ABSTRACT
Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.Copyright © 2022 Editorial Department of Journal of Shanghai Second Medical University. All rights reserved.
ABSTRACT
Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.
ABSTRACT
Objective: The Chinese herbal formula Huo-Xiang-Zheng-Qi (HXZQ) is effective in preventing and treating coronavirus disease 19 (COVID-19) infection;however, its mechanism remains unclear. This study used network pharmacology and molecular docking techniques to investigate the mechanism of action of HXZQ in preventing and treating COVID-19. Methods: The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) was used to search for the active ingredients and targets of the 10 traditional Chinese medicines (TCMs) of HXZQ prescription (HXZQP). GeneCards, Online Mendelian Inheritance in Man (OMIM), Pharmacogenomics Knowledge Base (PharmGKB), Therapeutic Target Database (TTD), and DrugBank databases were used to screen COVID-19-related genes and intersect them with the targets of HXZQP to obtain the drug efficacy targets. Cytoscape 3.8 software was used to construct the drug-active ingredient–target interaction network of HXZQP and perform protein–protein interaction (PPI) network construction and topology analysis. R software was used to perform Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Finally, AutoDock Vina was utilized for molecular docking of the active ingredients of TCM and drug target proteins. Results: A total of 151 active ingredients and 250 HXZQP targets were identified. Among these, 136 active ingredients and 67 targets of HXZQP were found to be involved in the prevention and treatment of COVID-19. The core proteins identified in the PPI network were MAPK1, MAPK3, MAPK8, MAPK14, STAT3, and PTGS2. Using GO and KEGG pathway enrichment analysis, HXZQP was found to primarily participate in biological processes such as defense response to a virus, cellular response to biotic stimulus, response to lipopolysaccharide, PI3K-Akt signaling pathway, Th17 cell differentiation, HIF-1 signaling pathway, and other signaling pathways closely related to COVID-19. Molecular docking results reflected that the active ingredients of HXZQP have a reliable affinity toward EGFR, MAPK1, MAPK3, MAPK8, and STAT3 proteins. Conclusion: Our study elucidated the main targets and pathways of HXZQP in the prevention and treatment of COVID-19. The study findings provide a basis for further investigation of the pharmacological effects of HXZQP.
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BACKGROUND & AIMS: Chronic hepatitis B is an incurable disease. Addressing the unmet medical need for therapies has been hampered by a lack of suitable cell culture models to investigate the HBV life cycle in a single experimental setup. We sought to develop a platform suitable to investigate all aspects of the entire HBV life cycle. METHODS: HepG2-NTCPsec+ cells were inoculated with HBV. Supernatants of infected cells were transferred to naïve cells. Inhibition of infection was determined in primary and secondary infected cells by high-content imaging of viral and cellular factors. Novel antivirals were triaged in cells infected with cell culture- or patient-derived HBV and in stably virus replicating cells. HBV internalisation and target-based receptor binding assays were conducted. RESULTS: We developed an HBV platform, screened 2,102 drugs and bioactives, and identified 3 early and 38 late novel HBV life cycle inhibitors using infectious HBV genotype D. Two early inhibitors, pranlukast (EC50 4.3 µM; 50% cytotoxic concentration [CC50] >50 µM) and cytochalasin D (EC50 0.07 µM; CC50 >50 µM), and 2 late inhibitors, fludarabine (EC50 0.1 µM; CC50 13.4 µM) and dexmedetomidine (EC50 6.2 µM; CC50 >50 µM), were further investigated. Pranlukast inhibited HBV preS1 binding, whereas cytochalasin D prevented the internalisation of HBV. Fludarabine inhibited the secretion of HBV progeny DNA, whereas dexmedetomidine interfered with the infectivity of HBV progeny. Patient-derived HBV genotype C was efficiently inhibited by fludarabine (EC50 0.08 µM) and dexmedetomidine (EC50 8.7 µM). CONCLUSIONS: The newly developed high-content assay is suitable to screen large-scale drug libraries, enables monitoring of the entire HBV life cycle, and discriminates between inhibition of early and late viral life cycle events. LAY SUMMARY: HBV infection is an incurable, chronic disease with few available treatments. Addressing this unmet medical need has been hampered by a lack of suitable cell culture models to study the entire viral life cycle in a single experimental setup. We developed an image-based approach suitable to screen large numbers of drugs, using a cell line that can be infected by HBV and produces large amounts of virus particles. By transferring viral supernatants from these infected cells to uninfected target cells, we could monitor the entire viral life cycle. We used this system to screen drug libraries and identified novel anti-HBV inhibitors that potently inhibit HBV in various phases of its life cycle. This assay will be an important new tool to study the HBV life cycle and accelerate the development of novel therapeutic strategies.
ABSTRACT
Chronic HCV infection and associated liver cancer impose a heavy burden on the healthcare system. Direct acting antivirals eliminate HCV, unless it is drug resistant, and partially reverse liver disease, but they cannot cure HCV-related cancer. A possible remedy could be a multi-component immunotherapeutic vaccine targeting both HCV-infected and malignant cells, but also those not infected with HCV. To meet this need we developed a two-component DNA vaccine based on the highly conserved core protein of HCV to target HCV-infected cells, and a renowned tumor-associated antigen telomerase reverse transcriptase (TERT) based on the rat TERT, to target malignant cells. Their synthetic genes were expression-optimized, and HCV core was truncated after aa 152 (Core152opt) to delete the domain interfering with immunogenicity. Core152opt and TERT DNA were highly immunogenic in BALB/c mice, inducing IFN-γ/IL-2/TNF-α response of CD4+ and CD8+ T cells. Additionally, DNA-immunization with TERT enhanced cellular immune response against luciferase encoded by a co-delivered plasmid (Luc DNA). However, DNA-immunization with Core152opt and TERT mix resulted in abrogation of immune response against both components. A loss of bioluminescence signal after co-delivery of TERT and Luc DNA into mice indicated that TERT affects the in vivo expression of luciferase directed by the immediate early cytomegalovirus and interferon-ß promoters. Panel of mutant TERT variants was created and tested for their expression effects. TERT with deleted N-terminal nucleoli localization signal and mutations abrogating telomerase activity still suppressed the IFN-ß driven Luc expression, while the inactivated reverse transcriptase domain of TERT and its analogue, enzymatically active HIV-1 reverse transcriptase, exerted only weak suppressive effects, implying that suppression relied on the presence of the full-length/nearly full-length TERT, but not its enzymatic activity. The effect(s) could be due to interference of the ectopically expressed xenogeneic rat TERT with biogenesis of mRNA, ribosomes and protein translation in murine cells, affecting the expression of immunogens. HCV core can aggravate this effect, leading to early apoptosis of co-expressing cells, preventing the induction of immune response.