Protein Phosphatase 1 Abrogates IRF7-Mediated Type I IFN Response In Antiviral Immunity

Interferon (IFN) regulatory factor 7 (IRF7) plays a key role in the production of IFN‐α in response to viral infection, and phosphorylation at IRF7 C‐terminal serine sites is prelude to its function. However, phosphatases that negatively regulate IRF7 phosphorylation and activity have not been reported. In this study, we have identified a conserved protein phosphatase 1 (PP1)‐binding motif in human and mouse IRF7 proteins, and shown that PP1 physically interacts with IRF7. Exogenous expression of PP1 subunits (PP1α, β, or γ) ablates IKKε‐stimulated IRF7 phosphorylation and dramatically attenuates IRF7 transcriptional activity. Inhibition of PP1 activity significantly increases IRF7 phosphorylation and IRF7‐mediated IFN‐α production in response to Newcastle disease virus (NDV) infection or Toll‐like receptor 7 (TLR7) challenge, leading to impaired viral replication. In addition, IFN treatment, TLR challenges and viral infection induce PP1 expression. Our findings disclose for the first time a pivotal role for PP1 in impeding IRF7‐mediated IFN‐α production in host immune responses

Neuromuscular Adaptations Following Training and Protein Supplementation in a Group of Trained Weightlifters

The purpose of this study was to examine the effects of a recovery supplement compared with a placebo on muscle morphology in trained weightlifters. Vastus lateralis and muscle fiber cross sectional area of type I and type II fibers were compared between groups using a series of 2 × 2 (group × time) repeated measure ANOVAs. Both groups on average improved cross-sectional area of the vastus lateralis, type I and type II muscle fibers from pre-to-post but individual response varied within both groups. Greater magnitude of changes in type I and type II muscle fibers were observed for the placebo group but not for vastus lateralis cross sectional area. Additionally, subjects were divided into large and small fiber groups based on combined fiber size at the start of the investigation. These findings indicate that the recovery supplement utilized provided no greater effect compared with a placebo in a 12-week block periodization protocol in trained weightlifters. The primary determinate of fiber size changes in the study was determined to be the initial fiber size of muscle fibers with larger practical changes observed in the small fiber group compared with the large fiber group in type I, II, and ultrasound cross-sectional area (CSA)

p62-mediated Selective Autophagy Endows Virus-Transformed Cells With Insusceptibility to DNA Damage Under Oxidative Stress

DNA damage response (DDR) and selective autophagy both can be activated by reactive oxygen/nitrogen species (ROS/RNS), and both are of paramount importance in cancer development. The selective autophagy receptor and ubiquitin (Ub) sensor p62 plays a key role in their crosstalk. ROS production has been well documented in latent infection of oncogenic viruses including Epstein-Barr Virus (EBV). However, p62-mediated selective autophagy and its interplay with DDR have not been investigated in these settings. In this study, we provide evidence that considerable levels of p62-mediated selective autophagy are spontaneously induced, and correlate with ROS-Keap1-NRF2 pathway activity, in virus-transformed cells. Inhibition of autophagy results in p62 accumulation in the nucleus, and promotes ROS-induced DNA damage and cell death, as well as downregulates the DNA repair proteins CHK1 and RAD51. In contrast, MG132-mediated proteasome inhibition, which induces rigorous autophagy, promotes p62 degradation but accumulation of the DNA repair proteins CHK1 and RAD51. However, pretreatment with an autophagy inhibitor offsets the effects of MG132 on CHK1 and RAD51 levels. These findings imply that p62 accumulation in the nucleus in response to autophagy inhibition promotes proteasome-mediated CHK1 and RAD51 protein instability. This claim is further supported by the findings that transient expression of a p62 mutant, which is constitutively localized in the nucleus, in B cell lines with low endogenous p62 levels recaptures the effects of autophagy inhibition on CHK1 and RAD51 protein stability. These results indicate that proteasomal degradation of RAD51 and CHK1 is dependent on p62 accumulation in the nucleus. However, small hairpin RNA (shRNA)-mediated p62 depletion in EBV-transformed lymphoblastic cell lines (LCLs) had no apparent effects on the protein levels of CHK1 and RAD51, likely due to the constitutive localization of p62 in the cytoplasm and incomplete knockdown is insufficient to manifest its nuclear effects on these proteins. Rather, shRNA-mediated p62 depletion in EBV-transformed LCLs results in significant increases of endogenous RNF168-γH2AX damage foci and chromatin ubiquitination, indicative of activation of RNF168-mediated DNA repair mechanisms. Our results have unveiled a pivotal role for p62-mediated selective autophagy that governs DDR in the setting of oncogenic virus latent infection, and provide a novel insight into virus-mediated oncogenesis

Enhanced effects of cigarette smoke extract on inflammatory cytokine expression in IL-1β-activated human mast cells were inhibited by Baicalein via regulation of the NF-κB pathway

Background: Human mast cells are capable of a wide variety of inflammatory responses and play a vital role in the pathogenesis of inflammatory diseases such as allergy, asthma, and atherosclerosis. We have reported that cigarette smoke extract (CSE) significantly increased IL-6 and IL-8 production in IL-1β-activated human mast cell line (HMC-1). Baicalein (BAI) has anti-inflammatory properties and inhibits IL-1β- and TNF-α-induced inflammatory cytokine production from HMC-1. The goal of the present study was to examine the effect of BAI on IL-6 and IL-8 production from CSE-treated and IL-1β-activated HMC-1.Methods: Main-stream (Ms) and Side-stream (Ss) cigarette smoke were collected onto fiber filters and extracted in RPMI-1640 medium. Two ml of HMC-1 at 1 × 10 6 cells/mL were cultured with CSE in the presence or absence of IL-1β (10 ng/mL) for 24 hrs. A group of HMC-1 cells stimulated with both IL-1β (10 ng/ml) and CSE was also treated with BAI. The expression of IL-6 and IL-8 was assessed by ELISA and RT-PCR. NF-κB activation was measured by electrophoretic mobility shift assay (EMSA) and IκBα degradation by Western blot.Results: Both Ms and Ss CSE significantly increased IL-6 and IL-8 production (p \u3c 0.001) in IL-1β-activated HMC-1. CSE increased NF-κB activation and decreased cytoplasmic IκBα proteins in IL-1β-activated HMC-1. BAI (1.8 to 30 μM) significantly inhibited production of IL-6 and IL-8 in a dose-dependent manner in IL-1β-activated HMC-1 with the optimal inhibition concentration at 30 μM, which also significantly inhibited the enhancing effect of CSE on IL-6 and IL-8 production in IL-1β-activated HMC-1. BAI inhibited NF-κB activation and increased cytoplasmic IκBα proteins in CSE-treated and IL-1β-activated HMC-1.Conclusions: Our results showed that CSE significantly increased inflammatory cytokines IL-6 and IL-8 production in IL-1β-activated HMC-1. It may partially explain why cigarette smoke contributes to lung and cardiovascular diseases. BAI inhibited the production of inflammatory cytokines through inhibition of NF-κB activation and IκBα phosphorylation and degradation. This inhibitory effect of BAI on the expression of inflammatory cytokines induced by CSE suggests its usefulness in the development of novel anti-inflammatory therapies

Cause of Death and Predictors of All-Cause Mortality in Anticoagulated Patients With Nonvalvular Atrial Fibrillation : Data From ROCKET AF

M. Kaste on työryhmän ROCKET AF Steering Comm jäsen.Background-Atrial fibrillation is associated with higher mortality. Identification of causes of death and contemporary risk factors for all-cause mortality may guide interventions. Methods and Results-In the Rivaroxaban Once Daily Oral Direct Factor Xa Inhibition Compared with Vitamin K Antagonism for Prevention of Stroke and Embolism Trial in Atrial Fibrillation (ROCKET AF) study, patients with nonvalvular atrial fibrillation were randomized to rivaroxaban or dose-adjusted warfarin. Cox proportional hazards regression with backward elimination identified factors at randomization that were independently associated with all-cause mortality in the 14 171 participants in the intention-to-treat population. The median age was 73 years, and the mean CHADS(2) score was 3.5. Over 1.9 years of median follow-up, 1214 (8.6%) patients died. Kaplan-Meier mortality rates were 4.2% at 1 year and 8.9% at 2 years. The majority of classified deaths (1081) were cardiovascular (72%), whereas only 6% were nonhemorrhagic stroke or systemic embolism. No significant difference in all-cause mortality was observed between the rivaroxaban and warfarin arms (P=0.15). Heart failure (hazard ratio 1.51, 95% CI 1.33-1.70, P= 75 years (hazard ratio 1.69, 95% CI 1.51-1.90, P Conclusions-In a large population of patients anticoagulated for nonvalvular atrial fibrillation, approximate to 7 in 10 deaths were cardiovascular, whereasPeer reviewe

Human Lung Fibroblasts Express Interleukin-6 in Response to Signaling After Mast Cell Contact

Asthma is a chronic inflammatory disease of the airways. Mast cell-derived cytokines may mediate both airway inflammation and remodeling. It has also been shown that fibroblasts can be the source of proinflammatory cytokines. In the human airways, mast cell-fibroblast interactions may have pivotal effects on modulating inflammation. To study this further, we cocultured normal human lung fibroblasts (NHLF) with a human mast cell line (HMC-1) and assayed for production of interleukin (IL)-6, an important proinflammatory cytokine. When cultured together, NHLF/HMC-1 contact Induced IL-6 secretion. Separation of HMC-1 and NHLF cells by a porous membrane inhibited this induction. HMC-1-derived cellular membranes caused an increase in IL-6 production in NHLF. Activation of p38 MAPK was also seen in cocultures by Western blot, whereas IL-6 production in cocultures was significantly inhibited by the p38 inhibitor SB203580. IL-6 production in cocultures was minimally inhibited by a chemical inhibitor of nuclear factor-κB (Bay11), indicating that nuclear factor-κB may have a minimal role in signaling IL-6 production in mast cell/fibroblasts cocultures. Blockade of intercellular adhesion molecule-1, tumor necrosis factor-RI, and surface IL-1β with neutralizing antibodies failed to significantly decrease IL-6 production in our coculture, indicating that other receptor-ligand associations may be responsible for this activation. These novel studies reveal the importance of cell-cell interactions in the complex milieu of airway inflammation

HIV-1 Tat Protein-Induced VCAM-1 Expression in Human Pulmonary Artery Endothelial Cells and Its Signaling

Expression of cell adhesion molecule in endothelial cells upon activation by human immunodeficiency virus (HIV) infection is associated with the development of atherosclerotic vasculopathy. We postulated that induction of vascular cell adhesion molecule-1 (VCAM-1) by HIV-1 Tat protein in endothelial cells might represent an early event that could culminate in inflammatory cell recruitment and vascular injury. We determined the role of HIV-1 Tat protein in VCAM-1 expression in human pulmonary artery endothelial cells (HPAEC). HIV-1 Tat protein treatment significantly increased cell-surface expression of VCAM-1 in HPAEC. Consistently, mRNA expression of VCAM-1 was also increased by HIV-1 Tat protein as measured by RT-PCR. HIV-1 Tat protein-induced VCAM-1 expression was abolished by the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) and the p38 MAPK inhibitor SB-203580. Furthermore, HIV-1 Tat protein enhanced DNA binding activity of NF-κB, facilitated nuclear translocation of NF-κB subunit p65, and increased production of reactive oxygen species (ROS). Similarly to VCAM-1 expression, HIV-1 Tat protein-induced NF-κB activation and ROS generation were abrogated by PDTC and SB-203580. These data indicate that HIV-1 Tat protein is able to induce VCAM-1 expression in HPAEC, which may represent a pivotal early molecular event in HIV-induced vascular/pulmonary injury. These data also suggest that the molecular mechanism underlying the HIV-1 Tat protein-induced VCAM-1 expression may involve ROS generation, p38 MAPK activation, and NF-κB translocation, which are the characteristics of pulmonary endothelial cell activation