33 research outputs found
Conformational Changes and Slow Dynamics through Microsecond Polarized Atomistic Molecular Simulation of an Integral Kv1.2 Ion Channel
Structure and dynamics of voltage-gated ion channels, in particular the motion of
the S4 helix, is a highly interesting and hotly debated topic in current
membrane protein research. It has critical implications for insertion and
stabilization of membrane proteins as well as for finding how transitions occur
in membrane proteins—not to mention numerous applications in drug
design. Here, we present a full 1 µs atomic-detail molecular dynamics
simulation of an integral Kv1.2 ion channel, comprising 120,000 atoms. By
applying 0.052 V/nm of hyperpolarization, we observe structural rearrangements,
including up to 120° rotation of the S4 segment, changes in
hydrogen-bonding patterns, but only low amounts of translation. A smaller
rotation (∼35°) of the extracellular end of all S4 segments is
present also in a reference 0.5 µs simulation without applied field,
which indicates that the crystal structure might be slightly different from the
natural state of the voltage sensor. The conformation change upon
hyperpolarization is closely coupled to an increase in 310 helix
contents in S4, starting from the intracellular side. This could support a model
for transition from the crystal structure where the hyperpolarization
destabilizes S4–lipid hydrogen bonds, which leads to the helix
rotating to keep the arginine side chains away from the hydrophobic phase, and
the driving force for final relaxation by downward translation is partly
entropic, which would explain the slow process. The coordinates of the
transmembrane part of the simulated channel actually stay closer to the recently
determined higher-resolution Kv1.2 chimera channel than the starting structure
for the entire second half of the simulation (0.5–1 µs).
Together with lipids binding in matching positions and significant thinning of
the membrane also observed in experiments, this provides additional support for
the predictive power of microsecond-scale membrane protein simulations
Dampened antiviral immunity to intravaginal exposure to RNA viral pathogens allows enhanced viral replication.
Understanding the host immune response to vaginal exposure to RNA viruses is required to combat sexual transmission of this class of pathogens. In this study, using lymphocytic choriomeningitis virus (LCMV) and Zika virus (ZIKV) in wild-type mice, we show that these viruses replicate in the vaginal mucosa with minimal induction of antiviral interferon and inflammatory response, causing dampened innate-mediated control of viral replication and a failure to mature local antigen-presenting cells (APCs). Enhancement of innate-mediated inflammation in the vaginal mucosa rescues this phenotype and completely inhibits ZIKV replication. To gain a better understanding of how this dampened innate immune activation in the lower female reproductive tract may also affect adaptive immunity, we modeled CD8 T cell responses using vaginal LCMV infection. We show that the lack of APC maturation in the vaginal mucosa leads to a delay in CD8 T cell activation in the draining lymph node and hinders the timely appearance of effector CD8 T cells in vaginal mucosa, thus further delaying viral control in this tissue. Our study demonstrates that vaginal tissue is exceptionally vulnerable to infection by RNA viruses and provides a conceptual framework for the male to female sexual transmission observed during ZIKV infection
Phagocytosis of live and dead Escherichia coli and Staphylococcus aureus in human whole blood is markedly reduced by combined inhibition of C5aR1 and CD14
Air Bubbles Activate Complement and Trigger Hemostasis and C3-Dependent Cytokine Release Ex Vivo in Human Whole Blood
Venous air embolism, which may complicate medical and surgical procedures, activates complement and triggers thromboinflammation. In lepirudin-anticoagulated human whole blood, we examined the effect of air bubbles on complement and its role in thromboinflammation. Whole blood from 16 donors was incubated with air bubbles without or with inhibitors of C3, C5, C5aR1, or CD14. Complement activation, hemostasis, and cytokine release were measured using ELISA and quantitative PCR. Compared with no air, incubating blood with air bubbles increased, on average, C3a 6.5-fold, C3bc 6-fold, C3bBbP 3.7-fold, C5a 4.6-fold, terminal complement complex sC5b9 3.6-fold, prothrombin fragments 1+2 (PTF1+2) 25-fold, tissue factor mRNA (TF-mRNA) 26-fold, microparticle tissue factor 6.1-fold, β-thromboglobulin 26-fold (all p < 0.05), and 25 cytokines 11-fold (range, 1.5–78-fold; all p < 0.0001). C3 inhibition attenuated complement and reduced PTF1+2 2-fold, TF-mRNA 5.4-fold, microparticle tissue factor 2-fold, and the 25 cytokines 2.7-fold (range, 1.4–4.9-fold; all p < 0.05). C5 inhibition reduced PTF1+2 2-fold and TF-mRNA 12-fold (all p < 0.05). C5 or CD14 inhibition alone reduced three cytokines, including IL-1β (p = 0.02 and p = 0.03). Combined C3 and CD14 inhibition reduced all cytokines 3.9-fold (range, 1.3–9.5-fold; p < 0.003) and was most pronounced for IL-1β (3.2- versus 6.4-fold), IL-6 (2.5- versus 9.3-fold), IL-8 (4.9- versus 8.6-fold), and IFN-γ (5- versus 9.5-fold). Antifoam activated complement and was avoided. PTF1+2 was generated in whole blood but not in plasma. In summary, air bubbles activated complement and triggered a C3-driven thromboinflammation. C3 inhibition reduced all mediators, whereas C5 inhibition reduced only TF-mRNA. Combined C5 and CD14 inhibition reduced IL-1β release. These data have implications for future mechanistic studies and possible pharmacological interventions in patients with air embolism
Air Bubbles Activate Complement and Trigger Hemostasis and C3-Dependent Cytokine Release Ex Vivo in Human Whole Blood
Venous air embolism, which may complicate medical and surgical procedures, activates complement and triggers thromboinflammation. In lepirudin-anticoagulated human whole blood, we examined the effect of air bubbles on complement and its role in thromboinflammation. Whole blood from 16 donors was incubated with air bubbles without or with inhibitors of C3, C5, C5aR1, or CD14. Complement activation, hemostasis, and cytokine release were measured using ELISA and quantitative PCR. Compared with no air, incubating blood with air bubbles increased, on average, C3a 6.5-fold, C3bc 6-fold, C3bBbP 3.7-fold, C5a 4.6- fold, terminal complement complex sC5b9 3.6-fold, prothrombin fragments 1+2 (PTF1+2) 25-fold, tissue factor mRNA (TF-mRNA) 26- fold, microparticle tissue factor 6.1-fold, b-thromboglobulin 26-fold (all p < 0.05), and 25 cytokines 11-fold (range, 1.5 78-fold; all p < 0.0001). C3 inhibition attenuated complement and reduced PTF1+2 2-fold, TF-mRNA 5.4-fold, microparticle tissue factor 2-fold, and the 25 cytokines 2.7-fold (range, 1.4 4.9-fold; all p < 0.05). C5 inhibition reduced PTF1+2 2-fold and TF-mRNA 12-fold (all p < 0.05). C5 or CD14 inhibition alone reduced three cytokines, including IL-1b (p 5 0.02 and p 5 0.03). Combined C3 and CD14 inhibition reduced all cytokines 3.9-fold (range, 1.3 9.5-fold; p < 0.003) and was most pronounced for IL-1b (3.2- versus 6.4-fold), IL-6 (2.5- versus 9.3-fold), IL-8 (4.9- versus 8.6-fold), and IFN-g (5- versus 9.5-fold). Antifoam activated complement and was avoided. PTF1+2 was generated in whole blood but not in plasma. In summary, air bubbles activated complement and triggered a C3-driven thromboinflammation. C3 inhibition reduced all mediators, whereas C5 inhibition reduced only TF-mRNA. Combined C5 and CD14 inhibition reduced IL-1b release. These data have implications for future mechanistic studies and possible pharmacological interventions in patients with air embolism
Air bubbles activate complement and trigger hemostasis and C3-dependent cytokine release ex vivo in human whole blood
Leucocyte expression of complement C5a receptors exacerbates infarct size after myocardial reperfusion injury
Aims Early reperfusion is mandatory for the treatment of acute myocardial infarction. This process, however, also induces additional loss of viable myocardium, called ischaemia-reperfusion (IR) injury. Complement activation plays an important role in IR injury, partly through binding of C5a to its major receptor (C5aR). We investigated the role of C5aR on infarct size and cardiac function in a model for myocardial IR injury