scholarly journals The Non-structural Protein NSs of SFTSV Causes Cytokine Storm Through the Hyper-activation of NF-κB

2020 ◽  
Author(s):  
Jumana Khalil ◽  
Shintaro Yamada ◽  
Yuta Tsukamoto ◽  
Hiroto Abe ◽  
Masayuki Shimojima ◽  
...  
Keyword(s):  
Inflammatory Cytokines ◽  
Pharmacological Intervention ◽  
Type I ◽  
Dependent Manner ◽  
Cytokine Storm ◽  
Structural Protein ◽  
Cytokines And Chemokines ◽  
Activity Dependent

Severe fever with thrombocytopenia syndrome (SFTS) virus (SFTSV) is an emerging highly pathogenic phlebovirus. The syndrome is characterized by the substantial production of inflammatory cytokines and chemokines, described as cytokine storm, which correlates with multi-organ failure and high mortality. SFSTV nonstructural (NSs) protein was suggested to mediate the pathogenesis by inhibiting antiviral interferon signaling in the host. However, whether SFTSV NSs protein mediates the induction of fatal cytokine storm remains unaddressed. We demonstrated that SFTSV NSs promotes the hyper-induction of cytokine/chemokine genes in vitro, reminiscent of cytokine storm. Using gene deletion and pharmacological intervention, we found that the induced cytokine storm is driven by the transcription factor NF-κB. Our investigation revealed that TANK-binding kinase 1 (TBK1) suppresses NF-κB signaling and cytokine/chemokine induction in its kinase activity-dependent manner, and that NSs sequesters TBK1 to prevent it from suppressing NF-κB, thereby promoting the activation of NF-κB and its target cytokine/chemokine genes. Of note, NF-κB inhibition suppressed the induction of pro-inflammatory cytokines in SFTSV-infected type I interferon (IFN-I) receptor 1-deficient (Ifnar1-/-) mice. These findings establish the essential role of NSs in SFTS pathogenesis and suggest NF-κB as a possible therapeutic target.

N-Linked Glycans within the A1A2A3 Domains of VWF Play a Critical Role in Modulating Macrophage-Mediated Clearance

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 469-469
Author(s):  
Alain Chion ◽  
Jamie O'Sullivan ◽  
Gudmundur Bergsson ◽  
Sean Keyes ◽  
Orla Rawley ◽  
...  
Keyword(s):  
Plasma Clearance ◽  
Full Length ◽  
Type I ◽  
Dependent Manner ◽  
Wild Type ◽  
Macrophage Receptors ◽  
A2 Domain

Abstract Enhanced plasma clearance of von Willebrand factor (VWF) plays an important role in the etiology of both type 1 and type 2 VWD. Nevertheless, although significant progress has been achieved in understanding the structure and functional properties of VWF, the mechanism(s) responsible for modulating VWF clearance from the plasma remain poorly understood. Accumulating recent data suggests that hepatic and splenic macrophages play key roles in modulating VWF clearance. A number of putative macrophage receptors for VWF have been also been described, including LRP1, β2-integrins and Siglec-5. In addition, it is well recognised that variation in VWF glycan expression significantly influences its clearance rate. In particular, terminal ABO(H) blood group determinants which are predominantly expressed on the N-linked glycans of human VWF significantly modulate its rate of clearance. Critically however, the molecular mechanisms through which specific macrophage receptors interact with particular regions of the complex VWF glycoprotein have not been defined. To investigate the role of VWF glycans and specific VWF domains in regulating VWF clearance, we expressed and purified a series of recombinant VWF variants and truncations with/without specific glycan sites. In addition, VWF glycosylation was modified using specific exoglycosidase digestions. Subsequently, recombinant VWF variants and glycoforms thereof were injected into VWF-/-mice, and plasma VWF clearance rates determined by ELISA. VWF-macrophage interactions were also quantified in vitro using phorbol ester-differentiated monocytic THP-1 cells, and primary human monocytes, in a High Content Analysis Imaging system. In keeping with previous reports, we observed that clearance of a truncated VWFA1A2A3 fragment in VWF-/-mice was very similar to that of full-length wild type (WT-) VWF (VWFA1A2A3; t1/2 = 6.3 min versus rWT-VWF; t1/2 = 7.9 min). Furthermore, chemical depletion of macrophages using clodronate liposomes administration significantly inhibited A1A2A3 clearance in vivo (1.7-fold at 10 min time point) to a similar extent to that observed with full length VWF. In vitro binding experiments confirmed that A1A2A3 bound to differentiated THP-1 cells in a dose- and time- dependent manner. Interestingly, this binding was significantly enhanced in the presence of ristocetin. Cumulatively, these data demonstrate that the A1A2A3 domains of VWF contain a critical receptor-binding site for macrophage-mediated clearance. Interestingly, we observed that the half-life of infused human plasma-derived VWF and recombinant VWF expressed in HEK293T cells in VWF-/- mice were significantly different. Furthermore, treatment with PNGase F to completely remove N-linked glycan structures markedly enhanced the clearance of full length VWF (t1/2 2.1 min; p<0.05). Collectively, these findings highlight the essential roles played by N-glycans in regulating VWF survival. Two N-linked glycan sites are located within A1A2A3 at N1515 and N1574 respectively. Importantly, we found that PNGase digestion of A1A2A3 resulted in markedly enhanced macrophage binding in vitro. Consequently we hypothesized that the two N-glycans located within the A2 domain might be important in regulating VWF clearance by macrophages. Targeted disruption of these individual N-glycan sites by site-directed mutagenesis (A1A2A3-N1515Q and A1A2A3-N1574Q respectively) resulted in significantly enhanced macrophage binding in vitro compared to wild type A1A2A3. Furthermore, following tail vein infusion in VWF-/-mice, full length VWFN1515Q and VWFN1574Q both demonstrated markedly reduced half-lives compared to wild type VWF (VWFN1515Q; t1/2 = 3.7 min, VWFN1574Q; t1/2 = 5.5 min). Finally, introduction of the N1515Q point mutation into truncated A1A2A3 also served to significantly enhance plasma clearance, (A1A2A3N1515Q-VWF; t1/2 = 3.1 min versus A1A2A3-VWF; t1/2 = 6.3 min). In conclusion, our novel data identify a crucial role of the VWF A domains in regulating macrophage-mediated VWF clearance. In addition, we further demonstrate that the N-linked glycans structures located at N1515 and N1574 within the A2 domain play specific roles in protecting VWF against in vivo clearance by macrophages. Given the important role played by enhanced VWF clearance in the etiology of type I VWD, these findings are of direct clinical importance. Disclosures No relevant conflicts of interest to declare.

scholarly journals Ebola Virus Binding to Tim-1 on T Lymphocytes Induces a Cytokine Storm

mBio ◽  
2017 ◽  
Vol 8 (5) ◽  
Author(s):  
Patrick Younan ◽  
Mathieu Iampietro ◽  
Andrew Nishida ◽  
Palaniappan Ramanathan ◽  
Rodrigo I. Santos ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
T Cell Activation ◽  
Cell Activation ◽  
Ebola Virus ◽  
Dependent Manner ◽  
Cytokine Storm

ABSTRACT Ebola virus (EBOV) disease (EVD) results from an exacerbated immunological response that is highlighted by a burst in the production of inflammatory mediators known as a “cytokine storm.” Previous reports have suggested that nonspecific activation of T lymphocytes may play a central role in this phenomenon. T-cell immunoglobulin and mucin domain-containing protein 1 (Tim-1) has recently been shown to interact with virion-associated phosphatidylserine to promote infection. Here, we demonstrate the central role of Tim-1 in EBOV pathogenesis, as Tim-1−/− mice exhibited increased survival rates and reduced disease severity; surprisingly, only a limited decrease in viremia was detected. Tim-1−/− mice exhibited a modified inflammatory response as evidenced by changes in serum cytokines and activation of T helper subsets. A series of in vitro assays based on the Tim-1 expression profile on T cells demonstrated that despite the apparent absence of detectable viral replication in T lymphocytes, EBOV directly binds to isolated T lymphocytes in a phosphatidylserine–Tim-1-dependent manner. Exposure to EBOV resulted in the rapid development of a CD4Hi CD3Low population, non-antigen-specific activation, and cytokine production. Transcriptome and Western blot analysis of EBOV-stimulated CD4+ T cells confirmed the induction of the Tim-1 signaling pathway. Furthermore, comparative analysis of transcriptome data and cytokine/chemokine analysis of supernatants highlight the similarities associated with EBOV-stimulated T cells and the onset of a cytokine storm. Flow cytometry revealed virtually exclusive binding and activation of central memory CD4+ T cells. These findings provide evidence for the role of Tim-1 in the induction of a cytokine storm phenomenon and the pathogenesis of EVD. IMPORTANCE Ebola virus infection is characterized by a massive release of inflammatory mediators, which has come to be known as a cytokine storm. The severity of the cytokine storm is consistently linked with fatal disease outcome. Previous findings have demonstrated that specific T-cell subsets are key contributors to the onset of a cytokine storm. In this study, we investigated the role of Tim-1, a T-cell-receptor-independent trigger of T-cell activation. We first demonstrated that Tim-1-knockout (KO) mice survive lethal Ebola virus challenge. We then used a series of in vitro assays to demonstrate that Ebola virus directly binds primary T cells in a Tim-1–phosphatidylserine-dependent manner. We noted that binding induces a cytokine storm-like phenomenon and that blocking Tim-1–phosphatidylserine interactions reduces viral binding, T-cell activation, and cytokine production. These findings highlight a previously unknown role of Tim-1 in the development of a cytokine storm and “immune paralysis.” IMPORTANCE Ebola virus infection is characterized by a massive release of inflammatory mediators, which has come to be known as a cytokine storm. The severity of the cytokine storm is consistently linked with fatal disease outcome. Previous findings have demonstrated that specific T-cell subsets are key contributors to the onset of a cytokine storm. In this study, we investigated the role of Tim-1, a T-cell-receptor-independent trigger of T-cell activation. We first demonstrated that Tim-1-knockout (KO) mice survive lethal Ebola virus challenge. We then used a series of in vitro assays to demonstrate that Ebola virus directly binds primary T cells in a Tim-1–phosphatidylserine-dependent manner. We noted that binding induces a cytokine storm-like phenomenon and that blocking Tim-1–phosphatidylserine interactions reduces viral binding, T-cell activation, and cytokine production. These findings highlight a previously unknown role of Tim-1 in the development of a cytokine storm and “immune paralysis.”

scholarly journals Noncanonical Wnt5a/JNK Signaling Contributes to the Development of D-Gal/LPS-Induced Acute Liver Failure

2021 ◽  
Author(s):  
Xiang-fen Ji ◽  
Yu-chen Fan ◽  
Fei Sun ◽  
Jing-wei Wang ◽  
kai wang
Keyword(s):  
Liver Failure ◽  
Acute Liver Failure ◽  
Inflammatory Cytokines ◽  
Dependent Manner ◽  
Jnk Signaling ◽  
Wnt5a Expression ◽  
Jnk Activation

Abstract Acute liver failure (ALF) is a deadly clinical disorder with few effective treatments and unclear pathogenesis. In our previous study, we demonstrated that aberrant Wnt5a expression was involved in acute on chronic liver failure. However, the role of Wnt5a in ALF is unknown. We investigated the expression of Wnt5a and its downstream signaling of c-jun N-terminal kinase (JNK) in a mouse model of ALF established by co-injection of D-galactosamine (D-Gal) and lipopolysaccharide (LPS) in C57BL/6 mice. We also investigated the role of Box5, a Wnt5a antagonist in vivo. Moreover, the effect of Wnt5a/JNK signaling on downstream inflammatory cytokines expression, phagocytosis and migration in THP-1 macrophages was studied in vitro. Aberrant Wnt5a expression and JNK activation were detected in D-Gal/LPS-induced ALF mice. Box5 pretreatment reversed JNK activation, and eventually decreased the mortality rate of D-Gal/LPS-treated mice with reduced hepatic necrosis and apoptosis, serum ALT and AST levels, and liver inflammatory cytokines expression, although the last was not significant. We further demonstrated that recombined Wnt5a (rWnt5a) induced tumor necrosis α (TNF-α) and Interleukin-6 (IL-6) mRNA expression, and increased the phagocytosis ability of THP-1 macrophages in a JNK-dependent manner, which could be restored by Box5. In addition, rWnt5a-induced migration of THP-1 macrophages was also turned by Box5. Our findings suggested that Wnt5a/JNK signaling play important role in the development of ALF, and Box5 could have particular hepatoprotecive effects in ALF.

scholarly journals RIC in COVID-19—a Clinical Trial to Investigate Whether Remote Ischemic Conditioning (RIC) Can Prevent Deterioration to Critical Care in Patients with COVID-19

2021 ◽  
Author(s):  
Sean M. Davidson ◽  
Kishal Lukhna ◽  
Diana A. Gorog ◽  
Alan D. Salama ◽  
Alejandro Rosell Castillo ◽  
...  
Keyword(s):  
South Africa ◽  
Clinical Trial ◽  
United Kingdom ◽  
Inflammatory Cytokines ◽  
In Vitro Cytotoxicity ◽  
Cytokine Storm ◽  
Remote Ischemic Conditioning ◽  
Ischemic Conditioning ◽  
The United Kingdom

Abstract Purpose Coronavirus disease 19 (COVID-19) has, to date, been diagnosed in over 130 million persons worldwide and is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Several variants of concern have emerged including those in the United Kingdom, South Africa, and Brazil. SARS-CoV-2 can cause a dysregulated inflammatory response known as a cytokine storm, which can progress rapidly to acute respiratory distress syndrome (ARDS), multi-organ failure, and death. Suppressing these cytokine elevations may be key to improving outcomes. Remote ischemic conditioning (RIC) is a simple, non-invasive procedure whereby a blood pressure cuff is inflated and deflated on the upper arm for several cycles. “RIC in COVID-19” is a pilot, multi-center, randomized clinical trial, designed to ascertain whether RIC suppresses inflammatory cytokine production. Methods A minimum of 55 adult patients with diagnosed COVID-19, but not of critical status, will be enrolled from centers in the United Kingdom, Brazil, and South Africa. RIC will be administered daily for up to 15 days. The primary outcome is the level of inflammatory cytokines that are involved in the cytokine storm that can occur following SARS-CoV-2 infection. The secondary endpoint is the time between admission and until intensive care admission or death. The in vitro cytotoxicity of patient blood will also be assessed using primary human cardiac endothelial cells. Conclusions The results of this pilot study will provide initial evidence on the ability of RIC to suppress the production of inflammatory cytokines in the setting of COVID-19. Trial Registration NCT04699227, registered January 7th, 2021.

scholarly journals Activation of mitochondrial TUFM ameliorates metabolic dysregulation through coordinating autophagy induction

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Dasol Kim ◽  
Hui-Yun Hwang ◽  
Eun Sun Ji ◽  
Jin Young Kim ◽  
Jong Shin Yoo ◽  
...  
Keyword(s):  
Metabolic Regulation ◽  
Elongation Factor ◽  
Dependent Manner ◽  
Diet Induced Obesity ◽  
Metabolic Dysregulation ◽  
Autophagy Induction ◽  
Transcription Factor Eb ◽  
Mitochondrial Elongation

AbstractDisorders of autophagy, a key regulator of cellular homeostasis, cause a number of human diseases. Due to the role of autophagy in metabolic dysregulation, there is a need to identify autophagy regulators as therapeutic targets. To address this need, we conducted an autophagy phenotype-based screen and identified the natural compound kaempferide (Kaem) as an autophagy enhancer. Kaem promoted autophagy through translocation of transcription factor EB (TFEB) without MTOR perturbation, suggesting it is safe for administration. Moreover, Kaem accelerated lipid droplet degradation in a lysosomal activity-dependent manner in vitro and ameliorated metabolic dysregulation in a diet-induced obesity mouse model. To elucidate the mechanism underlying Kaem’s biological activity, the target protein was identified via combined drug affinity responsive target stability and LC–MS/MS analyses. Kaem directly interacted with the mitochondrial elongation factor TUFM, and TUFM absence reversed Kaem-induced autophagy and lipid degradation. Kaem also induced mitochondrial reactive oxygen species (mtROS) to sequentially promote lysosomal Ca2+ efflux, TFEB translocation and autophagy induction, suggesting a role of TUFM in mtROS regulation. Collectively, these results demonstrate that Kaem is a potential therapeutic candidate/chemical tool for treating metabolic dysregulation and reveal a role for TUFM in autophagy for metabolic regulation with lipid overload.

scholarly journals Secreted KIAA1199 promotes the progression of rheumatoid arthritis by mediating hyaluronic acid degradation in an ANXA1-dependent manner

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Wei Zhang ◽  
Guoyu Yin ◽  
Heping Zhao ◽  
Hanzhi Ling ◽  
Zhen Xie ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Hyaluronic Acid ◽  
Dependent Manner ◽  
Collagen Induced Arthritis ◽  
Intracellular Degradation ◽  
Main Pathway ◽  
First Time

AbstractIn inflamed joints, enhanced hyaluronic acid (HA) degradation is closely related to the pathogenesis of rheumatoid arthritis (RA). KIAA1199 has been identified as a hyaladherin that mediates the intracellular degradation of HA, but its extracellular function remains unclear. In this study, we found that the serum and synovial levels of secreted KIAA1199 (sKIAA1199) and low-molecular-weight HA (LMW-HA, MW < 100 kDa) in RA patients were significantly increased, and the positive correlation between them was shown for the first time. Of note, treatment with anti-KIAA1199 mAb effectively alleviated the severity of arthritis and reduced serum LMW-HA levels and cytokine secretion in collagen-induced arthritis (CIA) mice. In vitro, sKIAA1199 was shown to mediate exogenous HA degradation by attaching to the cell membrane of RA fibroblast-like synoviosytes (RA FLS). Furthermore, the HA-degrading activity of sKIAA1199 depended largely on its adhesion to the membrane, which was achieved by its G8 domain binding to ANXA1. In vivo, kiaa1199-KO mice exhibited greater resistance to collagen-induced arthritis. Interestingly, this resistance could be partially reversed by intra-articular injection of vectors encoding full-length KIAA1199 instead of G8-deleted KIAA119 mutant, which further confirmed the indispensable role of G8 domain in KIAA1199 involvement in RA pathological processes. Mechanically, the activation of NF-κB by interleukin-6 (IL-6) through PI3K/Akt signaling is suggested to be the main pathway to induce KIAA1199 expression in RA FLS. In conclusion, our study supported the contribution of sKIAA1199 to RA pathogenesis, providing a new therapeutic target for RA by blocking sKIAA1199-mediated HA degradation.

scholarly journals Phaseolin Attenuates Lipopolysaccharide-Induced Inflammation in RAW 264.7 Cells and Zebrafish

Biomedicines ◽  
2021 ◽  
Vol 9 (4) ◽  
pp. 420
Author(s):  
Su-Jung Hwang ◽  
Ye-Seul Song ◽  
Hyo-Jong Lee
Keyword(s):  
Nitric Oxide ◽  
Nuclear Translocation ◽  
Raw 264.7 Cells ◽  
Network Pharmacology ◽  
Raw 264.7 ◽  
Dependent Manner ◽  
Bioactive Constituents

Kushen (Radix Sophorae flavescentis) is used to treat ulcerative colitis, tumors, and pruritus. Recently, phaseolin, formononetin, matrine, luteolin, and quercetin, through a network pharmacology approach, were tentatively identified as five bioactive constituents responsible for the anti-inflammatory effects of S. flavescentis. However, the role of phaseolin (one of the primary components of S. flavescentis) in the direct regulation of inflammation and inflammatory processes is not well known. In this study, the beneficial role of phaseolin against inflammation was explored in lipopolysaccharide (LPS)-induced inflammation models of RAW 264.7 macrophages and zebrafish larvae. Phaseolin inhibited LPS-mediated production of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS), without affecting cell viability. In addition, phaseolin suppressed pro-inflammatory mediators such as cyclooxygenase 2 (COX-2), interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α), monocyte chemoattractant protein-1 (MCP-1), and interleukin-6 (IL-6) in a dose-dependent manner. Furthermore, phaseolin reduced matrix metalloproteinase (MMP) activity as well as macrophage adhesion in vitro and the recruitment of leukocytes in vivo by downregulating Ninjurin 1 (Ninj1), an adhesion molecule. Finally, phaseolin inhibited the nuclear translocation of nuclear factor-kappa B (NF-κB). In view of the above, our results suggest that phaseolin could be a potential therapeutic candidate for the management of inflammation.

scholarly journals Establishment of a New Device for Electrical Stimulation of Non-Degenerative Cartilage Cells In Vitro

2021 ◽  
Vol 22 (1) ◽  
pp. 394
Author(s):  
Simone Krueger ◽  
Alexander Riess ◽  
Anika Jonitz-Heincke ◽  
Alina Weizel ◽  
Anika Seyfarth ◽  
...  
Keyword(s):  
Electrical Stimulation ◽  
Electric Fields ◽  
Cartilage Tissue ◽  
Type I ◽  
Dependent Manner ◽  
New Device ◽  
Alternating Electric Fields ◽  
Cartilage Cells ◽  
Stimulation Of

In cell-based therapies for cartilage lesions, the main problem is still the formation of fibrous cartilage, caused by underlying de-differentiation processes ex vivo. Biophysical stimulation is a promising approach to optimize cell-based procedures and to adapt them more closely to physiological conditions. The occurrence of mechano-electrical transduction phenomena within cartilage tissue is physiological and based on streaming and diffusion potentials. The application of exogenous electric fields can be used to mimic endogenous fields and, thus, support the differentiation of chondrocytes in vitro. For this purpose, we have developed a new device for electrical stimulation of chondrocytes, which operates on the basis of capacitive coupling of alternating electric fields. The reusable and sterilizable stimulation device allows the simultaneous use of 12 cavities with independently applicable fields using only one main supply. The first parameter settings for the stimulation of human non-degenerative chondrocytes, seeded on collagen type I elastin-based scaffolds, were derived from numerical electric field simulations. Our first results suggest that applied alternating electric fields induce chondrogenic re-differentiation at the gene and especially at the protein level of human de-differentiated chondrocytes in a frequency-dependent manner. In future studies, further parameter optimizations will be performed to improve the differentiation capacity of human cartilage cells.

scholarly journals PapRIV, a BV-2 microglial cell activating quorum sensing peptide

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yorick Janssens ◽  
Nathan Debunne ◽  
Anton De Spiegeleer ◽  
Evelien Wynendaele ◽  
Marta Planas ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Cell Model ◽  
Dependent Manner ◽  
Communication Signals ◽  
Gram Positive Bacteria ◽  
A Cell ◽  
Gastro Intestinal Tract

AbstractQuorum sensing peptides (QSPs) are bacterial peptides produced by Gram-positive bacteria to communicate with their peers in a cell-density dependent manner. These peptides do not only act as interbacterial communication signals, but can also have effects on the host. Compelling evidence demonstrates the presence of a gut-brain axis and more specifically, the role of the gut microbiota in microglial functioning. The aim of this study is to investigate microglial activating properties of a selected QSP (PapRIV) which is produced by Bacillus cereus species. PapRIV showed in vitro activating properties of BV-2 microglia cells and was able to cross the in vitro Caco-2 cell model and reach the brain. In vivo peptide presence was also demonstrated in mouse plasma. The peptide caused induction of IL-6, TNFα and ROS expression and increased the fraction of ameboid BV-2 microglia cells in an NF-κB dependent manner. Different metabolites were identified in serum, of which the main metabolite still remained active. PapRIV is thus able to cross the gastro-intestinal tract and the blood–brain barrier and shows in vitro activating properties in BV-2 microglia cells, hereby indicating a potential role of this quorum sensing peptide in gut-brain interaction.

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