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1.
Sci Rep ; 11(1): 20595, 2021 10 18.
Article in English | MEDLINE | ID: covidwho-1475487

ABSTRACT

The delivery of safe, visible wavelengths of light can be an effective, pathogen-agnostic, countermeasure that would expand the current portfolio of SARS-CoV-2 intervention strategies beyond the conventional approaches of vaccine, antibody, and antiviral therapeutics. Employing custom biological light units, that incorporate optically engineered light-emitting diode (LED) arrays, we harnessed monochromatic wavelengths of light for uniform delivery across biological surfaces. We demonstrated that primary 3D human tracheal/bronchial-derived epithelial tissues tolerated high doses of a narrow spectral band of visible light centered at a peak wavelength of 425 nm. We extended these studies to Vero E6 cells to understand how light may influence the viability of a mammalian cell line conventionally used for assaying SARS-CoV-2. The exposure of single-cell monolayers of Vero E6 cells to similar doses of 425 nm blue light resulted in viabilities that were dependent on dose and cell density. Doses of 425 nm blue light that are well-tolerated by Vero E6 cells also inhibited infection and replication of cell-associated SARS-CoV-2 by > 99% 24 h post-infection after a single five-minute light exposure. Moreover, the 425 nm blue light inactivated cell-free betacoronaviruses including SARS-CoV-1, MERS-CoV, and SARS-CoV-2 up to 99.99% in a dose-dependent manner. Importantly, clinically applicable doses of 425 nm blue light dramatically inhibited SARS-CoV-2 infection and replication in primary human 3D tracheal/bronchial tissue. Safe doses of visible light should be considered part of the strategic portfolio for the development of SARS-CoV-2 therapeutic countermeasures to mitigate coronavirus disease 2019 (COVID-19).


Subject(s)
COVID-19/drug therapy , COVID-19/prevention & control , Light , SARS-CoV-2 , Trachea/radiation effects , Virus Replication/radiation effects , Adult , Animals , Antiviral Agents/pharmacology , Bronchi , Calibration , Cell-Free System , Chlorocebus aethiops , Epithelium/pathology , Female , Humans , Respiratory Mucosa/radiation effects , Trachea/virology , Vero Cells
2.
PLoS Biol ; 19(3): e3001143, 2021 03.
Article in English | MEDLINE | ID: covidwho-1138557

ABSTRACT

There are currently limited Food and Drug Administration (FDA)-approved drugs and vaccines for the treatment or prevention of Coronavirus Disease 2019 (COVID-19). Enhanced understanding of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection and pathogenesis is critical for the development of therapeutics. To provide insight into viral replication, cell tropism, and host-viral interactions of SARS-CoV-2, we performed single-cell (sc) RNA sequencing (RNA-seq) of experimentally infected human bronchial epithelial cells (HBECs) in air-liquid interface (ALI) cultures over a time course. This revealed novel polyadenylated viral transcripts and highlighted ciliated cells as a major target at the onset of infection, which we confirmed by electron and immunofluorescence microscopy. Over the course of infection, the cell tropism of SARS-CoV-2 expands to other epithelial cell types including basal and club cells. Infection induces cell-intrinsic expression of type I and type III interferons (IFNs) and interleukin (IL)-6 but not IL-1. This results in expression of interferon-stimulated genes (ISGs) in both infected and bystander cells. This provides a detailed characterization of genes, cell types, and cell state changes associated with SARS-CoV-2 infection in the human airway.


Subject(s)
Bronchi/pathology , COVID-19/diagnosis , Gene Expression , SARS-CoV-2/isolation & purification , Single-Cell Analysis/methods , Adult , Bronchi/virology , COVID-19/immunology , COVID-19/pathology , COVID-19/virology , Cells, Cultured , Epithelium/pathology , Epithelium/virology , Humans , Immunity, Innate , Longitudinal Studies , SARS-CoV-2/genetics , Transcriptome , Viral Tropism
3.
Laryngoscope ; 131(6): E2013-E2017, 2021 06.
Article in English | MEDLINE | ID: covidwho-969763

ABSTRACT

OBJECTIVES/HYPOTHESIS: Intracellular entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) depends on the interaction between its spike protein with the cellular receptor angiotensin-converting enzyme 2 (ACE2) and depends on Furin-mediated spike protein cleavage and spike protein priming by host cell proteases, including transmembrane protease serine 2 (TMPRSS2). As the expression of ACE2, TMPRSS2, and Furin in the middle and inner ear remain unclear, we analyzed the expression of these proteins in mouse ear tissues. STUDY DESIGN: Animal Research. METHODS: We performed immunohistochemical analysis to examine the distribution of ACE2, TMPRSS2, and Furin in the Eustachian tube, middle ear spaces, and cochlea of mice. RESULTS: ACE2 was present in the nucleus of the epithelium of the middle ear and Eustachian tube, as well as in some nuclei of the hair cells in the organ of Corti, in the stria vascularis, and the spiral ganglion cells. ACE2 was also expressed in the cytoplasm of the stria vascularis. TMPRSS2 was expressed in both the nucleus and cytoplasm in the middle spaces, with the expression being stronger in the nucleus in the mucosal epithelium of the middle ear spaces and Eustachian tube. TMPRSS2 was present in the cytoplasm in the organ of Corti and stria vascularis and in the nucleus and cytoplasm in the spiral ganglion. Furin was expressed in the cytoplasm in the middle ear spaces, Eustachian tube, and cochlea. CONCLUSIONS: ACE2, TMPRSS2, and Furin are diffusely present in the Eustachian tube, middle ear spaces, and cochlea, suggesting that these tissues are susceptible to SARS-CoV-2 infection. LEVEL OF EVIDENCE: NA Laryngoscope, 131:E2013-E2017, 2021.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , COVID-19/pathology , Ear, Inner/pathology , Ear, Middle/pathology , Eustachian Tube/pathology , Furin/genetics , Gene Expression/genetics , Serine Endopeptidases/genetics , Animals , Cochlea/pathology , Epithelium/pathology , Immunohistochemistry , Mice , Mucous Membrane/pathology , Organ of Corti/pathology , Spiral Ganglion/pathology , Stria Vascularis/pathology , Temporal Bone/pathology
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