is likely a class I PI3K-independent process that incorporates some essential elements of the canonical pathway