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1.
Cell Transplant ; 30: 9636897211054481, 2021.
Article in English | MEDLINE | ID: covidwho-1511642

ABSTRACT

Biological and cellular interleukin-6 (IL-6)-related therapies have been used to treat severe COVID-19 pneumonia with hyperinflammatory syndrome and acute respiratory failure, which prompted further exploration of the role of IL-6 in human umbilical cord mesenchymal stem cell (hUCMSC) therapy. Peripheral blood mononuclear cells (PBMCs) were responders cocultured with hUCMSCs or exogenous IL-6. A PBMC suppression assay was used to analyze the anti-inflammatory effects via MTT assay. The IL-6 concentration in the supernatant was measured using ELISA. The correlation between the anti-inflammatory effect of hUCMSCs and IL-6 levels and the relevant roles of IL-6 and IL-6 mRNA expression was analyzed using the MetaCore functional network constructed from gene microarray data. The location of IL-6 and IL-6 receptor (IL-6R) expression was further evaluated. We reported that hUCMSCs did not initially exert any inhibitory effect on PHA-stimulated proliferation; however, a potent inhibitory effect on PHA-stimulated proliferation was observed, and the IL-6 concentration reached approximately 1000 ng/mL after 72 hours. Exogenous 1000 ng/mL IL-6 inhibited PHA-stimulated inflammation but less so than hUCMSCs. The inhibitory effects of hUCMSCs on PHA-stimulated PBMCs disappeared after adding an IL-6 neutralizing antibody or pretreatment with tocilizumab (TCZ), an IL-6R antagonist. hUCMSCs exert excellent anti-inflammatory effects by inducing higher IL-6 levels, which is different from TCZ. High concentration of IL-6 cytokine secretion plays an important role in the anti-inflammatory effect of hUCMSC therapy. Initial hUCMSC therapy, followed by TCZ, seems to optimize the therapeutic potential to treat COVID-19-related acute respiratory distress syndrome (ARDS).


Subject(s)
Antibodies, Monoclonal, Humanized/therapeutic use , COVID-19/complications , Interleukin-6/biosynthesis , Mesenchymal Stem Cell Transplantation , Mesenchymal Stem Cells/metabolism , Respiratory Distress Syndrome/therapy , SARS-CoV-2 , Antibodies, Monoclonal, Humanized/pharmacology , Antibodies, Neutralizing/immunology , Cells, Cultured , Coculture Techniques , Combined Modality Therapy , DNA, Complementary/genetics , Gene Expression Regulation/drug effects , Humans , Inflammation , Interleukin-6/genetics , Interleukin-6/pharmacology , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/metabolism , Lymphocyte Activation/drug effects , Phytohemagglutinins/pharmacology , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Receptors, Interleukin-6/antagonists & inhibitors , Receptors, Interleukin-6/biosynthesis , Receptors, Interleukin-6/genetics , Respiratory Distress Syndrome/drug therapy , Respiratory Distress Syndrome/etiology , Umbilical Cord/cytology
2.
Int J Mol Sci ; 22(20)2021 Oct 18.
Article in English | MEDLINE | ID: covidwho-1470894

ABSTRACT

Infection caused by the severe acute respiratory syndrome coronavirus (SARS-CoV-2) in many cases is accompanied by the release of a large amount of proinflammatory cytokines in an event known as "cytokine storm", which is associated with severe coronavirus disease 2019 (COVID-19) cases and high mortality. The excessive production of proinflammatory cytokines is linked, inter alia, to the enhanced activity of receptors capable of recognizing the conservative regions of pathogens and cell debris, namely TLRs, TREM-1 and TNFR1. Here we report that peptides derived from innate immunity protein Tag7 inhibit activation of TREM-1 and TNFR1 receptors during acute inflammation. Peptides from the N-terminal fragment of Tag7 bind only to TREM-1, while peptides from the C-terminal fragment interact solely with TNFR1. Selected peptides are capable of inhibiting the production of proinflammatory cytokines both in peripheral blood mononuclear cells (PBMCs) from healthy donors and in vivo in the mouse model of acute lung injury (ALI) by diffuse alveolar damage (DAD). Treatment with peptides significantly decreases the infiltration of mononuclear cells to lungs in animals with DAD. Our findings suggest that Tag7-derived peptides might be beneficial in terms of the therapy or prevention of acute lung injury, e.g., for treating COVID-19 patients with severe pulmonary lesions.


Subject(s)
Acute Lung Injury/pathology , Cytokines/chemistry , Peptides/metabolism , Receptors, Tumor Necrosis Factor, Type I/metabolism , Triggering Receptor Expressed on Myeloid Cells-1/metabolism , Acute Lung Injury/metabolism , Animals , Cytokines/genetics , Cytokines/metabolism , Disease Models, Animal , Humans , Interferon-gamma/genetics , Interferon-gamma/metabolism , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/metabolism , Lipopolysaccharides/pharmacology , Lung/metabolism , Lung/pathology , Lymphocyte Activation/drug effects , Male , Mice , Mice, Inbred ICR , Peptides/chemistry , Peptides/pharmacology , Protein Binding , Receptors, Tumor Necrosis Factor, Type I/antagonists & inhibitors , Triggering Receptor Expressed on Myeloid Cells-1/antagonists & inhibitors
3.
Int J Mol Sci ; 22(15)2021 Jul 26.
Article in English | MEDLINE | ID: covidwho-1374420

ABSTRACT

For the treatment of severe COVID-19, supplementation with human plasma-purified α-1 antitrypsin (AAT) to patients is currently considered. AAT inhibits host proteases that facilitate viral entry and possesses broad anti-inflammatory and immunomodulatory activities. Researchers have demonstrated that an interaction between SARS-CoV-2 spike protein (S) and lipopolysaccharides (LPS) enhances pro-inflammatory responses in vitro and in vivo. Hence, we wanted to understand the potential anti-inflammatory activities of plasma-derived and recombinant AAT (recAAT) in a model of human total peripheral blood mononuclear cells (PBMCs) exposed to a combination of CHO expressed trimeric spike protein and LPS, ex vivo. We confirmed that cytokine production was enhanced in PBMCs within six hours when low levels of LPS were combined with purified spike proteins ("spike"). In the presence of 0.5 mg/mL recAAT, however, LPS/spike-induced TNF-α and IL-1ß mRNA expression and protein release were significantly inhibited (by about 46-50%) relative to LPS/spike alone. Although without statistical significance, recAAT also reduced production of IL-6 and IL-8. Notably, under the same experimental conditions, the plasma-derived AAT preparation Respreeza (used in native and oxidized forms) did not show significant effects. Our findings imply that an early pro-inflammatory activation of human PBMCs is better controlled by the recombinant version of AAT than the human plasma-derived AAT used here. Considering the increasing clinical interest in AAT therapy as useful to ameliorate the hyper-inflammation seen during COVID-19 infection, different AAT preparations require careful evaluation.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Leukocytes, Mononuclear/metabolism , Spike Glycoprotein, Coronavirus/metabolism , alpha 1-Antitrypsin/pharmacology , Animals , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/immunology , CHO Cells , COVID-19/therapy , Cells, Cultured , Cricetulus , Cytokines/metabolism , Humans , Inflammation/metabolism , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Lipopolysaccharides/immunology , Lipopolysaccharides/toxicity , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , alpha 1-Antitrypsin/chemistry , alpha 1-Antitrypsin/immunology
4.
J Biol Chem ; 296: 100630, 2021.
Article in English | MEDLINE | ID: covidwho-1333548

ABSTRACT

Unchecked inflammation can result in severe diseases with high mortality, such as macrophage activation syndrome (MAS). MAS and associated cytokine storms have been observed in COVID-19 patients exhibiting systemic hyperinflammation. Interleukin-18 (IL-18), a proinflammatory cytokine belonging to the IL-1 family, is elevated in both MAS and COVID-19 patients, and its level is known to correlate with the severity of COVID-19 symptoms. IL-18 binds its specific receptor IL-1 receptor 5 (IL-1R5, also known as IL-18 receptor alpha chain), leading to the recruitment of the coreceptor, IL-1 receptor 7 (IL-1R7, also known as IL-18 receptor beta chain). This heterotrimeric complex then initiates downstream signaling, resulting in systemic and local inflammation. Here, we developed a novel humanized monoclonal anti-IL-1R7 antibody to specifically block the activity of IL-18 and its inflammatory signaling. We characterized the function of this antibody in human cell lines, in freshly obtained peripheral blood mononuclear cells (PBMCs) and in human whole blood cultures. We found that the anti-IL-1R7 antibody significantly suppressed IL-18-mediated NFκB activation, reduced IL-18-stimulated IFNγ and IL-6 production in human cell lines, and reduced IL-18-induced IFNγ, IL-6, and TNFα production in PBMCs. Moreover, the anti-IL-1R7 antibody significantly inhibited LPS- and Candida albicans-induced IFNγ production in PBMCs, as well as LPS-induced IFNγ production in whole blood cultures. Our data suggest that blocking IL-1R7 could represent a potential therapeutic strategy to specifically modulate IL-18 signaling and may warrant further investigation into its clinical potential for treating IL-18-mediated diseases, including MAS and COVID-19.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Antibodies, Monoclonal/pharmacology , Antibodies, Neutralizing/pharmacology , Immunologic Factors/pharmacology , Interleukin-18/genetics , Receptors, Interleukin-18/genetics , Anti-Inflammatory Agents/metabolism , Antibodies, Monoclonal/biosynthesis , Antibodies, Neutralizing/biosynthesis , COVID-19/drug therapy , Candida albicans/growth & development , Candida albicans/pathogenicity , Gene Expression Regulation , HEK293 Cells , Humans , Immunologic Factors/biosynthesis , Inflammation , Interferon-gamma/genetics , Interferon-gamma/immunology , Interleukin-18/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/microbiology , Lipopolysaccharides/antagonists & inhibitors , Lipopolysaccharides/pharmacology , Macrophage Activation Syndrome/drug therapy , NF-kappa B/genetics , NF-kappa B/immunology , Primary Cell Culture , Receptors, Interleukin-18/antagonists & inhibitors , Receptors, Interleukin-18/immunology , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Signal Transduction/drug effects , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/immunology
5.
Int J Mol Sci ; 22(15)2021 Jul 26.
Article in English | MEDLINE | ID: covidwho-1325683

ABSTRACT

For the treatment of severe COVID-19, supplementation with human plasma-purified α-1 antitrypsin (AAT) to patients is currently considered. AAT inhibits host proteases that facilitate viral entry and possesses broad anti-inflammatory and immunomodulatory activities. Researchers have demonstrated that an interaction between SARS-CoV-2 spike protein (S) and lipopolysaccharides (LPS) enhances pro-inflammatory responses in vitro and in vivo. Hence, we wanted to understand the potential anti-inflammatory activities of plasma-derived and recombinant AAT (recAAT) in a model of human total peripheral blood mononuclear cells (PBMCs) exposed to a combination of CHO expressed trimeric spike protein and LPS, ex vivo. We confirmed that cytokine production was enhanced in PBMCs within six hours when low levels of LPS were combined with purified spike proteins ("spike"). In the presence of 0.5 mg/mL recAAT, however, LPS/spike-induced TNF-α and IL-1ß mRNA expression and protein release were significantly inhibited (by about 46-50%) relative to LPS/spike alone. Although without statistical significance, recAAT also reduced production of IL-6 and IL-8. Notably, under the same experimental conditions, the plasma-derived AAT preparation Respreeza (used in native and oxidized forms) did not show significant effects. Our findings imply that an early pro-inflammatory activation of human PBMCs is better controlled by the recombinant version of AAT than the human plasma-derived AAT used here. Considering the increasing clinical interest in AAT therapy as useful to ameliorate the hyper-inflammation seen during COVID-19 infection, different AAT preparations require careful evaluation.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Leukocytes, Mononuclear/metabolism , Spike Glycoprotein, Coronavirus/metabolism , alpha 1-Antitrypsin/pharmacology , Animals , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/immunology , CHO Cells , COVID-19/therapy , Cells, Cultured , Cricetulus , Cytokines/metabolism , Humans , Inflammation/metabolism , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Lipopolysaccharides/immunology , Lipopolysaccharides/toxicity , SARS-CoV-2/drug effects , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , alpha 1-Antitrypsin/chemistry , alpha 1-Antitrypsin/immunology
6.
Int J Mol Sci ; 22(14)2021 Jul 14.
Article in English | MEDLINE | ID: covidwho-1314666

ABSTRACT

Proinflammatory cytokine production following infection with severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) is associated with poor clinical outcomes. Like SARS CoV-1, SARS CoV-2 enters host cells via its spike protein, which attaches to angiotensin-converting enzyme 2 (ACE2). As SARS CoV-1 spike protein is reported to induce cytokine production, we hypothesized that this pathway could be a shared mechanism underlying pathogenic immune responses. We herein compared the capabilities of Middle East Respiratory Syndrome (MERS), SARS CoV-1 and SARS CoV-2 spike proteins to induce cytokine expression in human peripheral blood mononuclear cells (PBMC). We observed that only specific commercial lots of SARS CoV-2 induce cytokine production. Surprisingly, recombinant SARS CoV-2 spike proteins from different vendors and batches exhibited different patterns of cytokine induction, and these activities were not inhibited by blockade of spike protein-ACE2 binding using either soluble ACE2 or neutralizing anti-S1 antibody. Moreover, commercial spike protein reagents contained varying levels of lipopolysaccharide (LPS), which correlated directly with their abilities to induce cytokine production. The LPS inhibitor, polymyxin B, blocked this cytokine induction activity. In addition, SARS CoV-2 spike protein avidly bound soluble LPS in vitro, rendering it a cytokine inducer. These results not only suggest caution in monitoring the purity of SARS CoV-2 spike protein reagents, but they indicate the possibility that interactions of SARS CoV-2 spike protein with LPS from commensal bacteria in virally infected mucosal tissues could promote pathogenic inflammatory cytokine production.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Cytokines/metabolism , Leukocytes, Mononuclear/metabolism , Lipopolysaccharides/pharmacology , Models, Biological , Spike Glycoprotein, Coronavirus/pharmacology , Healthy Volunteers , Humans , In Vitro Techniques , Leukocytes, Mononuclear/drug effects
7.
Parasit Vectors ; 14(1): 304, 2021 Jun 05.
Article in English | MEDLINE | ID: covidwho-1257961

ABSTRACT

BACKGROUND: Ivermectin is widely used in human and animal medicine to treat and prevent parasite nematode infections. It has been suggested that its mode of action requires the host immune system, as it is difficult to reproduce its clinical efficacy in vitro. We therefore studied the effects of a single dose of ivermectin (Stromectol®-0.15 mg/kg) on cytokine levels and immune cell gene expression in human volunteers. This dose reduces bloodstream microfilariae rapidly and for several months when given in mass drug administration programmes. METHODS: Healthy volunteers with no travel history to endemic regions were given 3-4 tablets, depending on their weight, of either ivermectin or a placebo. Blood samples were drawn immediately prior to administration, 4 h and 24 h afterwards, and complete blood counts performed. Serum levels of 41 cytokines and chemokines were measured using Luminex® and expression levels of 770 myeloid-cell-related genes determined using the NanoString nCounter®. Cytokine levels at 4 h and 24 h post-treatment were compared to the levels pre-treatment using simple t tests to determine if any individual results required further investigation, taking p = < 0.05 as the level of significance. NanoString data were analysed on the proprietary software, nSolver™. RESULTS: No significant differences were observed in complete blood counts or cytokine levels at either time point between people given ivermectin versus placebo. Only three genes showed a significant change in expression in peripheral blood mononuclear cells 4 h after ivermectin was given; there were no significant changes 24 h after drug administration or in polymorphonuclear cells at either time point. Leukocytes isolated from those participants given ivermectin showed no difference in their ability to kill Brugia malayi microfilariae in vitro. CONCLUSIONS: Overall, our data do not support a direct effect of ivermectin, when given at the dose used in current filarial elimination programmes, on the human immune system. Trial registration ClinicalTrials.gov NCT03459794 Registered 9th March 2018, Retrospectively registered https://clinicaltrials.gov/ct2/show/NCT03459794?term=NCT03459794&draw=2&rank=1 .


Subject(s)
Antiparasitic Agents/administration & dosage , Antiparasitic Agents/immunology , Cytokines/blood , Immunity, Innate/drug effects , Ivermectin/administration & dosage , Ivermectin/immunology , Leukocytes, Mononuclear/drug effects , Neutrophils/drug effects , Adolescent , Adult , Aged , Animals , Brugia malayi/drug effects , Cytokines/immunology , Gene Expression/drug effects , Human Experimentation , Humans , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/parasitology , Middle Aged , Neutrophils/immunology , Neutrophils/parasitology , Young Adult
8.
Int J Mol Sci ; 22(13)2021 Jun 23.
Article in English | MEDLINE | ID: covidwho-1282518

ABSTRACT

The usefulness of anti-inflammatory drugs as an adjunct therapy to improve outcomes in COVID-19 patients is intensely discussed in this paper. Willow bark (Salix cortex) has been used for centuries to relieve pain, inflammation, and fever. Its main active ingredient, salicin, is metabolized in the human body into salicylic acid, the precursor of the commonly used pain drug acetylsalicylic acid (ASA). Here, we report on the in vitro anti-inflammatory efficacy of two methanolic Salix extracts, standardized to phenolic compounds, in comparison to ASA in the context of a SARS-CoV-2 peptide challenge. Using SARS-CoV-2 peptide/IL-1ß- or LPS-activated human PBMCs and an inflammatory intestinal Caco-2/HT29-MTX co-culture, Salix extracts, and ASA concentration-dependently suppressed prostaglandin E2 (PGE2), a principal mediator of inflammation. The inhibition of COX-2 enzyme activity, but not protein expression was observed for ASA and one Salix extract. In activated PBMCs, the suppression of relevant cytokines (i.e., IL-6, IL-1ß, and IL-10) was seen for both Salix extracts. The anti-inflammatory capacity of Salix extracts was still retained after transepithelial passage and liver cell metabolism in an advanced co-culture model system consisting of intestinal Caco-2/HT29-MTX cells and differentiated hepatocyte-like HepaRG cells. Taken together, our in vitro data suggest that Salix extracts might present an additional anti-inflammatory treatment option in the context of SARS-CoV-2 peptides challenge; however, more confirmatory data are needed.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Aspirin/pharmacology , COVID-19/drug therapy , COVID-19/immunology , Plant Extracts/pharmacology , Anti-Inflammatory Agents/chemistry , Benzyl Alcohols/metabolism , COVID-19/virology , Caco-2 Cells , Cyclooxygenase 2/drug effects , Cytokines/metabolism , Dinoprostone/metabolism , Glucosides/metabolism , HT29 Cells , Humans , Inflammation , Leukocytes, Mononuclear/drug effects , Lipopolysaccharides/immunology , Plant Bark/chemistry , Plant Extracts/chemistry , SARS-CoV-2/immunology
9.
Inflammation ; 44(5): 1865-1877, 2021 Oct.
Article in English | MEDLINE | ID: covidwho-1188133

ABSTRACT

An understanding of the pathological inflammatory mechanisms involved in SARS-CoV-2 virus infection is necessary in order to discover new molecular pharmacological targets for SARS-CoV-2 cytokine storm. In this study, the effects of a recombinant SARS-CoV-2 spike glycoprotein S1 was investigated in human peripheral blood mononuclear cells (PBMCs). Stimulation of PBMCs with spike glycoprotein S1 (100 ng/mL) resulted in significant elevation in the production of TNFα, IL-6, IL-1ß and IL-8. However, pre-treatment with dexamethasone (100 nM) caused significant reduction in the release of these cytokines. Further experiments revealed that S1 stimulation of PBMCs increased phosphorylation of NF-κB p65 and IκBα, and IκBα degradation. DNA binding of NF-κB p65 was also significantly increased following stimulation with spike glycoprotein S1. Treatment of PBMCs with dexamethasone (100 nM) or BAY11-7082 (1 µM) resulted in inhibition of spike glycoprotein S1-induced NF-κB activation. Activation of p38 MAPK by S1 was blocked in the presence of dexamethasone and SKF 86002. CRID3, but not dexamethasone pre-treatment, produced significant inhibition of S1-induced activation of NLRP3/caspase-1. Further experiments revealed that S1-induced increase in the production of TNFα, IL-6, IL-1ß and IL-8 was reduced in the presence of BAY11-7082 and SKF 86002, while CRID3 pre-treatment resulted in the reduction of IL-1ß production. These results suggest that SARS-CoV-2 spike glycoprotein S1 stimulated PBMCs to release pro-inflammatory cytokines through mechanisms involving activation of NF-κB, p38 MAPK and NLRP3 inflammasome. It is proposed that the clinical benefits of dexamethasone in COVID-19 are possibly due to its anti-inflammatory activity in reducing SARS-CoV-2 cytokine storm.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Cytokine Release Syndrome/virology , Cytokines/metabolism , Dexamethasone/pharmacology , Leukocytes, Mononuclear/metabolism , SARS-CoV-2/pathogenicity , Spike Glycoprotein, Coronavirus/immunology , Anti-Inflammatory Agents/therapeutic use , Biomarkers/metabolism , Blotting, Western , COVID-19/drug therapy , COVID-19/immunology , COVID-19/virology , Cells, Cultured , Cytokine Release Syndrome/drug therapy , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/metabolism , Dexamethasone/therapeutic use , Enzyme-Linked Immunosorbent Assay , Humans , Leukocytes, Mononuclear/drug effects , Recombinant Proteins/immunology , SARS-CoV-2/immunology
10.
EBioMedicine ; 66: 103341, 2021 Apr.
Article in English | MEDLINE | ID: covidwho-1184943

ABSTRACT

BACKGROUND: Despite an impressive effort in clinical research, no standard therapeutic approach for coronavirus disease 2019 (COVID-19) patients has been established, highlighting the need to identify early biomarkers for predicting disease progression and new therapeutic interventions for patient management. The present study aimed to evaluate the involvement of the human endogenous retrovirus -W envelope (HERV-W ENV) in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection considering recent findings that HERVs are activated in response to infectious agents and lead to various immunopathological effects. We analysed HERV-W ENV expression in blood cells of COVID-19 patients in correlation with clinical characteristics and have discussed its potential role in the outcome of the disease. METHODS: We analysed HERV-W ENV expression in blood samples of COVID-19 patients and healthy donors by flow cytometry and quantitative reverse transcriptase PCR analysis, and evaluated its correlation with clinical signs, inflammatory markers, cytokine expression, and disease progression. FINDINGS: HERV-W ENV was highly expressed in the leukocytes of COVID-19 patients but not in those of healthy donors. Its expression correlated with the markers of T-cell differentiation and exhaustion and blood cytokine levels. The percentage of HERV-W ENV-positive lymphocytes correlated with inflammatory markers and pneumonia severity in COVID-19 patients. Notably, HERV-W ENV expression reflects the respiratory outcome of patients during hospitalization. INTERPRETATION: Given the known immuno- and neuro-pathogenicity of HERV-W ENV protein, it could promote certain pathogenic features of COVID-19 and therefore serve as a biomarker to predict clinical progression of disease and open to further studies for therapeutic intervention. FUNDING: Information available at the end of the manuscript.


Subject(s)
COVID-19/virology , Gene Products, env/metabolism , Pregnancy Proteins/metabolism , T-Lymphocytes/virology , Aged , Antiviral Agents/therapeutic use , COVID-19/etiology , COVID-19/therapy , Case-Control Studies , Cell Differentiation , Cytokines/metabolism , Endogenous Retroviruses , Female , Gene Products, env/genetics , Hospitalization , Humans , Interleukin-6/blood , Interleukin-6/pharmacology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/metabolism , Male , Middle Aged , Pneumonia, Viral/diagnostic imaging , Pneumonia, Viral/therapy , Pneumonia, Viral/virology , Pregnancy Proteins/genetics , Severity of Illness Index , T-Lymphocytes/metabolism , Treatment Outcome
11.
Molecules ; 25(11)2020 Jun 11.
Article in English | MEDLINE | ID: covidwho-981163

ABSTRACT

Flavonoids are widely used as phytomedicines. Here, we report on flavonoid phytomedicines with potential for development into prophylactics or therapeutics against coronavirus disease 2019 (COVID-19). These flavonoid-based phytomedicines include: caflanone, Equivir, hesperetin, myricetin, and Linebacker. Our in silico studies show that these flavonoid-based molecules can bind with high affinity to the spike protein, helicase, and protease sites on the ACE2 receptor used by the severe acute respiratory syndrome coronavirus 2 to infect cells and cause COVID-19. Meanwhile, in vitro studies show potential of caflanone to inhibit virus entry factors including, ABL-2, cathepsin L, cytokines (IL-1ß, IL-6, IL-8, Mip-1α, TNF-α), and PI4Kiiiß as well as AXL-2, which facilitates mother-to-fetus transmission of coronavirus. The potential for the use of smart drug delivery technologies like nanoparticle drones loaded with these phytomedicines to overcome bioavailability limitations and improve therapeutic efficacy are discussed.


Subject(s)
Antiviral Agents/pharmacology , Betacoronavirus/drug effects , Coronavirus Infections/drug therapy , Coronavirus OC43, Human/drug effects , Flavonoids/pharmacology , Peptidyl-Dipeptidase A/chemistry , Pneumonia, Viral/drug therapy , Spike Glycoprotein, Coronavirus/chemistry , Angiotensin-Converting Enzyme 2 , Animals , Antiviral Agents/chemistry , Betacoronavirus/chemistry , Betacoronavirus/growth & development , Binding Sites , COVID-19 , Chloroquine/chemistry , Chloroquine/pharmacology , Coronavirus Infections/genetics , Coronavirus OC43, Human/chemistry , Coronavirus OC43, Human/growth & development , Drug Carriers/administration & dosage , Drug Carriers/chemistry , Flavonoids/chemistry , Humans , Interleukins/antagonists & inhibitors , Interleukins/chemistry , Interleukins/genetics , Interleukins/metabolism , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/virology , Lung/drug effects , Lung/pathology , Lung/virology , Mice , Molecular Docking Simulation , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Pandemics , Peptidyl-Dipeptidase A/genetics , Peptidyl-Dipeptidase A/metabolism , Phytotherapy/methods , Pneumonia, Viral/genetics , Primary Cell Culture , Protein Binding , Protein Interaction Domains and Motifs , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/chemistry , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/antagonists & inhibitors , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Thermodynamics , Virus Internalization/drug effects
12.
J Transl Med ; 18(1): 452, 2020 11 30.
Article in English | MEDLINE | ID: covidwho-948411

ABSTRACT

BACKGROUND: Dysregulation of transcription and cytokine expression has been implicated in the pathogenesis of a variety inflammatory diseases. The resulting imbalance between inflammatory and resolving transcriptional programs can cause an overabundance of pro-inflammatory, classically activated macrophage type 1 (M1) and/or helper T cell type 1 (Th1) products, such as IFNγ, TNFα, IL1-ß, and IL12, that prevent immune switching to resolution and healing. The low molecular weight fraction of human serum albumin (LMWF5A) is a novel biologic drug that is currently under clinical investigation for the treatment of osteoarthritis and the hyper-inflammatory response associated with COVID-19. This study aims to elucidate transcriptional mechanisms of action involved with the ability of LMWF5A to reduce pro-inflammatory cytokine release. METHODS: ELISA arrays were used to identify cytokines and chemokines influenced by LMWF5A treatment of LPS-stimulated peripheral blood mononuclear cells (PBMC). The resulting profiles were analyzed by gene enrichment to gain mechanistic insight into the biologic processes and transcription factors (TFs) underlying the identified differentially expressed cytokines. DNA-binding ELISAs, luciferase reporter assays, and TNFα or IL-1ß relative potency were then employed to confirm the involvement of enriched pathways and TFs. RESULTS: LMWF5A was found to significantly inhibit a distinct set of pro-inflammatory cytokines (TNFα, IL-1ß, IL-12, CXCL9, CXCL10, and CXCL11) associated with pro-inflammatory M1/Th1 immune profiles. Gene enrichment analysis also suggests these cytokines are, in part, regulated by NF-κB and STAT transcription factors. Data from DNA-binding and reporter assays support this with LMWF5A inhibition of STAT1α DNA-binding activity as well as a reduction in overall NF-κB-driven luciferase expression. Experiments using antagonists specific for the immunomodulatory and NF-κB/STAT-repressing transcription factors, peroxisome proliferator-activated receptor (PPAR)γ and aryl hydrocarbon receptor (AhR), indicate these pathways are involved in the LMWF5A mechanisms of action by reducing LMWF5A drug potency as measured by TNFα and IL-1ß release. CONCLUSION: In this report, we provide evidence that LMWF5A reduces pro-inflammatory cytokine release by activating the immunoregulatory transcription factors PPARγ and AhR. In addition, our data indicate that LMWF5A suppresses NF-κB and STAT1α pro-inflammatory pathways. This suggests that LMWF5A acts through these mechanisms to decrease pro-inflammatory transcription factor activity and subsequent inflammatory cytokine production.


Subject(s)
Cytokines/metabolism , Inflammation/prevention & control , Leukocytes, Mononuclear/drug effects , Serum Albumin, Human/pharmacology , Anti-Inflammatory Agents/pharmacology , COVID-19/drug therapy , COVID-19/immunology , COVID-19/pathology , Cells, Cultured , Gene Expression Regulation/drug effects , HEK293 Cells , Humans , Inflammation/genetics , Inflammation/metabolism , Inflammation Mediators/metabolism , Interferon-Stimulated Gene Factor 3/metabolism , Leukocytes, Mononuclear/metabolism , Lipopolysaccharides , Lymphocyte Activation/drug effects , Molecular Weight , NF-kappa B/metabolism , Serum Albumin, Human/chemistry , Signal Transduction/drug effects , Signal Transduction/genetics , Signal Transduction/immunology , Transcription Factors/metabolism
13.
EBioMedicine ; 61: 103039, 2020 Nov.
Article in English | MEDLINE | ID: covidwho-838297

ABSTRACT

The signalling receptor for LPS, CD14, is a key marker of, and facilitator for, pro-inflammatory macrophage function. Pro-inflammatory macrophage differentiation remains a process facilitating a broad array of disease pathologies, and has recently emerged as a potential target against cytokine storm in COVID19. Here, we perform a whole-genome CRISPR screen to identify essential nodes regulating CD14 expression in myeloid cells, using the differentiation of THP-1 cells as a starting point. This strategy uncovers many known pathways required for CD14 expression and regulating macrophage differentiation while additionally providing a list of novel targets either promoting or limiting this process. To speed translation of these results, we have then taken the approach of independently validating hits from the screen using well-curated small molecules. In this manner, we identify pharmacologically tractable hits that can either increase CD14 expression on non-differentiated monocytes or prevent CD14 upregulation during macrophage differentiation. An inhibitor for one of these targets, MAP2K3, translates through to studies on primary human monocytes, where it prevents upregulation of CD14 following M-CSF induced differentiation, and pro-inflammatory cytokine production in response to LPS. Therefore, this screening cascade has rapidly identified pharmacologically tractable nodes regulating a critical disease-relevant process.


Subject(s)
Cell Differentiation/drug effects , Lipopolysaccharide Receptors/metabolism , Macrophages/immunology , Macrophages/metabolism , Biomarkers , Cells, Cultured , Cytokines/metabolism , Humans , Immunophenotyping , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Lipopolysaccharides/adverse effects , Macrophages/drug effects , THP-1 Cells
14.
Biomed Pharmacother ; 131: 110643, 2020 Nov.
Article in English | MEDLINE | ID: covidwho-726408

ABSTRACT

Coronavirus Disease 2019 (COVID-19) caused by Severe Acute Respiratory Syndrome Coronavirus -2 (SARS-CoV-2) has been widely spread in the world with a high mortality. Cytokine storm syndrome (CSS) and acute lung injury caused by SARS-CoV-2 infection severely threaten the patients. With the purpose to find effective and low-toxic drugs to mitigate CSS, entecavir and imipenem were identified to reduce TNF-α using a LPS-induced macrophage model from the anti-infective drug library. Entecavir and imipenem efficiently suppressed the release of inflammatory cytokines by partly intervention of NF-κB activity. The acute lung injury was also alleviated and the survival time was prolonged in mice. In addition, entecavir and imipenem inhibited the release of TNF-α and IL-10 in human peripheral blood mononuclear cells (hPBMCs). Collectively, we proposed that entecavir and imipenem might be candidates for the treatment of CSS.


Subject(s)
Coronavirus Infections/drug therapy , Cytokine Release Syndrome/drug therapy , Guanine/analogs & derivatives , Imipenem/pharmacology , Pneumonia, Viral/drug therapy , Acute Lung Injury/drug therapy , Acute Lung Injury/virology , Animals , COVID-19 , Coronavirus Infections/complications , Cytokine Release Syndrome/virology , Cytokines/immunology , Drug Repositioning , Guanine/pharmacology , Humans , Interleukin-10/immunology , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/immunology , Lipopolysaccharides , Macrophages/immunology , Male , Mice , Mice, Inbred C57BL , Pandemics , Pneumonia, Viral/complications , Tumor Necrosis Factor-alpha/drug effects , Tumor Necrosis Factor-alpha/metabolism
15.
Sci Adv ; 6(31)2020 07.
Article in English | MEDLINE | ID: covidwho-725277

ABSTRACT

The outbreak of the highly contagious and deadly severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), also known as coronavirus disease 2019 (COVID-19), has posed a serious threat to public health across the globe, calling for the development of effective diagnostic markers and therapeutics. Here, we report a highly reliable severity diagnostic biomarker, acetylated 676th lysine transforming growth factor-beta-induced protein (TGFBIp K676Ac). TGFBIp K676Ac was consistently elevated in the blood of patients with SARS-CoV-2 pneumonia (n = 113), especially in patients in the intensive care unit (ICU) compared to non-ICU patients. Patients' blood samples showed increased cytokines and lymphopenia, which are exemplary indicators of SARS-CoV-2 pneumonia. Treatment with TGFBIp neutralizing antibodies suppressed the cytokine storm. The increased level of TGFBIp K676Ac in ICU patients suggests the promise of this protein as a reliable severity diagnostic biomarker for severe SARS-CoV-2 disease.


Subject(s)
Betacoronavirus/pathogenicity , Coronavirus Infections/diagnosis , Cytokine Release Syndrome/diagnosis , Extracellular Matrix Proteins/immunology , Leukocytes, Mononuclear/immunology , Pneumonia, Viral/diagnosis , Protein Processing, Post-Translational , Respiratory Insufficiency/diagnosis , Transforming Growth Factor beta/immunology , Acetylation , Antibodies, Neutralizing/pharmacology , Betacoronavirus/immunology , Biomarkers/blood , COVID-19 , Case-Control Studies , Coronavirus Infections/blood , Coronavirus Infections/immunology , Coronavirus Infections/pathology , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/immunology , Cytokine Release Syndrome/pathology , Extracellular Matrix Proteins/antagonists & inhibitors , Extracellular Matrix Proteins/genetics , Gene Expression , Humans , Intensive Care Units , Leukocyte Count , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/pathology , Leukocytes, Mononuclear/virology , Lung/blood supply , Lung/drug effects , Lung/pathology , Lung/virology , Lysine/metabolism , NF-kappa B/genetics , NF-kappa B/immunology , Pandemics , Pneumonia, Viral/blood , Pneumonia, Viral/immunology , Pneumonia, Viral/pathology , Primary Cell Culture , Prognosis , Respiratory Insufficiency/blood , Respiratory Insufficiency/immunology , Respiratory Insufficiency/pathology , SARS-CoV-2 , Severity of Illness Index , Transforming Growth Factor beta/antagonists & inhibitors , Transforming Growth Factor beta/genetics
16.
Biomed Pharmacother ; 131: 110653, 2020 Nov.
Article in English | MEDLINE | ID: covidwho-722385

ABSTRACT

BACKGROUND: Angiotensin receptor blockers (ARBs) reducing inflammation and protecting lung and brain function, could be of therapeutic efficacy in COVID-19 patients. METHODS: Using GSEA, we compared our previous transcriptome analysis of neurons injured by glutamate and treated with the ARB Candesartan (GSE67036) with transcriptional signatures from SARS-CoV-2 infected primary human bronchial epithelial cells (NHBE) and lung postmortem (GSE147507), PBMC and BALF samples (CRA002390) from COVID-19 patients. RESULTS: Hundreds of genes upregulated in SARS-CoV-2/COVID-19 transcriptomes were similarly upregulated by glutamate and normalized by Candesartan. Gene Ontology analysis revealed expression profiles with greatest significance and enrichment, including proinflammatory cytokine and chemokine activity, the NF-kappa B complex, alterations in innate and adaptive immunity, with many genes participating in the COVID-19 cytokine storm. CONCLUSIONS: There are similar injury mechanisms in SARS-CoV-2 infection and neuronal injury, equally reduced by ARB treatment. This supports the hypothesis of a therapeutic role for ARBs, ameliorating the COVID-19 cytokine storm.


Subject(s)
Angiotensin II Type 1 Receptor Blockers/pharmacology , Benzimidazoles/pharmacology , Coronavirus Infections/drug therapy , Cytokine Release Syndrome/drug therapy , Pneumonia, Viral/drug therapy , Tetrazoles/pharmacology , Betacoronavirus/genetics , Betacoronavirus/isolation & purification , Biphenyl Compounds , Bronchi/cytology , Bronchoalveolar Lavage Fluid/virology , COVID-19 , Coronavirus Infections/complications , Coronavirus Infections/virology , Cytokine Release Syndrome/virology , Epithelial Cells/drug effects , Epithelial Cells/virology , Gene Expression Profiling , Humans , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/virology , Pandemics , Pneumonia, Viral/complications , Pneumonia, Viral/virology , SARS-CoV-2 , Transcriptome
17.
J Thromb Thrombolysis ; 51(1): 226-231, 2021 Jan.
Article in English | MEDLINE | ID: covidwho-648995

ABSTRACT

Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection has been linked to a higher risk of mortality compared to influenza, which is mainly due to severe secondary diseases, such as acute respiratory distress syndrome (ARDS). In turn, ARDS is characterized by an acute inflammation and an excessive activity of the coagulation cascade, rising the vulnerability for venous thromboembolic events. In order to investigate the relation of inflammation and the influence of coagulation factors on their release, human peripheral mononuclear blood cells (PBMCs) were treated with autologous serum, heparinized plasma and different doses of fibrin. Thereafter, the concentration of pro-inflammatory cytokines and chemokines in the secretome of PBMCs was measured by enzyme-linked immunosorbent assay. Our analyses revealed autologous serum to significantly increase the secretion of cytokines and chemokines after 24 h of incubation time. Furthermore, the addition of fibrin markedly increased the secretion of cytokines and chemokines by PBMCs in a dose-dependent manner. Consequently, in accordance with previous studies, our study outlines that anti-coagulation may constitute a promising tool for the treatment of SARS-CoV-2, reducing both, the cytokine storm, as well as the risk for thrombotic complications.


Subject(s)
Blood Coagulation , COVID-19/therapy , Cytokine Release Syndrome , Fibrin , Inflammation , Blood Coagulation/drug effects , Blood Coagulation/immunology , COVID-19/blood , COVID-19/drug therapy , COVID-19/immunology , Cells, Cultured , Chemokines/immunology , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/drug therapy , Cytokine Release Syndrome/etiology , Dose-Response Relationship, Drug , Fibrin/immunology , Fibrin/pharmacology , Fibrinolytic Agents/pharmacology , Heparin/pharmacology , Humans , Immunization, Passive , Inflammation/blood , Inflammation/therapy , Leukocytes, Mononuclear/drug effects , SARS-CoV-2
18.
Circ Arrhythm Electrophysiol ; 13(8): e008627, 2020 08.
Article in English | MEDLINE | ID: covidwho-641777

ABSTRACT

BACKGROUND: During acute infections, the risk of malignant ventricular arrhythmias is increased, partly because of a higher propensity to develop QTc prolongation. Although it is generally believed that QTc changes almost exclusively result from concomitant treatment with QT-prolonging antimicrobials, direct effects of inflammatory cytokines on ventricular repolarization are increasingly recognized. We hypothesized that systemic inflammation per se can significantly prolong QTc during acute infections, via cytokine-mediated changes in K+ channel expression. METHODS: We evaluated (1) the frequency of QTc prolongation and its association with inflammatory markers, in patients with different types of acute infections, during active disease and remission; (2) the prevalence of acute infections in a cohort of consecutive patients with Torsades de Pointes; (3) the relationship between K+ channel mRNA levels in ventricles and peripheral blood mononuclear cells and their changes in patients with acute infection over time. RESULTS: In patients with acute infections, regardless of concomitant QT-prolonging antimicrobial treatments, QTc was significantly prolonged but rapidly normalized in parallel to CRP (C-reactive protein) and cytokine level reduction. Consistently in the Torsades de Pointes cohort, concomitant acute infections were highly prevalent (30%), despite only a minority (25%) of these cases were treated with QT-prolonging antimicrobials. KCNJ2 K+ channel expression in peripheral blood mononuclear cell, which strongly correlated to that in ventricles, inversely associated to CRP and IL (interleukin)-1 changes in acute infection patients. CONCLUSIONS: During acute infections, systemic inflammation rapidly induces cytokine-mediated ventricular electrical remodeling and significant QTc prolongation, regardless concomitant antimicrobial therapy. Although transient, these changes may significantly increase the risk of life-threatening ventricular arrhythmia in these patients. It is timely and warranted to transpose these findings to the current coronavirus disease 2019 (COVID-19) pandemic, in which both increased amounts of circulating cytokines and cardiac arrhythmias are demonstrated along with a frequent concomitant treatment with several QT-prolonging drugs. Graphic Abstract: A graphic abstract is available for this article.


Subject(s)
Communicable Diseases/metabolism , Cytokines/metabolism , Heart Arrest/metabolism , Heart Rate , Heart Ventricles/metabolism , Inflammation/metabolism , Leukocytes, Mononuclear/metabolism , Potassium Channels, Inwardly Rectifying/metabolism , Torsades de Pointes/metabolism , Action Potentials , Acute Disease , Adult , Aged , Aged, 80 and over , Anti-Infective Agents/adverse effects , Communicable Diseases/drug therapy , Communicable Diseases/epidemiology , Communicable Diseases/physiopathology , Female , Heart Arrest/epidemiology , Heart Arrest/physiopathology , Heart Rate/drug effects , Heart Ventricles/drug effects , Heart Ventricles/physiopathology , Humans , Inflammation/epidemiology , Inflammation/physiopathology , Leukocytes, Mononuclear/drug effects , Male , Middle Aged , Potassium Channels, Inwardly Rectifying/genetics , Prevalence , Risk Factors , Signal Transduction , Time Factors , Torsades de Pointes/epidemiology , Torsades de Pointes/physiopathology , Young Adult
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