Electrical Pulse Stimulation of Cultured Human Skeletal Muscle Cells as an In Vitro Model of Exercise

Background and Aims Physical exercise leads to substantial adaptive responses in skeletal muscles and plays a central role in a healthy life style. Since exercise induces major systemic responses, underlying cellular mechanisms are difficult to study in vivo. It was therefore desirable to develop an in vitro model that would resemble training in cultured human myotubes. Methods Electrical pulse stimulation (EPS) was applied to adherent human myotubes. Cellular contents of ATP, phosphocreatine (PCr) and lactate were determined. Glucose and oleic acid metabolism were studied using radio-labeled substrates, and gene expression was analyzed using real-time RT-PCR. Mitochondrial content and function were measured by live imaging and determination of citrate synthase activity, respectively. Protein expression was assessed by electrophoresis and immunoblotting. Results High-frequency, acute EPS increased deoxyglucose uptake and lactate production, while cell contents of both ATP and PCr decreased. Chronic, low-frequency EPS increased oxidative capacity of cultured myotubes by increasing glucose metabolism (uptake and oxidation) and complete fatty acid oxidation. mRNA expression level of pyruvate dehydrogenase complex 4 (PDK4) was significantly increased in EPS-treated cells, while mRNA expressions of interleukin 6 (IL-6), cytochrome C and carnitin palmitoyl transferase b (CPT1b) also tended to increase. Intensity of MitoTracker®Red FM was doubled after 48 h of chronic, low-frequency EPS. Protein expression of a slow fiber type marker (MHCI) was increased in EPS-treated cells. Conclusions Our results imply that in vitro EPS (acute, high-frequent as well as chronic, low-frequent) of human myotubes may be used to study effects of exercise.This work was funded by the University of Oslo, Oslo University College, the Norwegian Diabetes Foundation, the Freia Chocolade Fabriks Medical Foundation and the Anders Jahre’s Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Prostacyclin reverses platelet stress fibre formation causing platelet aggregate instability

Prostacyclin (PGI2) modulates platelet activation to regulate haemostasis. Evidence has emerged to suggest that thrombi are dynamic structures with distinct areas of differing platelet activation. It was hypothesised that PGI2 could reverse platelet spreading by actin cytoskeletal modulation, leading to reduced capability of platelet aggregates to withstand a high shear environment. Our data demonstrates that post-flow of PGI2 over activated and spread platelets on fibrinogen, identified a significant reduction in platelet surface area under high shear. Exploration of the molecular mechanisms underpinning this effect revealed that PGI2 reversed stress fibre formation in adherent platelets, reduced platelet spreading, whilst simultaneously promoting actin nodule formation. The effects of PGI2 on stress fibres were mimicked by the adenylyl cyclase activator forskolin and prevented by inhibitors of protein kinase A (PKA). Stress fibre formation is a RhoA dependent process and we found that treatment of adherent platelets with PGI2 caused inhibitory phosphorylation of RhoA, reduced RhoA GTP-loading and reversal of myosin light chain phosphorylation. Phospho-RhoA was localised in actin nodules with PKA type II and a number of other phosphorylated PKA substrates. This study demonstrates that PGI2 can reverse key platelet functions after their initial activation and identifies a novel mechanism for controlling thrombosis

IgE allergy diagnostics and other relevant tests in allergy, a World Allergy Organization position paper

Currently, testing for immunoglobulin E (IgE) sensitization is the cornerstone of diagnostic evaluation in suspected allergic conditions. This review provides a thorough and updated critical appraisal of the most frequently used diagnostic tests, both in vivo and in vitro. It discusses skin tests, challenges, and serological and cellular in vitro tests, and provides an overview of indications, advantages and disadvantages of each in conditions such as respiratory, food, venom, drug, and occupational allergy. Skin prick testing remains the first line approach in most instances; the added value of serum specific IgE to whole allergen extracts or components, as well as the role of basophil activation tests, is evaluated. Unproven, non-validated, diagnostic tests are also discussed. Throughout the review, the reader must bear in mind the relevance of differentiating between sensitization and allergy; the latter entails not only allergic sensitization, but also clinically relevant symptoms triggered by the culprit allergen.info:eu-repo/semantics/publishedVersio

Electronic and atomic structures of the Sr3Ir4Sn13 single crystal: A possible charge density wave material

[[abstract]]X-ray scattering (XRS), x-ray absorption near-edge structure (XANES) and extended x-ray absorption fine structure (EXAFS) spectroscopic techniques were used to study the electronic and atomic structures of the high-quality Sr3Ir4Sn13 (SIS) single crystal below and above the transition temperature (T* ≈ 147 K). The evolution of a series of modulated satellite peaks below the transition temperature in the XRS experiment indicated the formation of a possible charge density wave (CDW) in the (110) plane. The EXAFS phase derivative analysis supports the CDW-like formation by revealing different bond distances [Sn1(2)-Sn2] below and above T* in the (110) plane. XANES spectra at the Ir L3-edge and Sn K-edge demonstrated an increase (decrease) in the unoccupied (occupied) density of Ir 5d-derived states and a nearly constant density of Sn 5p-derived states at temperatures T < T* in the (110) plane. These observations clearly suggest that the Ir 5d-derived states are closely related to the anomalous resistivity transition. Accordingly, a close relationship exists between local electronic and atomic structures and the CDW-like phase in the SIS single crystal.[[notice]]補正完