Kinin B1 receptor homo-oligomerization is required for receptor trafficking to the cell surface.

The kinin B1 receptor (B1R) is a G protein-coupled receptor with pro-inflammatory activity that is latent in healthy tissues but induced by tissue insult. Here, we investigated if B1R homo-oligomerization is a possible mechanism regulating the presentation of this receptor at the level of maturation and trafficking to the cell surface. To this end, we used HEK293 cells stably expressing N-terminal FLAG and HA epitope-tagged wild-type human B1R and an N-terminal receptor fragment, B1stop135, which terminates at the C-terminal end of the third transmembrane domain and has previously been shown to oligomerize with B1R. Receptors were monitored by immunoblotting and immunoprecipitation, receptor function by agonist binding and agonist-promoted phosphoinositide hydrolysis, and receptor trafficking by confocal immunofluorescence microscopy. When expressed alone, B1R is core N-glycosylated and forms oligomers localized intracellularly and on the cell surface. B1stop135 also exists as core N-glycosylated oligomers but is localized exclusively intracellularly. When co-expressed, B1stop135 prevents specifically B1R homo-oligomerization by forming nonfunctional B1R-B1stop135 hetero-oligomers, retains B1R intracellularly at least in part in the endoplasmatic reticulum (ER), increases calnexin binding to the receptor, and increases receptor degradation. We conclude that B1R homo-oligomerization is necessary for B1R maturation and trafficking to the cell surface. Modulating this mechanism may be a novel therapeutic avenue in inflammatory disease

Lönsam mjölkproduktion

Ravelsmarks gård have now reached the stage when there are few options for the future. The options are either a construction of completely new farm buildings or a winding up of the milk production. Present on the farm today there is 65 dairy cows plus recruitment. 44 cows stand in long-stalls and 20 cows stand in short-stalls. The objective of this study was to examine the profitability of a completely new cowshed. In order to get costs for investments different milking systems and equipment for feedstuff have been compared. The systems in the investment budget have been chosen on recommendations from advisers, salesmen and on the basis on what is suitable for the farm. One system where a so-called mix feeder wagon is used for the roughage seemed to be best suited for Ravelsmark. The different prices obtained for the manure well show that there are possibilities to decrease the investment costs. By using the rubber cloth from MPG one could save about 300 000 crowns. In the summarizing working plan one can see that the alternative with milking parlour can cope better with decreased milk yield and decreased milk price than the alternative with milk robot. The cost estimates for the robot cannot cope with a milk yield of 9500 litres and a milk price of 2.70 crowns. It differs approximately 200 000 crowns in annual profit between the milking parlour alternative and the robot alternative.Ravelsmarks gård befinner sig i dag i ett skede då det inte finns så många alternativ inför framtiden. Antingen är det en nybyggnation som gäller eller på sikt en avveckling av mjölkproduktionen. På gården finns det 65 mjölkkor samt ungdjur. 44 kor står på långbås och 20 kor står på kortbås. Målet med detta examensarbete var att undersöka hur lönsamheten skulle kunna bli med ett nybyggt stall. För att få reda på investerings kostnader har jag tittat på olika mjölkningssystem och utrustning för utfodring. De val av system som är med i investeringsbudgeten, har valts utifrån synpunkter från rådgivare, säljare och vad som passar till Ravelsmark. Ett system där man använder en såkallad mixfeeder för att utfodra grovfoder, verkar kunna passa bäst in på Ravelsmark. Med detta system får man möjlighet att utfodra olika djurgrupper på ett bra sätt. De prisuppgifterna jag fick på gödselbrunnar, visar att det finns möjligheter att få ned investeringskostnaden. Genom att använda sig av MPGs tätskikts duk skulle man kunna spara ca 300 000 kr. I sammanställningen av driftsplanen ser man att alternativet med mjölkgrop klarar minskad avkastning och ett sänkt mjölkpris bättre än alternativet med en mjölkrobot. Vid en avkastning på 9500 liter mjölk och ett mjölk pris på 2,70 kr går inte robot kalkylen ihop. Det skiljer ca 200 000 kr i resultat mellan de båda alternativen i de olika scenarion som valts

Binding of von Willebrand factor by coagulase-negative staphylococci

Coagulase-negative staphylococci (CNS) are the most common infectious micro-organisms isolated from prosthetic devices. To determine whether von Willebrand factor (vWF) acts as an adhesin in bacterial recognition, bacterial binding of recombinant vWF (rvWF) was studied. Eleven CNS strains, belonging to S. epidermidis, S. haemolyticus and S. hominis species, bound soluble rvWF, but to a lesser extent than S. aureus. S. epidermidis strain H2-W bound 125I-labelled rvWF in a dose-dependent manner. The binding could be inhibited by unlabelled rvWF and thrombospondin, but not by fibrinogen, vitronectin or the carbohydrates N-acetylgalactoseamine, d-galactose, d-glucose, and d-fucose. Pre-incubation of rvWF with type I collagen and Arg-Gly-Asp-Ser (RGDS) peptides did not inhibit binding, whereas pre-incubation of rvWF with heparin decreased binding significantly. The interaction between CNS and rvWF was sensitive to proteinase treatment of bacterial cells. CNS strains bound to immobilised rvWF an extent greater or equal to the positive control strain S. aureus Cowan I. rvWF binding structures from bacterial cell wall were detected by immunoblot. Cowan I strain had 140-, 90- and 38-kDa binding molecules. S. haemolyticus strain SM131 and S. epidermidis strain H2-W had two (120 and 60 kDa) and five (120, 90, 60, 52 and 38 kDa) binding molecules, respectively. Similar binding structures were formed when cell wall extracts from these strains were incubated with thrombospondin. These results indicate that specific ligand–receptor interaction between CNS and rvWF may contribute to bacterial adhesion and colonisation on biomaterial surfaces. Heparin-binding domains of rvWF might be the crucial regions for bacterial attachment. rvWF and thrombospondin may recognise similar molecules in staphylococcal cell wall extracts

Overregulation of Health Care: Musings on Disruptive Innovation Theory

Disruptive innovation theory provides one lens through which to describe how regulations may stifle innovation and increase costs. Basing their discussion on this theory, Curtis and Schulman consider some of the effects that regulatory controls may have on innovation in the health sector

Podvertising : om sponsring i ett nytt medieformat

Firstly, the title of this paper is Podvertising: advertising in a new media format (in Swedish: Podvertising: om sponsring i ett nytt medieformat) and the authors are Holger Brundin and Fredrik Bröndum. The paper is authored for Media- and communication studies at The institution for communication and media, Lunds University. The podcast is a fairly new and growing format of media with the internet as its platform of distribution. In comparison to traditional media such as the television or the radio, the podcast is young and the podcast producers are still experimenting with the commercial potential of the format. The commercial podcast had its breakthrough in Sweden as late as 2010. Therefore, no clear rules or guidelines on advertisment exists for the podcast format. Our main question formulations regards how the external sponsors are talked about and presented in the podcast. With this in mind we have chosen a number of theoretical viewpoints to apply on our research and analysis. Our primary theory of choice is the media convergence theory. Thereafter we use theoretical arguments regarding advertorials and study podcasts in the contexts of commerce and history. Methodically, we use a quantitative analysis of content and a text analysis for the qualitative analysis. The results of our analysis, which is predominantly qualitative, shows that the podcast producers often portray their relationship to the sponsor as an amicable one, using words that indicates that they have a close relationship, and the producers prefer to adress their sponsors as friends rather than commercial partners

G protein-coupled Estrogen Receptor 1 (GPER1)/GPR30 Increases ERK1/2 Activity Through PDZ-dependent and -independent Mechanisms

G protein-coupled receptor 30 (GPR30), also called G protein-coupled estrogen receptor 1 (GPER1), is thought to play important roles in breast cancer and cardiometabolic regulation, but many questions remain about ligand activation, effector coupling, and subcellular localization. We showed recently that GPR30 interacts through the C-terminal type I PDZ motif with SAP97 and protein kinase A (PKA)-anchoring protein (AKAP) 5, which anchor the receptor in the plasma membrane and mediate an apparently constitutive decrease in cAMP production independently of Gi/o. Here, we show that GPR30 also constitutively increases ERK1/2 activity. Removing the receptor PDZ motif or knocking down specifically AKAP5 inhibited the increase, showing that this increase also requires the PDZ interaction. However, the increase was inhibited by pertussis toxin (PTX) as well as by wortmannin, but not by AG1478, indicating that Gi/o and phosphoinositide 3-kinase (PI3K) mediate the increase independently of epidermal growth factor receptor (EGFR) transactivation. FK506 and okadaic acid also inhibited the increase, implying that a protein phosphatase is involved. The proposed GPR30 agonist G-1 also increased ERK1/2 activity, but this increase was only observed at a level of receptor expression below that required for the constitutive increase. Furthermore, deleting the PDZ motif did not inhibit the G-1-stimulated increase. Based on these results, we propose that GPR30 increases ERK1/2 activity via two Gi/o-mediated mechanisms; a PDZ-dependent apparently constitutive mechanism, and a PDZ-independent G-1-stimulated mechanism

International Union of Pharmacology. XLV. Classification of the Kinin Receptor Family: from Molecular Mechanisms to Pathophysiological Consequences

Kinins are proinflammatory peptides that mediate numerous vascular and pain responses to tissue injury. Two pharmacologically distinct kinin receptor subtypes have been identified and characterized for these peptides, which are named B1 and B2 and belong to the rhodopsin family of G protein-coupled receptors. The B2 receptor mediates the action of bradykinin (BK) and lysyl-bradykinin (Lys-BK), the first set of bioactive kinins formed in response to injury from kininogen precursors through the actions of plasma and tissue kallikreins, whereas the B(1) receptor mediates the action of des-Arg9-BK and Lys-des-Arg9-BK, the second set of bioactive kinins formed through the actions of carboxypeptidases on BK and Lys-BK, respectively. The B2 receptor is ubiquitous and constitutively expressed, whereas the B1 receptor is expressed at a very low level in healthy tissues but induced following injury by various proinflammatory cytokines such as interleukin-1beta. Both receptors act through G alpha(q) to stimulate phospholipase C beta followed by phosphoinositide hydrolysis and intracellular free Ca2+ mobilization and through G alpha(i) to inhibit adenylate cyclase and stimulate the mitogen-activated protein kinase pathways. The use of mice lacking each receptor gene and various specific peptidic and nonpeptidic antagonists have implicated both B1 and B2 receptors as potential therapeutic targets in several pathophysiological events related to inflammation such as pain, sepsis, allergic asthma, rhinitis, and edema, as well as diabetes and cancer. This review is a comprehensive presentation of our current understanding of these receptors in terms of molecular and cell biology, physiology, pharmacology, and involvement in human disease and drug development

The G Protein-Coupled Estrogen Receptor 1 (GPER1/GPR30) Agonist G-1 Regulates Vascular Smooth Muscle Cell Ca Handling.

The G protein-coupled estrogen receptor GPER1/GPR30 is implicated in blood pressure regulation but the mechanisms are not identified. Here, we hypothesize that GPER1 controls blood pressure by regulating vascular smooth muscle cell Ca(2+) handling. Treatment with the GPER1 agonist G-1 (in the µM concentration range) acutely reduced spontaneous and synchronous Ca(2+) spike activity in A7r5 vascular smooth muscle cells expressing mRNA for GPER1. Furthermore, G-1 (1 µM) attenuated the thromboxane A2 analogue U46619-stimulated Ca(2+) spike activity but had no effect on the U46619-induced increase in the basal level of Ca(2+). The voltage-sensitive L-type Ca(2+) channel blocker nifedipine (100 nM) reduced Ca(2+) spike activity similar to G-1. Pharmacological, but not physiological, concentrations of the estrogen 17β-estradiol reduced Ca(2+) spike activity. The GPER1 antagonist G-15 blocked G-1-induced downregulation of Ca(2+) spike activity, supporting a GPER1-dependent mechanism. G-1 (1 µM) and nifedipine (100 nM) attenuated the 30-mM KCl-evoked rise in intracellular Ca(2+) concentration, suggesting that G-1 blocks inflow of Ca(2+) via voltage-sensitive Ca(2+) channels. In conclusion, we demonstrate that the GPER1 agonist G-1 regulates vascular smooth muscle cell Ca(2+) handling by lowering Ca(2+) spike activity, suggesting a role for this mechanism in GPER1-mediated control of blood pressure. © 2013 S. Karger AG, Basel