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1.
Arch Virol ; 166(8): 2285-2289, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1826502

ABSTRACT

Mesenchymal stromal cells (MSCs) are considered multipotent progenitors with the capacity to differentiate into mesoderm-like cells in many species. The immunosuppressive properties of MSCs are important for downregulating inflammatory responses. Turkey coronavirus (TCoV) is the etiological agent of a poult mortality syndrome that affects intestinal epithelial cells. In this study, poult MSCs were isolated, characterized, and infected with TCoV after in vitro culture. The poult-derived MSCs showed fibroblast-like morphology and the ability to undergo differentiation into mesodermal-derived cells and to support virus replication. Infection with TCoV resulted in cytopathic effects and the loss of cell viability. TCoV antigens and new viral progeny were detected at high levels, as were transcripts of the pro-inflammatory factors INFγ, IL-6, and IL-8. These findings suggest that the cytokine storm phenomenon is not restricted to one genus of the family Coronaviridae and that MSCs cannot always balance the process.


Subject(s)
Coronavirus, Turkey/physiology , Cytokines/metabolism , Virus Replication , Animals , Cell Differentiation , Cell Survival , Cytopathogenic Effect, Viral , Interferon-gamma/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/virology , Turkeys , Up-Regulation
2.
Oxid Med Cell Longev ; 2022: 9366494, 2022.
Article in English | MEDLINE | ID: covidwho-1807713

ABSTRACT

Trehalose, a natural disaccharide, is synthesized by many organisms when cells are exposed to stressful stimuli. On the basis of its ability to modulate autophagy, trehalose has been considered an innovative drug for ameliorating many diseases, but its molecular mechanism is not well described. Previous findings demonstrated that trehalose plays a photoprotective role against ultraviolet (UV) B-induced damage through autophagy induction in keratinocytes. In this study, coimmunoprecipitation, label-free quantitative proteomic and parallel reaction monitoring, and western blot analysis demonstrated that trehalose promotes the interaction between tissue inhibitor of metalloproteinase (TIMP) 3 and Beclin1. Western blot and immunofluorescence staining analysis suggested that trehalose increases ATG9A localization in lysosomes and decreases its localization in the endoplasmic reticulum. Furthermore, in the presence or absence of UVB radiation, we evaluated the influence of TIMP3 and ATG9A small interfering RNA (siRNA) on the effect of trehalose on autophagy, cell death, migration, or interleukin-8 expression in keratinocytes, including HaCaT, A431, and human epidermal keratinocytes. The results revealed that in HaCaT cells, TIMP3 and ATG9A siRNA resulted in attenuation of trehalose-induced autophagy and inhibited cell death. In A431 cells, TIMP3 and ATG9A siRNA led to attenuation of trehalose-induced autophagy and cell death and inhibited migration. In human epidermal keratinocytes, trehalose-induced autophagy and inhibition of the interleukin-8 expression were blocked by ATG9A but not TIMP3 siRNA. In addition, the results of quantitative real-time PCR and immunohistochemistry analysis demonstrated the abnormal expression of TIMP3 and ATG9A in actinic keratosis and cutaneous squamous cell carcinoma skin tissues. These findings suggest the protective effects of trehalose in normal keratinocytes and its inhibitory effects on cancerous keratinocytes, possibly mediated by activation of autophagy and regulation of TIMP3 and ATG9A, providing the mechanistic basis for the potential use of trehalose in the prevention or treatment of UVB-induced skin diseases.


Subject(s)
Carcinoma, Squamous Cell , Skin Neoplasms , Autophagy , Autophagy-Related Proteins/metabolism , Carcinoma, Squamous Cell/pathology , Humans , Interleukin-8/metabolism , Keratinocytes/metabolism , Membrane Proteins/metabolism , Proteomics , RNA, Small Interfering/metabolism , Skin Neoplasms/metabolism , Tissue Inhibitor of Metalloproteinase-3/genetics , Tissue Inhibitor of Metalloproteinase-3/metabolism , Tissue Inhibitor of Metalloproteinase-3/pharmacology , Trehalose/pharmacology , Ultraviolet Rays/adverse effects , Vesicular Transport Proteins/metabolism
3.
PLoS Pathog ; 18(4): e1010468, 2022 04.
Article in English | MEDLINE | ID: covidwho-1779781

ABSTRACT

An overreactive inflammatory response and coagulopathy are observed in patients with severe form of COVID-19. Since increased levels of D-dimer (DD) are associated with coagulopathy in COVID-19, we explored whether DD contributes to the aberrant cytokine responses. Here we show that treatment of healthy human monocytes with DD induced a dose dependent increase in production of pyrogenic mediator, Prostaglandin E2 (PGE2) and inflammatory cytokines, IL-6 and IL-8. The DD-induced PGE2 and inflammatory cytokines were enhanced significantly by co-treatment with immune complexes (IC) of SARS CoV-2 recombinant S protein or of pseudovirus containing SARS CoV-2 S protein (PVCoV-2) coated with spike-specific chimeric monoclonal antibody (MAb) containing mouse variable and human Fc regions. The production of PGE2 and cytokines in monocytes activated with DD and ICs was sensitive to the inhibitors of ß2 integrin and FcγRIIa, and to the inhibitors of calcium signaling, Mitogen-Activated Protein Kinase (MAPK) pathway, and tyrosine-protein kinase. Importantly, strong increase in PGE2 and in IL-6/IL-8/IL-1ß cytokines was observed in monocytes activated with DD in the presence of IC of PVCoV-2 coated with plasma from hospitalized COVID-19 patients but not from healthy donors. The IC of PVCoV-2 with convalescent plasma induced much lower levels of PGE2 and cytokines compared with plasma from hospitalized COVID-19 patients. PGE2 and IL-6/IL-8 cytokines produced in monocytes activated with plasma-containing IC, correlated well with the levels of spike binding antibodies and not with neutralizing antibody titers. Our study suggests that a combination of high levels of DD and high titers of spike-binding antibodies that can form IC with SARS CoV-2 viral particles might accelerate the inflammatory status of lung infiltrating monocytes leading to increased lung pathology in patients with severe form of COVID-19.


Subject(s)
COVID-19 , Monocytes , Animals , Antigen-Antibody Complex , COVID-19/therapy , Cytokines/metabolism , Dinoprostone/metabolism , Fibrin Fibrinogen Degradation Products , Humans , Immunization, Passive , Immunologic Factors/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Mice , Spike Glycoprotein, Coronavirus/metabolism
4.
Int J Mol Sci ; 23(7)2022 Apr 06.
Article in English | MEDLINE | ID: covidwho-1776254

ABSTRACT

In the novel pandemic of Coronavirus Disease 2019, high levels of pro-inflammatory cytokines lead to endothelial activation and dysfunction, promoting a pro-coagulative state, thrombotic events, and microvasculature injuries. The aim of the present work was to investigate the effect of SARS-CoV-2 on pro-inflammatory cytokines, tissue factor, and chemokine release, with Human Microvascular Endothelial Cells (HMEC-1). ACE2 receptor expression was evaluated by western blot analysis. SARS-CoV-2 infection was assessed by one-step RT-PCR until 7 days post-infection (p.i.), and by Transmission Electron Microscopy (TEM). IL-6, TNF-α, IL-8, IFN-α, and hTF mRNA expression levels were detected by RT-PCR, while cytokine release was evaluated by ELISA. HMEC-1 expressed ACE2 receptor and SARS-CoV-2 infection showed a constant viral load. TEM analysis showed virions localized in the cytoplasm. Expression of IL-6 at 24 h and IFN-α mRNA at 24 h and 48 h p.i. was higher in infected than uninfected HMEC-1 (p < 0.05). IL-6 levels were significantly higher in supernatants from infected HMEC-1 (p < 0.001) at 24 h, 48 h, and 72 h p.i., while IL-8 levels were significantly lower at 24 h p.i. (p < 0.001). These data indicate that in vitro microvascular endothelial cells are susceptible to SARS-CoV-2 infection but slightly contribute to viral amplification. However, SARS-CoV-2 infection might trigger the increase of pro-inflammatory mediators.


Subject(s)
COVID-19 , Angiotensin-Converting Enzyme 2 , Chemokines/genetics , Chemokines/metabolism , Cytokines/metabolism , Endothelial Cells/metabolism , Humans , Interleukin-6/genetics , Interleukin-6/metabolism , Interleukin-8/genetics , Interleukin-8/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , SARS-CoV-2
5.
J Reprod Immunol ; 151: 103501, 2022 Jun.
Article in English | MEDLINE | ID: covidwho-1763858

ABSTRACT

While COVID-19 infection during pregnancy is common, fetal transmission is rare, suggesting that intrauterine mechanisms form an effective blockade against SARS-CoV-2. Key among these is the decidual immune environment of the placenta. We hypothesize that decidual leukocytes are altered by maternal SARS-CoV-2 infection in pregnancy and that this decidual immune response is shaped by the timing of infection during gestation. To address this hypothesis, we collected decidua basalis tissues at delivery from women with symptomatic COVID-19 during second (2nd Tri COVID, n = 8) or third trimester (3rd Tri COVID, n = 8) and SARS-CoV-2-negative controls (Control, n = 8). Decidual natural killer (NK) cells, macrophages and T cells were evaluated using quantitative microscopy, and pro- and anti-inflammatory cytokine mRNA expression was evaluated using quantitative reverse transcriptase PCR (qRT-PCR). When compared with the Control group, decidual tissues from 3rd Tri COVID exhibited significantly increased macrophages, NK cells and T cells, whereas 2nd Tri COVID only had significantly increased T cells. In evaluating decidual cytokine expression, we noted that IL-6, IL-8, IL-10 and TNF-α were significantly correlated with macrophage cell abundance. However, in 2nd Tri COVID tissues, there was significant downregulation of IL-6, IL-8, IL-10, and TNF-α. Taken together, these results suggest innate and adaptive immune responses are present at the maternal-fetal interface in maternal SARS-CoV-2 infections late in pregnancy, and that infections earlier in pregnancy show evidence of a resolving immune response. Further studies are warranted to characterize the full scope of intrauterine immune responses in pregnancies affected by maternal COVID-19.


Subject(s)
COVID-19 , Pregnancy Complications, Infectious , Cytokines/metabolism , Decidua , Female , Humans , Immunity , Interleukin-10/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Pregnancy , Pregnancy Complications, Infectious/metabolism , SARS-CoV-2 , Tumor Necrosis Factor-alpha/metabolism
6.
J Virol ; 96(5): e0208621, 2022 03 09.
Article in English | MEDLINE | ID: covidwho-1736026

ABSTRACT

Coronavirus infections induce the expression of multiple proinflammatory cytokines and chemokines. We have previously shown that in cells infected with gammacoronavirus infectious bronchitis virus (IBV), interleukin 6 (IL-6), and IL-8 were drastically upregulated, and the MAP kinase p38 and the integrated stress response pathways were implicated in this process. In this study, we report that coronavirus infection activates a negative regulatory loop that restricts the upregulation of a number of proinflammatory genes. As revealed by the initial transcriptomic and subsequent validation analyses, the anti-inflammatory adenine-uridine (AU)-rich element (ARE)-binding protein, zinc finger protein 36 (ZFP36), and its related family members were upregulated in cells infected with IBV and three other coronaviruses, alphacoronaviruses porcine epidemic diarrhea virus (PEDV), human coronavirus 229E (HCoV-229E), and betacoronavirus HCoV-OC43, respectively. Characterization of the functional roles of ZFP36 during IBV infection demonstrated that ZFP36 promoted the degradation of transcripts coding for IL-6, IL-8, dual-specificity phosphatase 1 (DUSP1), prostaglandin-endoperoxide synthase 2 (PTGS2) and TNF-α-induced protein 3 (TNFAIP3), through binding to AREs in these transcripts. Consistently, knockdown and inhibition of JNK and p38 kinase activities reduced the expression of ZFP36, as well as the expression of IL-6 and IL-8. On the contrary, overexpression of mitogen-activated protein kinase kinase 3 (MKK3) and MAPKAP kinase-2 (MK2), the upstream and downstream kinases of p38, respectively, increased the expression of ZFP36 and decreased the expression of IL-8. Taken together, this study reveals an important regulatory role of the MKK3-p38-MK2-ZFP36 axis in coronavirus infection-induced proinflammatory response. IMPORTANCE Excessive and uncontrolled induction and release of proinflammatory cytokines and chemokines, the so-called cytokine release syndrome (CRS), would cause life-threatening complications and multiple organ failure in severe coronavirus infections, including severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS) and COVID-19. This study reveals that coronavirus infection also induces the expression of ZFP36, an anti-inflammatory ARE-binding protein, promoting the degradation of ARE-containing transcripts coding for IL-6 and IL-8 as well as a number of other proteins related to inflammatory response. Furthermore, the p38 MAP kinase, its upstream kinase MKK3 and downstream kinase MK2 were shown to play a regulatory role in upregulation of ZFP36 during coronavirus infection cycles. This MKK3-p38-MK2-ZFP36 axis would constitute a potential therapeutic target for severe coronavirus infections.


Subject(s)
Coronavirus Infections/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Tristetraprolin/metabolism , p38 Mitogen-Activated Protein Kinases/metabolism , Adenine/metabolism , Animals , Cell Line , Chlorocebus aethiops , Coronavirus Infections/genetics , Gene Expression Regulation , Humans , Infectious bronchitis virus/metabolism , Infectious bronchitis virus/pathogenicity , Interleukin-6/genetics , Interleukin-8/genetics , Intracellular Signaling Peptides and Proteins/metabolism , Phosphorylation , Transcriptional Activation , Up-Regulation , Uridine/metabolism , Vero Cells
7.
Int J Mol Sci ; 23(2)2022 Jan 15.
Article in English | MEDLINE | ID: covidwho-1631216

ABSTRACT

Angiotensin II receptor type 1 (AT1R) and endothelin-1 receptor type A (ETAR) are G-protein-coupled receptors (GPCRs) expressed on the surface of a great variety of cells: immune cells, vascular smooth cells, endothelial cells, and fibroblasts express ETAR and AT1R, which are activated by endothelin 1 (ET1) and angiotensin II (AngII), respectively. Certain autoantibodies are specific for these receptors and can regulate their function, thus being known as functional autoantibodies. The function of these antibodies is similar to that of natural ligands, and it involves not only vasoconstriction, but also the secretion of proinflammatory cytokines (such as interleukin-6 (IL6), IL8 and TNF-α), collagen production by fibroblasts, and reactive oxygen species (ROS) release by fibroblasts and neutrophils. The role of autoantibodies against AT1R and ETAR (AT1R-AAs and ETAR-AAs, respectively) is well described in the pathogenesis of many medical conditions (e.g., systemic sclerosis (SSc) and SSc-associated pulmonary hypertension, cystic fibrosis, and allograft dysfunction), but their implications in cardiovascular diseases are still unclear. This review summarizes the current evidence regarding the effects of AT1R-AAs and ETAR-AAs in cardiovascular pathologies, highlighting their roles in heart transplantation and mechanical circulatory support, preeclampsia, and acute coronary syndromes.


Subject(s)
Autoantibodies/metabolism , Cardiovascular Diseases/immunology , Receptor, Angiotensin, Type 1/immunology , Receptor, Endothelin A/immunology , Cardiovascular Diseases/metabolism , Cardiovascular Diseases/pathology , Collagen/metabolism , Humans , Interleukin-6/metabolism , Interleukin-8/metabolism , Reactive Oxygen Species/metabolism , Receptor, Angiotensin, Type 1/metabolism , Tumor Necrosis Factor-alpha/metabolism
8.
Sci Rep ; 11(1): 20793, 2021 10 21.
Article in English | MEDLINE | ID: covidwho-1479813

ABSTRACT

In Europe, multiple waves of infections with SARS-CoV-2 (COVID-19) have been observed. Here, we have investigated whether common patterns of cytokines could be detected in individuals with mild and severe forms of COVID-19 in two pandemic waves, and whether machine learning approach could be useful to identify the best predictors. An increasing trend of multiple cytokines was observed in patients with mild or severe/critical symptoms of COVID-19, compared with healthy volunteers. Linear Discriminant Analysis (LDA) clearly recognized the three groups based on cytokine patterns. Classification and Regression Tree (CART) further indicated that IL-6 discriminated controls and COVID-19 patients, whilst IL-8 defined disease severity. During the second wave of pandemics, a less intense cytokine storm was observed, as compared with the first. IL-6 was the most robust predictor of infection and discriminated moderate COVID-19 patients from healthy controls, regardless of epidemic peak curve. Thus, serum cytokine patterns provide biomarkers useful for COVID-19 diagnosis and prognosis. Further definition of individual cytokines may allow to envision novel therapeutic options and pave the way to set up innovative diagnostic tools.


Subject(s)
COVID-19/blood , COVID-19/epidemiology , Cytokines/blood , Aged , Biomarkers/blood , COVID-19 Testing , Case-Control Studies , Cytokines/metabolism , Discriminant Analysis , Female , Humans , Interleukin-6/metabolism , Interleukin-8/metabolism , Italy/epidemiology , Machine Learning , Male , Middle Aged , Pandemics , Regression Analysis , SARS-CoV-2
9.
JCI Insight ; 6(18)2021 09 22.
Article in English | MEDLINE | ID: covidwho-1435144

ABSTRACT

Neutrophils provide a critical line of defense in immune responses to various pathogens, inflicting self-damage upon transition to a hyperactivated, procoagulant state. Recent work has highlighted proinflammatory neutrophil phenotypes contributing to lung injury and acute respiratory distress syndrome (ARDS) in patients with coronavirus disease 2019 (COVID-19). Here, we use state-of-the art mass spectrometry-based proteomics and transcriptomic and correlative analyses as well as functional in vitro and in vivo studies to dissect how neutrophils contribute to the progression to severe COVID-19. We identify a reinforcing loop of both systemic and neutrophil intrinsic IL-8 (CXCL8/IL-8) dysregulation, which initiates and perpetuates neutrophil-driven immunopathology. This positive feedback loop of systemic and neutrophil autocrine IL-8 production leads to an activated, prothrombotic neutrophil phenotype characterized by degranulation and neutrophil extracellular trap (NET) formation. In severe COVID-19, neutrophils directly initiate the coagulation and complement cascade, highlighting a link to the immunothrombotic state observed in these patients. Targeting the IL-8-CXCR-1/-2 axis interferes with this vicious cycle and attenuates neutrophil activation, degranulation, NETosis, and IL-8 release. Finally, we show that blocking IL-8-like signaling reduces severe acute respiratory distress syndrome of coronavirus 2 (SARS-CoV-2) spike protein-induced, human ACE2-dependent pulmonary microthrombosis in mice. In summary, our data provide comprehensive insights into the activation mechanisms of neutrophils in COVID-19 and uncover a self-sustaining neutrophil-IL-8 axis as a promising therapeutic target in severe SARS-CoV-2 infection.


Subject(s)
COVID-19/metabolism , Interleukin-8/metabolism , Lung/immunology , Neutrophils/immunology , SARS-CoV-2 , Thrombosis/etiology , Animals , COVID-19/complications , COVID-19/pathology , Humans , Lung/pathology , Mice , Neutrophil Activation , Neutrophils/pathology , Phenotype , Thrombosis/pathology
10.
Int Immunopharmacol ; 101(Pt A): 108192, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1433412

ABSTRACT

The mounting evidence regarding the pathogenesis of COVID-19 indicated that the cytokine storm has an axial role in the severity of this disease, which may lead to thrombotic complications, acute respiratory distress syndrome (ARDS), and myocardial damage, among other consequences. It has recently been demonstrated that statins are known to have anti-viral, anti-inflammatory, anti-thrombotic, and immunomodulatory features; however, their advantage has not been evaluated in COVID-19. This study aimed to investigate the protective effects of lovastatin in intensive care unit (ICU) patients with COVID-19. The case-control study consists of 284 ICU patients, which classified into three groups as follows: 1) the patients who no received lovastatin as a control (92 patients), 2) patients received 20 mg per day lovastatin (99 patients), and 3) patients received 40 mg per day lovastatin (93 patients). Each group's demographic and clinical parameters, along with CRP, interleukin (IL)-6, IL-8 levels, and mortality rate, were studied in three-time points. The results showed that there was no statistically significant difference between our study groups in terms of age and sex. (P > 0.05). Besides, in patients, receiving lovastatin the CRP, IL-6, IL-8 levels were significantly decreased from T1 to T3 than to the control group. Our results also showed that the use of lovastatin in COVID-19 patients significantly reduced the length of hospitalization in the ICU compared with the control group. In addition, our results showed that the mortality rate in patients receiving lovastatin was lower when compared to the control group; however, this difference was not statistically significant. Since the cytokine storm is a significant factor in the pathology of SARS-CoV-2, our findings highlighted the potential use of lovastatin to mitigate the inflammatory response induced by SARS-CoV-2 infection.


Subject(s)
Anti-Inflammatory Agents/pharmacology , COVID-19/drug therapy , Lovastatin/pharmacology , Adult , Anti-Inflammatory Agents/therapeutic use , COVID-19/blood , Case-Control Studies , Critical Care/methods , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/drug therapy , Cytokines/drug effects , Female , Hospitalization , Humans , Intensive Care Units , Interleukin-6/metabolism , Interleukin-8/metabolism , Lovastatin/therapeutic use , Male , Middle Aged , Receptors, Immunologic/metabolism , Sex Factors
11.
J Clin Immunol ; 41(7): 1607-1620, 2021 10.
Article in English | MEDLINE | ID: covidwho-1298388

ABSTRACT

The complement system, a network of highly-regulated proteins, represents a vital part of the innate immune response. Over-activation of the complement system plays an important role in inflammation, tissue damage, and infectious disease severity. The prevalence of MERS-CoV in Saudi Arabia remains significant and cases are still being reported. The role of complement in Middle East Respiratory Syndrome coronavirus (MERS-CoV) pathogenesis and complement-modulating treatment strategies has received limited attention, and studies involving MERS-CoV-infected patients have not been reported. This study offers the first insight into the pulmonary expression profile including seven complement proteins, complement regulatory factors, IL-8, and RANTES in MERS-CoV infected patients without underlying chronic medical conditions. Our results significantly indicate high expression levels of complement anaphylatoxins (C3a and C5a), IL-8, and RANTES in the lungs of MERS-CoV-infected patients. The upregulation of lung complement anaphylatoxins, C5a, and C3a was positively correlated with IL-8, RANTES, and the fatality rate. Our results also showed upregulation of the positive regulatory complement factor P, suggesting positive regulation of the complement during MERS-CoV infection. High levels of lung C5a, C3a, factor P, IL-8, and RANTES may contribute to the immunopathology, disease severity, ARDS development, and a higher fatality rate in MERS-CoV-infected patients. These findings highlight the potential prognostic utility of C5a, C3a, IL-8, and RANTES as biomarkers for MERS-CoV disease severity and mortality. To further explore the prediction of functional partners (proteins) of highly expressed proteins (C5a, C3a, factor P, IL-8, and RANTES), the computational protein-protein interaction (PPI) network was constructed, and six proteins (hub nodes) were identified.


Subject(s)
Chemokine CCL5/genetics , Chemokine CCL5/metabolism , Complement C3a/metabolism , Complement C5a/metabolism , Coronavirus Infections/diagnosis , Interleukin-8/metabolism , Lung/metabolism , Middle East Respiratory Syndrome Coronavirus/physiology , Aged , Biomarkers/metabolism , Complement C3a/genetics , Complement C5a/genetics , Coronavirus Infections/metabolism , Coronavirus Infections/mortality , Female , Humans , Interleukin-8/genetics , Male , Middle Aged , Prognosis , Severity of Illness Index , Survival Analysis , Up-Regulation
12.
Arch Virol ; 166(8): 2285-2289, 2021 Aug.
Article in English | MEDLINE | ID: covidwho-1245240

ABSTRACT

Mesenchymal stromal cells (MSCs) are considered multipotent progenitors with the capacity to differentiate into mesoderm-like cells in many species. The immunosuppressive properties of MSCs are important for downregulating inflammatory responses. Turkey coronavirus (TCoV) is the etiological agent of a poult mortality syndrome that affects intestinal epithelial cells. In this study, poult MSCs were isolated, characterized, and infected with TCoV after in vitro culture. The poult-derived MSCs showed fibroblast-like morphology and the ability to undergo differentiation into mesodermal-derived cells and to support virus replication. Infection with TCoV resulted in cytopathic effects and the loss of cell viability. TCoV antigens and new viral progeny were detected at high levels, as were transcripts of the pro-inflammatory factors INFγ, IL-6, and IL-8. These findings suggest that the cytokine storm phenomenon is not restricted to one genus of the family Coronaviridae and that MSCs cannot always balance the process.


Subject(s)
Coronavirus, Turkey/physiology , Cytokines/metabolism , Virus Replication , Animals , Cell Differentiation , Cell Survival , Cytopathogenic Effect, Viral , Interferon-gamma/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/virology , Turkeys , Up-Regulation
13.
Int J Mol Sci ; 22(11)2021 May 26.
Article in English | MEDLINE | ID: covidwho-1244042

ABSTRACT

Infection induces the production of proinflammatory cytokines and chemokines such as interleukin-8 (IL-8) and IL-6. Although they facilitate local antiviral immunity, their excessive release leads to life-threatening cytokine release syndrome, exemplified by the severe cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In this study, we investigated the roles of the integrated stress response (ISR) and activator protein-1 (AP-1) family proteins in regulating coronavirus-induced IL-8 and IL-6 upregulation. The mRNA expression of IL-8 and IL-6 was significantly induced in cells infected with infectious bronchitis virus (IBV), a gammacoronavirus, and porcine epidemic diarrhea virus, an alphacoronavirus. Overexpression of a constitutively active phosphomimetic mutant of eukaryotic translation initiation factor 2α (eIF2α), chemical inhibition of its dephosphorylation, or overexpression of its upstream double-stranded RNA-dependent protein kinase (PKR) significantly enhanced IL-8 mRNA expression in IBV-infected cells. Overexpression of the AP-1 protein cJUN or its upstream kinase also increased the IBV-induced IL-8 mRNA expression, which was synergistically enhanced by overexpression of cFOS. Taken together, this study demonstrated the important regulatory roles of ISR and AP-1 proteins in IL-8 production during coronavirus infection, highlighting the complex interactions between cellular stress pathways and the innate immune response.


Subject(s)
Coronavirus Infections/metabolism , Endoplasmic Reticulum Stress/genetics , Eukaryotic Initiation Factor-2/metabolism , Interleukin-8/metabolism , Unfolded Protein Response/genetics , Alphacoronavirus/metabolism , Alphacoronavirus/pathogenicity , Animals , Cell Line , Chlorocebus aethiops , Coronavirus Infections/genetics , Gammacoronavirus/metabolism , Gammacoronavirus/pathogenicity , Gene Expression Regulation , Humans , Immunity, Innate , Infectious bronchitis virus/metabolism , Infectious bronchitis virus/pathogenicity , Interleukin-8/genetics , Phosphorylation , Porcine epidemic diarrhea virus/metabolism , Porcine epidemic diarrhea virus/pathogenicity , Proto-Oncogene Proteins c-fos/genetics , Proto-Oncogene Proteins c-fos/metabolism , Proto-Oncogene Proteins c-jun/genetics , Proto-Oncogene Proteins c-jun/metabolism , Signal Transduction/genetics , Transcription Factor AP-1/genetics , Transcription Factor AP-1/metabolism , Up-Regulation , Vero Cells , eIF-2 Kinase/genetics , eIF-2 Kinase/metabolism
14.
J Enzyme Inhib Med Chem ; 36(1): 659-668, 2021 Dec.
Article in English | MEDLINE | ID: covidwho-1109085

ABSTRACT

Human intestinal epithelial cell line-6 (HIEC-6) cells and primary human hepatocytes (PHHs) were treated with 3-amidinophenylalanine-derived inhibitors of trypsin-like serine proteases for 24 hours. It was proven that treatment with MI-1900 and MI-1907 was tolerated up to 50 µM in HIEC-6. These inhibitors did not cause elevations in extracellular H2O2 levels and in the concentrations of interleukin (IL)-6 and IL-8 and did not alter occludin distribution in HIEC-6. It was also found that MI-1900 and MI-1907 up to 50 µM did not affect cell viability, IL-6 and IL-8 and occludin levels of PHH. Based on our findings, these inhibitors could be safely applicable at 50 µM in HIEC-6 and in PHH; however, redox status was disturbed in case of PHH. Moreover, it has recently been demonstrated that MI-1900 prevents the replication and spread of the new SARS-CoV-2 in infected Calu-3 cells, most-likely via an inhibition of the membrane-bound host protease TMPRSS2.


Subject(s)
Antiviral Agents/pharmacology , Epithelial Cells/drug effects , Hepatocytes/drug effects , Phenylalanine/pharmacology , Protease Inhibitors/pharmacology , Serine Endopeptidases/metabolism , Cell Line , Cell Survival/drug effects , Epithelial Cells/cytology , Epithelial Cells/enzymology , Gene Expression Regulation/drug effects , Hepatocytes/cytology , Hepatocytes/enzymology , Humans , Hydrogen Peroxide/metabolism , Interleukin-6/genetics , Interleukin-6/metabolism , Interleukin-8/genetics , Interleukin-8/metabolism , Intestinal Mucosa/cytology , Intestinal Mucosa/drug effects , Intestinal Mucosa/enzymology , Occludin/genetics , Occludin/metabolism , Oxidation-Reduction/drug effects , Phenylalanine/analogs & derivatives , Primary Cell Culture , Serine Endopeptidases/genetics
15.
J Am Coll Nutr ; 40(4): 327-332, 2021.
Article in English | MEDLINE | ID: covidwho-1087590

ABSTRACT

Objective: Vitamin D deficiency is common in the general population and diabetic patients, and supplementation with vitamin D is widely used to help lower oxidative stress and inflammation. The cytokine storm in SARS-CoV2 infection has been linked with both diabetes and Vitamin D deficiency. This study examined the hypothesis that supplementation with vitamin D, in combination with l-cysteine (LC), is better at reducing oxidative stress and thereby, more effective, at inhibiting the secretion of the pro-inflammatory cytokines, Interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1) in U937 monocytes exposed to high glucose concentrations. Methods: U937 monocytes were pretreated with 1,25 (OH)2 vitamin D (VD, 10 nM) or LC (250 µM) or VD + LC for 24 h and then exposed to control or high glucose (HG, 25 mM) for another 24 h. Results: There were significantly greater reactive oxygen species (ROS) levels in monocytes treated with HG than those in controls. Combined supplementation with VD and LC showed a more significant reduction in ROS (46%) in comparison with treatment with LC (19%) or VD (26%) alone in monocytes exposed to HG. Similarly, VD supplementation, together with LC, caused a more significant inhibition in the secretion of IL-8 (36% versus 16%) and MCP-1 (46% versus 26%) in comparison with that of VD (10 nM) alone in high-glucose treated monocytes. Conclusions: These results suggest that combined supplementation with vitamin D and LC has the potential to be more effective than either VD or LC alone in lowering the risk of oxidative stress and inflammation associated with type 2 diabetes or COVID-19 infection. Further, this combined vitamin D with LC/N-acetylcysteine may be a potent alternative therapy for SARS-CoV2 infected subjects. This approach can prevent cellular damage due to cytokine storm in comorbid systemic inflammatory conditions, such as diabetes, obesity, and hypertension.


Subject(s)
COVID-19/drug therapy , Cysteine/administration & dosage , Oxidative Stress/drug effects , SARS-CoV-2/immunology , Vitamin D/administration & dosage , COVID-19/immunology , Chemokine CCL2/metabolism , Diabetes Mellitus, Type 2/drug therapy , Dietary Supplements , Drug Therapy, Combination , Glucose/administration & dosage , Humans , Interleukin-8/metabolism , Monocytes/immunology , Monocytes/virology , U937 Cells , Vitamin D Deficiency/drug therapy , Vitamin D Deficiency/immunology , Vitamin D Deficiency/virology
16.
Int J Mol Sci ; 22(3)2021 Jan 29.
Article in English | MEDLINE | ID: covidwho-1055071

ABSTRACT

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is currently a global public health emergency. Periodontitis, the most prevalent disease that leads to tooth loss, is caused by infection by periodontopathic bacteria. Periodontitis is also a risk factor for pneumonia and the exacerbation of chronic obstructive pulmonary disease, presumably because of the aspiration of saliva contaminated with periodontopathic bacteria into the lower respiratory tract. Patients with these diseases have increased rates of COVID-19 aggravation and mortality. Because periodontopathic bacteria have been isolated from the bronchoalveolar lavage fluid of patients with COVID-19, periodontitis may be a risk factor for COVID-19 aggravation. However, the molecular links between periodontitis and COVID-19 have not been clarified. In this study, we found that the culture supernatant of the periodontopathic bacterium Fusobacterium nucleatum (CSF) upregulated the SARS-CoV-2 receptor angiotensin-converting enzyme 2 in A549 alveolar epithelial cells. In addition, CSF induced interleukin (IL)-6 and IL-8 production by both A549 and primary alveolar epithelial cells. CSF also strongly induced IL-6 and IL-8 expression by BEAS-2B bronchial epithelial cells and Detroit 562 pharyngeal epithelial cells. These results suggest that when patients with mild COVID-19 frequently aspirate periodontopathic bacteria, SARS-CoV-2 infection is promoted, and inflammation in the lower respiratory tract may become severe in the presence of viral pneumonia.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Culture Media, Conditioned/chemistry , Cytokines/metabolism , Fusobacterium nucleatum/metabolism , Angiotensin-Converting Enzyme 2/genetics , COVID-19/pathology , COVID-19/virology , Cell Line , Culture Media, Conditioned/pharmacology , Epithelial Cells/cytology , Epithelial Cells/metabolism , Epithelial Cells/microbiology , Humans , Interleukin-6/metabolism , Interleukin-8/metabolism , SARS-CoV-2/isolation & purification , Up-Regulation/drug effects
17.
Cell Death Dis ; 11(11): 957, 2020 11 06.
Article in English | MEDLINE | ID: covidwho-912894

ABSTRACT

A global effort is currently undertaken to restrain the COVID-19 pandemic. Host immunity has come out as a determinant for COVID-19 clinical outcomes, and several studies investigated the immune profiling of SARS-CoV-2 infected people to properly direct the clinical management of the disease. Thus, lymphopenia, T-cell exhaustion, and the increased levels of inflammatory mediators have been described in COVID-19 patients, in particular in severe cases1. Age represents a key factor in COVID-19 morbidity and mortality2. Understanding age-associated immune signatures of patients are therefore important to identify preventive and therapeutic strategies. In this study, we investigated the immune profile of COVID-19 hospitalized patients identifying a distinctive age-dependent immune signature associated with disease severity. Indeed, defined circulating factors - CXCL8, IL-10, IL-15, IL-27, and TNF-α - positively correlate with older age, longer hospitalization, and a more severe form of the disease and may thus represent the leading signature in critical COVID-19 patients.


Subject(s)
Coronavirus Infections/pathology , Cytokines/metabolism , Pneumonia, Viral/pathology , Age Factors , Aged , Aged, 80 and over , Antibodies, Viral/blood , Betacoronavirus/immunology , Betacoronavirus/isolation & purification , COVID-19 , Cluster Analysis , Coronavirus Infections/immunology , Coronavirus Infections/virology , Female , Humans , Immunoglobulin G/blood , Interleukin-10/metabolism , Interleukin-8/metabolism , Length of Stay , Leukocytes, Mononuclear/cytology , Leukocytes, Mononuclear/metabolism , Male , Middle Aged , Pandemics , Pneumonia, Viral/immunology , Pneumonia, Viral/virology , SARS-CoV-2 , Severity of Illness Index , Tumor Necrosis Factor-alpha/metabolism
18.
Front Immunol ; 11: 2145, 2020.
Article in English | MEDLINE | ID: covidwho-776206

ABSTRACT

SARS-CoV-2 infection has recently been declared a pandemic. Some patients showing severe symptoms exhibit drastic inflammation and airway damage. In this study, we re-analyzed published scRNA-seq data of COVID-19 patient bronchoalveolar lavage fluid to further classify and compare immunological features according to the patient's disease severity. Patients with severe symptoms showed DNA damage and apoptotic features of epithelial cells. Our results suggested that epithelial damage was associated with neutrophil infiltration. Myeloid cells of severe patients showed higher expression of proinflammatory cytokines and chemokines such as CXCL8. As a result, neutrophils were abundant in lungs of patients from the severe group. Furthermore, recruited neutrophils highly expressed genes related to neutrophil extracellular traps. Neutrophil-mediated inflammation was regulated by glucocorticoid receptor expression and activity. Based on these results, we suggest that severe COVID-19 symptoms may be determined by differential expression of glucocorticoid receptors and neutrophils.


Subject(s)
Betacoronavirus/immunology , Coronavirus Infections/immunology , Interleukin-8/genetics , Neutrophils/immunology , Pneumonia, Viral/immunology , Receptors, Glucocorticoid/genetics , Severity of Illness Index , Transcriptome , Adult , Aged , Bronchoalveolar Lavage Fluid/immunology , COVID-19 , Coronavirus Infections/virology , Epithelial Cells/pathology , Extracellular Traps/immunology , Female , Gene Expression Profiling/methods , Humans , Inflammation/immunology , Interleukin-8/metabolism , Male , Middle Aged , Myeloid Cells/immunology , Neutrophil Infiltration/immunology , Pandemics , Pneumonia, Viral/virology , RNA-Seq , Receptors, Glucocorticoid/metabolism , SARS-CoV-2 , Single-Cell Analysis/methods
19.
Int J Mol Sci ; 21(15)2020 Jul 31.
Article in English | MEDLINE | ID: covidwho-693525

ABSTRACT

The COVID-19 pandemic caused by the SARS-CoV-2 virus, overlaps with the ongoing epidemics of cigarette smoking and electronic cigarette (e-cig) vaping. However, there is scarce data relating COVID-19 risks and outcome with cigarette or e-cig use. In this study, we mined three independent RNA expression datasets from smokers and vapers to understand the potential relationship between vaping/smoking and the dysregulation of key genes and pathways related to COVID-19. We found that smoking, but not vaping, upregulates ACE2, the cellular receptor that SARS-CoV-2 requires for infection. Both smoking and use of nicotine and flavor-containing e-cigs led to upregulation of pro-inflammatory cytokines and inflammasome-related genes. Specifically, chemokines including CCL20 and CXCL8 are upregulated in smokers, and CCL5 and CCR1 are upregulated in flavor/nicotine-containing e-cig users. We also found genes implicated in inflammasomes, such as CXCL1, CXCL2, NOD2, and ASC, to be upregulated in smokers and these e-cig users. Vaping flavor and nicotine-less e-cigs, however, did not lead to significant cytokine dysregulation and inflammasome activation. Release of inflammasome products, such as IL-1B, and cytokine storms are hallmarks of COVID-19 infection, especially in severe cases. Therefore, our findings demonstrated that smoking or vaping may critically exacerbate COVID-19-related inflammation or increase susceptibility to COVID-19.


Subject(s)
Electronic Nicotine Delivery Systems , Immune System/metabolism , Peptidyl-Dipeptidase A/metabolism , Tobacco Smoking , Adult , Angiotensin-Converting Enzyme 2 , Betacoronavirus/isolation & purification , Bronchi/cytology , COVID-19 , Chemokine CCL20/genetics , Chemokine CCL20/metabolism , Coronavirus Infections/pathology , Coronavirus Infections/virology , Epithelial Cells/cytology , Epithelial Cells/metabolism , Humans , Interleukin-1beta/metabolism , Interleukin-8/genetics , Interleukin-8/metabolism , Middle Aged , Nod2 Signaling Adaptor Protein/genetics , Nod2 Signaling Adaptor Protein/metabolism , Pandemics , Pneumonia, Viral/pathology , Pneumonia, Viral/virology , SARS-CoV-2 , Up-Regulation , Young Adult
20.
JAMA Cardiol ; 5(11): 1281-1285, 2020 11 01.
Article in English | MEDLINE | ID: covidwho-676377

ABSTRACT

Importance: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be documented in various tissues, but the frequency of cardiac involvement as well as possible consequences are unknown. Objective: To evaluate the presence of SARS-CoV-2 in the myocardial tissue from autopsy cases and to document a possible cardiac response to that infection. Design, Setting, and Participants: This cohort study used data from consecutive autopsy cases from Germany between April 8 and April 18, 2020. All patients had tested positive for SARS-CoV-2 in pharyngeal swab tests. Exposures: Patients who died of coronavirus disease 2019. Main Outcomes and Measures: Incidence of SARS-CoV-2 positivity in cardiac tissue as well as CD3+, CD45+, and CD68+ cells in the myocardium and gene expression of tumor necrosis growth factor α, interferon γ, chemokine ligand 5, as well as interleukin-6, -8, and -18. Results: Cardiac tissue from 39 consecutive autopsy cases were included. The median (interquartile range) age of patients was 85 (78-89) years, and 23 (59.0%) were women. SARS-CoV-2 could be documented in 24 of 39 patients (61.5%). Viral load above 1000 copies per µg RNA could be documented in 16 of 39 patients (41.0%). A cytokine response panel consisting of 6 proinflammatory genes was increased in those 16 patients compared with 15 patients without any SARS-CoV-2 in the heart. Comparison of 15 patients without cardiac infection with 16 patients with more than 1000 copies revealed no inflammatory cell infiltrates or differences in leukocyte numbers per high power field. Conclusions and Relevance: In this analysis of autopsy cases, viral presence within the myocardium could be documented. While a response to this infection could be reported in cases with higher virus load vs no virus infection, this was not associated with an influx of inflammatory cells. Future investigations should focus on evaluating the long-term consequences of this cardiac involvement.


Subject(s)
Autopsy/methods , COVID-19/complications , Cardiovascular Infections/etiology , SARS-CoV-2/genetics , Aged , Aged, 80 and over , COVID-19/diagnosis , COVID-19/epidemiology , COVID-19/virology , Cardiovascular Infections/metabolism , Cardiovascular Infections/virology , Chemokines/metabolism , Cohort Studies , Female , Germany/epidemiology , Humans , Incidence , Interferon-gamma/metabolism , Interleukin-18/metabolism , Interleukin-6/metabolism , Interleukin-8/metabolism , Male , Myocarditis/etiology , Myocarditis/metabolism , Myocarditis/virology , Myocardium/immunology , Myocardium/metabolism , Pandemics , Peptide Fragments/metabolism , SARS-CoV-2/isolation & purification , Tumor Necrosis Factor-alpha/metabolism , Viral Load/statistics & numerical data
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