ABSTRACT
The SARS-CoV-2 pandemic has become a severe global public health event, and the development of protective and therapeutic strategies is urgently needed. Downregulation of angiotensin converting enzyme 2 (ACE2; one of the important SARS-CoV-2 entry receptors) and aberrant inflammatory responses (cytokine storm) are the main targets to inhibit and control COVID-19 invasion. Silver nanomaterials have well-known pharmaceutical properties, including antiviral, antibacterial, and anticancer properties. Here, based on a self-established metal evaporation-condensation-size graded collection system, smaller silver particles reaching the Ångstrom scale (AgÅPs) were fabricated and coated with fructose to obtain a stabilized AgÅP solution (F-AgÅPs). F-AgÅPs potently inactivated SARS-CoV-2 and prevented viral infection. Considering the application of anti-SARS-CoV-2, a sterilized F-AgÅP solution was produced via spray formulation. In our model, the F-AgÅP spray downregulated ACE2 expression and attenuated proinflammatory factors. Moreover, F-AgÅPs were found to be rapidly eliminated to avoid respiratory and systemic toxicity in this study as well as our previous studies. This work presents a safe and potent anti-SARS-CoV-2 agent using an F-AgÅP spray.
Subject(s)
Angiotensin-Converting Enzyme 2 , COVID-19 Drug Treatment , Humans , Peptidyl-Dipeptidase A/metabolism , SARS-CoV-2 , Silver/pharmacologyABSTRACT
Coronavirus disease 2019 (COVID19), caused by the severe acute respiratory syndrome coronavirus2 (SARSCoV2), led to an outbreak of viral pneumonia in December 2019. The present study aimed to investigate the host inflammatory response signaturecaused by SARSCoV2 in human corneal epithelial cells (HCECs). The expression level of angiotensinconverting enzyme 2 (ACE2) in the human cornea was determined via immunofluorescence. In vitro experiments were performed in HCECs stimulated with the SARSCoV2 spike protein. Moreover, the expression levels of ACE2, IL8, TNFα, IL6, gasdermin D (GSDMD) and IL1ß in HCECs were detected using reverse transcriptionquantitative PCR and/or western blotting. It was identified that ACE2 was expressed in normal human corneal epithelium and HCECs cultured in vitro. Furthermore, the expression levels of IL8, TNFα and IL6 in HCECs were decreased following SARSCoV2 spike protein stimulation, while the expression levels of GSDMD and IL1ß were increased. In conclusion, the present results demonstrated that the SARSCoV2 spike protein suppressed the host inflammatory response and induced pyroptosis in HCECs. Therefore, blocking the ACE2 receptor in HCECs may reduce the infection rate of COVID19.
Subject(s)
Epithelium, Corneal/metabolism , Spike Glycoprotein, Coronavirus/metabolism , Adult , Aged , Angiotensin-Converting Enzyme 2/genetics , Angiotensin-Converting Enzyme 2/metabolism , Cells, Cultured , Cornea/cytology , Epithelial Cells/cytology , Epithelial Cells/metabolism , Epithelial Cells/virology , Epithelium, Corneal/virology , Female , Humans , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Interleukin-6/genetics , Interleukin-6/metabolism , Intracellular Signaling Peptides and Proteins/genetics , Intracellular Signaling Peptides and Proteins/metabolism , Male , Middle Aged , Phosphate-Binding Proteins/genetics , Phosphate-Binding Proteins/metabolism , Pyroptosis , Spike Glycoprotein, Coronavirus/genetics , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolism , Up-RegulationABSTRACT
Infectious bovine viral diarrhea virus (BVDV) cDNA clones have been used for the expression of classical swine fever virus (CSFV) genes for immune prevention and control. However, can it be used for the expression of an allogenetic fragment? To answer this question, a BVDV chimeric virus expressing the spike (S) antigen fragment of porcine epidemic diarrhea virus (PEDV) was constructed. Antigen S499-602 was inserted into pig-derived BVDV-2 infectious cDNA clone pASH28 in tandem by overlapping PCR, located between the seventh and eighth amino acids at the N-terminus of the capsid (C) protein of BVDV. Indirect immunofluorescence assay confirmed that the chimeric virus vASH-dS312 containing double S499-602 sequences was successfully assembled, which could react with the monoclonal antibody (MAb) against BVDV E2 and PEDV S proteins. Further western blot analysis confirmed that the exogenous S499-602 double protein could be stably expressed. Next, the chimeric virus vASH-dS312 was administered to BALB/C mice either orally or by intramuscular injection. The immunized mice were healthy and showed no signs of toxicity. IgG against BVDV and PEDV antibodies could be detected in the mice administered vASH-dS312 by intramuscular injection, which had neutralization activity against BVDV and PEDV. Thus, this study reported a new insertion site in the BVDV infectious cDNA clone that could successfully express an allogenetic antigen.