TNFR1 Absence Is Not Crucial for Different Types of Cell Reaction to TNF: A Study of the TNFR1-Knockout Cell Model.

BACKGROUND: One of the mechanisms regulating the biological activity of tumor necrosis factor (TNF) in cells is the co-expression of TNFR1/TNFR2 receptors. A model with a differential level of receptor expression is required to evaluate the contribution of these mechanisms. AIM: The development of a cellular model to compare the effects of TNF on cells depending on the presence of both receptors and TNFR2 alone. METHODS: TNFR1 absence modifications of ZR-75/1 and K-562 cell lines were obtained by TNFR1 knockout. The presence of deletions was confirmed by Sanger sequencing, and the absence of cell membrane receptor expression was confirmed by flow cytometry. The dose-dependent effect of TNF on intact and knockout cells was comparatively evaluated by the effect on the cell cycle, the type of cell death, and the profile of expressed genes. RESULTS: Knockout of TNFR1 resulted in a redistribution of TNFR2 receptors with an increased proportion of TNFR2+ cells in both lines and a multidirectional change in the density of expression in the lines (increased in K562 and decreased in ZR75/1). The presence of a large number of cells with high TNFR2 density in the absence of TNFR1 in the K562 cells was associated with greater sensitivity to TNF-stimulating doses and increased proliferation but did not result in a significant change in cell death parameters. A twofold increase in TNFR2+ cell distribution in this cell line at a reduced expression density in ZR75/1 cells was associated with a change in sensitivity to low cytokine concentrations in terms of proliferation; an overall increase in cell death, most pronounced at standard stimulating concentrations; and increased expression of the lymphocyte-activation gene groups, host-pathogen interaction, and innate immunity. CONCLUSIONS: The absence of TNFR1 leads to different variants of compensatory redistribution of TNFR2 in cellular models, which affects the type of cell response and the threshold level of sensitivity. The directionality of cytokine action modulation and sensitivity to TNF levels depends not only on the fraction of cells expressing TNFR2 but also on the density of expression.

Recombinant SARS-CoV-2 S Protein Binds to Glycans of the Lactosamine Family in vitro.

Many viruses, beside binding to their main cell target, interact with other molecules that promote virus adhesion to the cell; often, these additional targets are glycans. The main receptor for SARS-CoV-2 is a peptide motif in the ACE2 protein. We studied interaction of the recombinant SARS-CoV-2 spike (S) protein with an array of glycoconjugates, including various sialylated, sulfated, and other glycans, and found that the S protein binds some (but not all) glycans of the lactosamine family. We suggest that parallel influenza infection will promote SARS-CoV-2 adhesion to the respiratory epithelial cells due to the unmasking of lactosamine chains by the influenza virus neuraminidase.

Amphiphilic Poly(N-Vinylpyrrolidone) Nanoparticles Loaded with DNA Plasmids Encoding Gn and Gc Glycoproteins of the Rift Valley Fever Virus: Preparation and In Vivo Evaluation.

The aim of the study was to develop amphiphilic poly(N-vinylpyrrolidone) (PVP) nanoparticles (NPs) loaded with DNA plasmids encoding Gn and Gc glycoproteins of the Rift Valley fever virus (RVFV) and to study the humoral response in vivo. DNA plasmids were protected from extracellular nucleases by loading in NPs from PVP derivatives modified with amino acids ß-alanine (Ala7-PVPOD4000) or glycine (Gly7.5-PVP-OD4000) fabricated by the original self-assembly technique. The obtained NPs were administered in mice and the enhancement of humoral response compared to this one in case of immunization with native DNA plasmids was demonstrated. The NPs loaded with DNA plasmids are promising for the fabrication of various DNA particulate vaccines.

Microencapsulated plasmids expressing Gn and Gc glycoproteins of Rift Valley Fever virus enhance humoral immune response in mice.

OBJECTIVES: The aim of the current study was to develop biodegradable alginate (ALG)/poly-L-lysine (PLL) microcapsules (MC) with entrapped plasmids expressing Gn and Gc glycoproteins of Rift Valley Fever virus (RVFV) and to evaluate the humoral immune response in mice. RESULTS: Expressing phRVF/Gn and phRVF/Gc plasmids which encode full-sized Gn and Gc glycoproteins and contain signal fusion protein F sequences of human parainfluenza (HPIV-1) were constructed. To protect the plasmids from cleavage by extracellular nucleases, they were entrapped into multilayer ALG/PLL microcapsules by layer-by-layer technique. To study the efficacy of humoral immune response, both native and microencapsulated plasmids were injected intramuscular into BALB/c mice. The humoral response in the mice immunized with free plasmids was characterized by virus-neutralizing antibody induction (with titres 1:4 to 1:8), while the injection of microencapsulated plasmids allowed to increase the titre level (from 1:16 to 1:32). CONCLUSION: The plasmids microencapsulated in biodegradable MC could be promising for development of DNA vaccines against RVFV.