scholarly journals In-House Immunofluorescence Assay for Detection of SARS-CoV-2 Antigens in Cells from Nasopharyngeal Swabs as a Diagnostic Method for COVID-19

Diagnostics ◽  
2021 ◽  
Vol 11 (12) ◽  
pp. 2346
Author(s):  
Athene Hoi-Ying Lam ◽  
Jian-Piao Cai ◽  
Ka-Yi Leung ◽  
Ricky-Ruiqi Zhang ◽  
Danlei Liu ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Nucleocapsid Protein ◽  
Diagnostic Method ◽  
Polyclonal Antibodies ◽  
Respiratory Viruses ◽  
Cross Reactivity ◽  
Immunofluorescence Assay ◽  
Respiratory Epithelial Cells ◽  
N Protein ◽  
Respiratory Epithelial

Immunofluorescence is a traditional diagnostic method for respiratory viruses, allowing rapid, simple and accurate diagnosis, with specific benefits of direct visualization of antigens-of-interest and quality assessment. This study aims to evaluate the potential of indirect immunofluorescence as an in-house diagnostic method for SARS-CoV-2 antigens from nasopharyngeal swabs (NPS). Three primary antibodies raised from mice were used for immunofluorescence staining, including monoclonal antibody against SARS-CoV nucleocapsid protein, and polyclonal antibodies against SARS-CoV-2 nucleocapsid protein and receptor-binding domain of SARS-CoV-2 spike protein. Smears of cells from NPS of 29 COVID-19 patients and 20 non-infected individuals, and cells from viral culture were stained by the three antibodies. Immunofluorescence microscopy was used to identify respiratory epithelial cells with positive signals. Polyclonal antibody against SARS-CoV-2 N protein had the highest sensitivity and specificity among the three antibodies tested, detecting 17 out of 29 RT-PCR-confirmed COVID-19 cases and demonstrating no cross-reactivity with other tested viruses except SARS-CoV. Detection of virus-infected cells targeting SARS-CoV-2 N protein allow identification of infected individuals, although accuracy is limited by sample quality and number of respiratory epithelial cells. The potential of immunofluorescence as a simple diagnostic method was demonstrated, which could be applied by incorporating antibodies targeting SARS-CoV-2 into multiplex immunofluorescence panels used clinically, such as for respiratory viruses, thus allowing additional routine testing for diagnosis and surveillance of SARS-CoV-2 even after the epidemic has ended with low prevalence of COVID-19.

scholarly journals Respiratory Epithelial Cells in Innate Immunity against Respiratory Viruses

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
David Rodriguez-Mier ◽  
Ernesto Torres-Lopez ◽  
Mario C. Salinas-Carmona ◽  
Adrián G. Rosas-Taraco
Keyword(s):  
Innate Immunity ◽  
Epithelial Cells ◽  
Respiratory Viruses ◽  
Respiratory Epithelial Cells ◽  
Respiratory Epithelial

scholarly journals SARS-CoV-2 drives JAK1/2-dependent local complement hyperactivation

2021 ◽  
Vol 6 (58) ◽  
pp. eabg0833
Author(s):  
Bingyu Yan ◽  
Tilo Freiwald ◽  
Daniel Chauss ◽  
Luopin Wang ◽  
Erin West ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Clinical Manifestations ◽  
Lung Epithelial Cells ◽  
Complement Factor ◽  
Respiratory Epithelial Cells ◽  
Signature Genes ◽  
Wide Range ◽  
Lung Epithelial ◽  
Gene Transcripts ◽  
Respiratory Epithelial

Patients with coronavirus disease 2019 (COVID-19) present a wide range of acute clinical manifestations affecting the lungs, liver, kidneys and gut. Angiotensin converting enzyme (ACE) 2, the best-characterized entry receptor for the disease-causing virus SARS-CoV-2, is highly expressed in the aforementioned tissues. However, the pathways that underlie the disease are still poorly understood. Here, we unexpectedly found that the complement system was one of the intracellular pathways most highly induced by SARS-CoV-2 infection in lung epithelial cells. Infection of respiratory epithelial cells with SARS-CoV-2 generated activated complement component C3a and could be blocked by a cell-permeable inhibitor of complement factor B (CFBi), indicating the presence of an inducible cell-intrinsic C3 convertase in respiratory epithelial cells. Within cells of the bronchoalveolar lavage of patients, distinct signatures of complement activation in myeloid, lymphoid and epithelial cells tracked with disease severity. Genes induced by SARS-CoV-2 and the drugs that could normalize these genes both implicated the interferon-JAK1/2-STAT1 signaling system and NF-κB as the main drivers of their expression. Ruxolitinib, a JAK1/2 inhibitor, normalized interferon signature genes and all complement gene transcripts induced by SARS-CoV-2 in lung epithelial cell lines, but did not affect NF-κB-regulated genes. Ruxolitinib, alone or in combination with the antiviral remdesivir, inhibited C3a protein produced by infected cells. Together, we postulate that combination therapy with JAK inhibitors and drugs that normalize NF-κB-signaling could potentially have clinical application for severe COVID-19.

scholarly journals Lycopene Inhibits Toll-Like Receptor 4-Mediated Expression of Inflammatory Cytokines in House Dust Mite-Stimulated Respiratory Epithelial Cells

Molecules ◽  
2021 ◽  
Vol 26 (11) ◽  
pp. 3127
Author(s):  
Jiyeon Choi ◽  
Joo Weon Lim ◽  
Hyeyoung Kim
Keyword(s):  
Epithelial Cells ◽  
A549 Cells ◽  
Cytokine Expression ◽  
Ros Production ◽  
Toll Like Receptor ◽  
Respiratory Epithelial Cells ◽  
Toll Like Receptor 4 ◽  
Mitochondrial Ros ◽  
Tlr4 Activation ◽  
Respiratory Epithelial

House dust mites (HDM) are critical factors in airway inflammation. They activate respiratory epithelial cells to produce reactive oxygen species (ROS) and activate Toll-like receptor 4 (TLR4). ROS induce the expression of inflammatory cytokines in respiratory epithelial cells. Lycopene is a potent antioxidant nutrient with anti-inflammatory activity. The present study aimed to investigate whether HDM induce intracellular and mitochondrial ROS production, TLR4 activation, and pro-inflammatory cytokine expression (IL-6 and IL-8) in respiratory epithelial A549 cells. Additionally, we examined whether lycopene inhibits HDM-induced alterations in A549 cells. The treatment of A549 cells with HDM activated TLR4, induced the expression of IL-6 and IL-8, and increased intracellular and mitochondrial ROS levels. TAK242, a TLR4 inhibitor, suppressed both HDM-induced ROS production and cytokine expression. Furthermore, lycopene inhibited the HDM-induced TLR4 activation and cytokine expression, along with reducing the intracellular and mitochondrial ROS levels in HDM-treated cells. These results collectively indicated that the HDM induced TLR4 activation and increased intracellular and mitochondrial ROS levels, thus resulting in the induction of cytokine expression in respiratory epithelial cells. The antioxidant lycopene could inhibit HDM-induced cytokine expression, possibly by suppressing TLR4 activation and reducing the intracellular and mitochondrial ROS levels in respiratory epithelial cells.

scholarly journals Dolosigranulum pigrum Modulates Immunity against SARS-CoV-2 in Respiratory Epithelial Cells

Pathogens ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 634
Author(s):  
Md. Aminul Islam ◽  
Leonardo Albarracin ◽  
Vyacheslav Melnikov ◽  
Bruno G. N. Andrade ◽  
Rafael R. C. Cuadrat ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Specific Effect ◽  
Commensal Bacteria ◽  
Cellular Damage ◽  
Poly I:C ◽  
Respiratory Epithelial Cells ◽  
Promising Alternative ◽  
Immunological Mechanisms ◽  
Syncytial Virus ◽  
Respiratory Epithelial

In a previous work, we demonstrated that nasally administered Dolosigranulum pigrum 040417 beneficially modulated the respiratory innate immune response triggered by the activation of Toll-like receptor 3 (TLR3) and improved protection against Respiratory Syncytial Virus (RSV) in mice. In this work, we aimed to evaluate the immunomodulatory effects of D. pigrum 040417 in human respiratory epithelial cells and the potential ability of this immunobiotic bacterium to increase the protection against Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The respiratory commensal bacterium D. pigrum 040417 differentially modulated the production of IFN-β, IL-6, CXCL8, CCL5 and CXCL10 in the culture supernatants of Calu-3 cells stimulated with poly(I:C) or challenged with SARS-CoV-2. The differential cytokine profile induced by the 040417 strain was associated with a significant reduction in viral replication and cellular damage after coronavirus infection. Of note, D. pigrum 030918 was not able to modify the resistance of Calu-3 cells to SARS-CoV-2 infection, indicating a strain-specific immunomodulatory effect for respiratory commensal bacteria. The findings of this work improve our understanding of the immunological mechanisms involved in the modulation of respiratory immunity induced by respiratory commensal bacteria, by demonstrating their specific effect on respiratory epithelial cells. In addition, the results suggest that particular strains such as D. pigrum 040417 could be used as a promising alternative for combating SARS-CoV-2 and reducing the severity of COVID-19.

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