Background: Although mRNA vaccines encapsulated in lipid nanoparticles (LNPs) have demonstrated a safety profile with minimal serious adverse events in clinical trials, there is opportunity to further reduce mRNA reactogenicity. The development of naked mRNA vaccines could improve vaccine tolerability. Naked nucleic acid delivery using the jet injection method may be a solution. Methods: In the first part of the study, the optimal conditions providing low traumatization and high expression of the model mRNA-GFP molecule in the tissues of laboratory animals were determined. Then, we used the selected protocol to immunize BALB/c mice with mRNA-RBD encoding the SARS-CoV-2 receptor-binding domain (RBD). It was demonstrated that mice vaccinated with naked mRNA-RBD developed a high level of specific antibodies with virus-neutralizing activity. The vaccine also induced a strong RBD-specific T-cell response and reduced the viral load in the lungs of the animals after infection with the SARS-CoV-2 virus. The level of immune response in mice immunized with mRNA-RBD using a spring-loaded jet injector was comparable to that in animals immunized with mRNA-RBD encapsulated in LNPs. Results: In this study, the efficacy of an inexpensive, simple, and safe method of mRNA delivery using a spring-loaded jet injector was evaluated and validated. Conclusions: Our findings suggest that the jet injection method may be a possible alternative to LNPs for delivering mRNA vaccines against SARS-CoV-2 infection.

BackgroundAt present, there are no universal markers of tumor stem cells known, including for B-lymphomas. Previously, we have shown that Epstein-Barr virus-induced B-cell lymphoma culture contains cells capable of internalizing TAMRA-labeled DNA. These cells form sphere-forming centers and are essential for the development of xenografts genetically identical to the initial culture.ObjectiveTo analyze the stem characteristics of cells that internalize DNA.MethodsSorting and RNA sequencing of two subpopulations (TAMRA + and TAMRA-) of Epstein-Barr virus-induced B-cell lymphoma culture and a series of quantitative real-time reverse transcription PCR were performed.ResultsTAMRA + cells were shown to have increased synthesis of mRNA of genes associated with the maintenance of a poorly differentiated state (SOX2, NANOG, POU5F1, CYP26A1), self-renewal (FZD5, FZD7, TCF3, LEF1) and epithelial-mesenchymal transition (MMP2, ITGB7). Transcriptomic analysis revealed that in TAMRA + cells, the synthesis of mitochondrial genes, as well as caspases and some apoptosis inhibitors, is reduced. TAMRA + cells possess clonogenic properties, increased level of synthesis of mRNA for key genes associated with self-renewal and poorly differentiated state maintenance.ConclusionsInternalization of the TAMRA-DNA probe is the marker of B-lymphoma cancer stem cells and can be used to detect tumor stem cells and develop new approaches to targeted treatment of B-lymphoma.

BACKGROUND: The emergence of the coronavirus (SARS-CoV-2) caused a pandemic that led to an increase in morbidity and mortality among infected individuals. The SARS-CoV-2 variants circulating during that period, including Delta, were characterized by a broad spectrum of pathological effects at both the tissue and cellular levels. In 2025, the Omicron variant predominates, producing milder forms of respiratory conditions. Among the key research questions related to SARS-CoV-2, considerable attention has been devoted to its pathogenicity, which is characterized by high virulence and the ability to induce systemic pathological processes. Transmission electron microscopy is used to visualize virus–cell receptor interactions and to study the destructive impact of the pathogen on host cells. The findings of such studies form the basis for understanding the course and pathogenesis of the disease and contribute to vaccine development. AIM: The work aimed to investigate SARS-CoV-2 virion reproduction and the trends of cytopathic effects in Vero E6 cell culture infected with Delta and Omicron variants. METHODS: Ultrastructural analysis was performed on Vero E6 cell cultures infected with the Delta and Omicron SARS-CoV-2 variants. Infected cell cultures were fixed at three time points: 6, 18, and 24 hours after viral inoculation. RESULTS: Most samples in both groups (Delta and Omicron) retained near-normal morphology 6 hours post-inoculation and exhibited numerous transport vesicles. At 18 hours, Delta and Omicron groups showed similar morphological changes, including vacuolization of the endoplasmic reticulum and Golgi complex. The Omicron group demonstrated altered mitochondrial morphology. Marked destructive changes were observed 24 hours post-inoculation. Both groups displayed cytoplasmic condensation, extensive organelle vacuolization, and numerous vesicles containing viral particles. Analysis of virion reproduction trends revealed an increase in viral particle numbers at 18 hours, followed by a decline in viral virion replication by 24 hours. CONCLUSION: The study suggests similar mechanisms of cytopathic pattern formation in Vero E6 cells infected with the Delta and Omicron SARS-CoV-2 variants, along with active virion reproduction in cell culture between 6 and 18 hours post-infection.

Despite the end of the COVID-19 pandemic, there still remain risks of new aggressive strains of coronavirus. As the human population increases progressively, it is mandatory to ensure both preventive measures and an immediate response to emerging infectious threats. Another essential component for rapidly restraining a new possible pandemic is the development of new anticoronaviral therapeutics. In the present study, the anticoronaviral capabilities of Gc protein-derived macrophage-activating factor (GcMAF) are characterized. It is demonstrated that the administration of GcMAF to Syrian hamsters infected with SARS-CoV-2 within the first phase of infection (six days postinfection) is accompanied by (i) a statistically significant reduction in the viral load of the lung tissue and (ii) the switching of the inflammatory status of the lung tissue to a neutral one in terms of mRNA expression levels of the groups of pro/anti-inflammatory cytokines and chemokines. The potential mechanism for this antiviral action and the containment of the inflammatory response by the drug associated with the engagement of terminal N-acetylgalactosamine GcMAF and C-type lectin domain containing 10A expressed at the surface of lung-infiltrating macrophages and pneumocytes, which simultaneously express angiotensin-converting enzyme 2, is discussed.

Introduction. RNA-containing viruses, especially influenza viruses, are of high epidemiological significance. The manifestation of COVID-19 has led to the registration of coinfection cases, the pathogenesis of which is poorly studied. The Vero (E6) cell line is widely used to study the morphogenesis of various viruses, including influenza and coronavirus. The aim of the work is to study the ultrastructure of Vero (E6) cells and the reproduction of viral particles during monoinfection with the influenza A virus and coinfection of this virus with two SARS-CoV-2 genovariants in dynamics 6, 18 and 24 hours after inoculation. Materials and methods. The Vero (E6) cell line model was used for in vitro study of the viral infection effects and an analysis of the dynamics of changes in the number of intracellular viral particles. The study involved 4 experimental groups: Vero (E6) cells mono-infected with the influenza virus strain A/H1N1 pmd09 at a dose of 0.1 MOI; Vero (E6) cells co-infected with the influenza virus strain A/H1N1 pmd09 and Delta strain of SARS-CoV-2 at a total dose of 0.1 MOI; Vero (E6) cells co-infected with the influenza virus strain A/H1N1 pmd09 and Omicron strain of SARS-CoV-2 at a total dose of 0.1 MOI. In each study group, cells were monitored at time points of 6, 18, and 24 hours. Results. After 6 h, no pathological structures were detected in all groups, except for virus-containing transport vesicles. After 18 h, vacuolization of the ER of varying degree was noted in all the studied groups. After 24 h, ultrastructural changes, namely vacuolization of organelles and/or compaction of the cytoplasm, were encountered in all groups comparatively more frequently than at 6 h and 18 h time points. . The dynamics of the number of viral particles increased significantly by 24 h time point in the monoinfection group. However, none of the coinfection groups demonstrated a tendency for the number of viral particles to change, since no statistically significant differences were found between the 6 h, 18 h, and 24 h stages. Conclusion. The results obtained suggested that the interaction between A/H1N1 pmd09 and SARS-CoV-2 viruses contributed to an overall decrease in the formation of new virions in Vero (E6) cells in both cases of coinfection.

In this study, we investigated the features of co-infection with SARS-CoV-2 and the enterovirus vaccine strain LEV8 of coxsackievirus A7 or enterovirus A71 for Vero E6 cells and Syrian hamsters. The investigation of co-infection with SARS-CoV-2 and LEV-8 or EV-A71 in the cell model showed that a competitive inhibitory effect for these viruses was especially significant against SARS-CoV-2. Pre-infection with enteroviruses in the animals caused more than a 100-fold decrease in the levels of SARS-CoV-2 virus replication in the respiratory tract and more rapid clearance of infectious SARS-CoV-2 from the lower respiratory tract. Co-infection with SARS-CoV-2 and LEV-8 or EV-A71 also reduced the severity of clinical manifestations of the SARS-CoV-2 infection in the animals. Additionally, the histological data illustrated that co-infection with strain LEV8 of coxsackievirus A7 decreased the level of pathological changes induced by SARS-CoV-2 in the lungs. Research into the chemokine/cytokine profile demonstrated that the studied enteroviruses efficiently triggered this part of the antiviral immune response, which is associated with the significant inhibition of SARS-CoV-2 infection. These results demonstrate that there is significant viral interference between the studied strain LEV-8 of coxsackievirus A7 or enterovirus A71 and SARS-CoV-2 in vitro and in vivo.

IntroductionRestoring immune tolerance is a promising area of therapy for autoimmune diseases. One method that helps restore immunological tolerance is the approach using tolerogenic dendritic cells (tolDCs). In our study, we analyzed the effectiveness of using dendritic cells transfected with DNA constructs encoding IL-10, type II collagen, and CCR9 to induce immune tolerance in an experimental model of arthritis.MethodsDendritic cell cultures were obtained from bone marrow cells of Balb/c mice. Dendritic cells (DCs) cultures were transfected with pmaxCCR9, pmaxIL-10, and pmaxCollagen type II by electroporation. The phenotype and functions of DCs were studied using enzyme-linked immunosorbent assay (ELISA) and flow cytometry. Migration of electroporated DCs was assessed in vitro. Induction of antigen-collagen induced arthritis (ACIA) was carried out according to the protocol in Balb/c mice. DCs were then administered to ACIA mice. The development of arthritis was monitored by measuring paw swelling with a caliper at different time points. The immunological changes were assessed by analyzing the content of antibodies to type II collagen using enzyme immunoassay. Additionally, a histological examination of the joint tissue was conducted, followed by data analysis.The results are as followsDCs were obtained, characterized by reduced expression of CD80, CD86, and H-2Db (MHC class I), increased expression of CCR9, as well as producing IL-10 and having migratory activity to thymus cells. Transfected DCs induced T-regulatory cells (T-reg) and increased the intracellular content of IL-10 and TGF-β in CD4+T cells in their co-culture, and also suppressed their proliferative activity in response to antigen. The administration of tolDCs transfected with DNA constructs encoding type II collagen, IL-10, and CCR9 to mice with ACIA demonstrated a reduction in paw swelling, a reduction in the level of antibodies to type II collagen, and a regression of histological changes.ConclusionThe study presents an approach by which DCs transfected with DNA constructs encoding epitopes of type II collagen, IL-10 and CCR9 promote the development of antigen-specific tolerance, control inflammation and reduce the severity of experimental arthritis through the studied mechanisms: induction of T-reg, IL-10, TGF-β.

Vitamin D3 transporter (DBP) is a multifunctional protein. Site-specific deglycosylation results in its conversion to group-specific component protein-derived macrophage activating factor (GcMAF), which is capable of activating macrophages. It has been shown that depending on precursor conversion conditions, the resulting GcMAF activates mouse peritoneal macrophages towards synthesis of either pro- (IL-1β, TNF-α—M1 phenotype) or anti-inflammatory (TGF-β, IL-10—M2 phenotype) cytokines. The condition for the transition of the direction of the inflammatory response of macrophages when exposed to GcMAF is the initial glycosylated state of the population of DBP molecules and the associated effective deglycosylation of DBP by β-galactosidase. In vivo experiments with GcMAF exhibiting anti-inflammatory properties on models of induced arthritis in mice and cystitis in rats indicate a significant anti-inflammatory effect of the macrophage activator. The feasibility of unidirectional induction of anti-inflammatory properties of macrophages allows creation of combined therapeutic platforms where M2 macrophages are among the key therapeutic components.

Immature hematopoietic progenitors are a constant source for renewal of hemocyte populations and the basic component of the tissue and cell repair apparatus. A unique property of these cells of internalizing extracellular double-stranded DNA has been previously shown. The leukostimulatory effect demonstrated in our pioneering studies was considered to be due to the feature of this cell. In the present research, we have analyzed the effects of DNA genome reconstructor preparation (DNAgr), DNAmix, and human recombinant angiogenin on both hematopoietic stem cells and multipotent progenitors. Treatment with bone marrow cells of experimental mice with these preparations stimulates colony formation by hematopoietic stem cells and proliferation of multipotent descendants. The main lineage responsible for this is the granulocyte-macrophage hematopoietic lineage. Using fluorescent microscopy as well as FACS assay, co-localization of primitive c-Kit- and Sca-1-positive progenitors and the TAMRA-labeled double-stranded DNA has been shown. Human recombinant angiogenin was used as a reference agent. Cells with specific markers were quantified in intact bone marrow and colonies grown in the presence of inducers. Quantitative analysis revealed that a total of 14,000 fragment copies of 500 bp, which is 0.2% of the haploid genome, can be delivered into early progenitors. Extracellular double-stranded DNA fragments stimulated the colony formation in early hematopoietic progenitors from the bone marrow, which assumed their effect on cells in G0. The observed number of Sca1+/c-Kit+ cells in colonies testifies to the possibility of both symmetrical and asymmetrical division of the initial hematopoietic stem cell and its progeny.

The SARS-CoV-2 pandemic has underscored the necessity for functional transgenic animal models for testing. Mouse lines with overexpression of the human receptor ACE2 serve as the common animal model to study COVID-19 infection. Overexpression of ACE2 under a strong ubiquitous promoter facilitates convenient and sensitive testing of COVID-19 pathology. We performed pronuclear microinjections using a 5 kb CAG-ACE2 linear transgene construct and identified three founder lines with 140, 72, and 73 copies, respectively. Two of these lines were further analyzed for ACE2 expression profiles and sensitivity to SARS-CoV-2 infection. Both lines expressed ACE2 in all organs analyzed. Embryonic fibroblast cell lines derived from transgenic embryos demonstrated severe cytopathic effects following infection, even at low doses of SARS-CoV-2 (0,1–1.0 TCID50). Infected mice from the two lines began to show COVID-19 clinical signs three days post-infection and succumbed between days 4 and 7. Histological examination of lung tissues from terminally ill mice revealed severe pathological alterations. To further characterize the integration site in one of the lines, we applied nanopore sequencing combined with Cas9 enrichment to examine the internal transgene concatemer structure. Oxford Nanopore sequencing (ONT) is becoming the gold standard for transgene insert characterization, but it is relatively inefficient without targeted region enrichment. We digested genomic DNA with Cas9 and gRNA against the ACE2 transgene to create ends suitable for ONT adapter ligation. ONT data analysis revealed that most of the transgene copies were arranged in a head-to-tail configuration, with palindromic junctions being rare. We also detected occasional plasmid backbone fragments within the concatemer, likely co-purified during transgene gel extraction, which is a common occurrence in pronuclear microinjections.

Introduction. The main characteristic of pathogenicity of SARS-CoV-2 virus is its ability to cause death in sensitive laboratory animals. The absence of lethal animal infection models requires the search for other approaches to assess pathogenicity. Studying infectious processes in guinea pig lungs allows us to identify the features and patterns of the disease, determine significant histological characteristics, and identify the main pathogenic criteria. We aimed to conduct a comparative pathomorphological study of lung tissue in guinea pigs infected with various SARS-CoV-2 variants using morphometric standardized criteria. Materials and methods. We looked at structural changes in the lungs of 96 outbred guinea pigs depending on COVID-19 pathogenesis. The animals were randomly divided into seven experimental and one control groups of 6 males and 6 females each. The experimental groups were intranasally infected with the following SARS-CoV-2 strains: Wuhan, Alpha, Beta, Gamma, Delta, and Omicron (the last one represented by two genetic lineages BA.5.2 and EG.5). The control group was intranasally injected with 0.9% NaCl. The viral load over time was determined with RT-PCR on nasal flushes and infection titer (on Vero E6 cell culture). The animals that survived were removed from the experiment on day 15. Pathomorphological examination was performed with optical microscopy of histological lung specimens. Results. We detected diffuse alveolar damage as a similar pathological feature in the lungs of both patients with COVID-19 and guinea pigs. Significant differences were revealed in the number of pathomorphological signs caused by various genetic variants of COVID-19. The most considerable pathological effect of SARS-CoV-2 on lung tissue was caused by the Gamma variant. We suppose guinea pig lung dystelectasis to be the main marker of pathogenicity of SARS-CoV-2 strains, since this sign differs significantly in SARS-CoV-2 variants and is associated with excessive cytokine activity. Alveolar hemorrhagic syndrome is characteristic of all the studied coronavirus variants except for Omicron EG.5. The genetic lineages of the omicron variant have a minimal pathological effect on lung tissue and do not cause death in guinea pigs. Conclusion. Guinea pig is an appropriate animal for a SARS-CoV-2 model. Pathomorphological changes in the lungs depend on the SARS-CoV-2 variant causing them. Keywords: SARS-CoV-2, COVID-19, guinea pig, lungs, pathomorphology, SARS-CoV-2 variants

The aim of the work was to study the features of the infectious process in the lungs of animals used as models for assessing SARS-CoV-2 pathogenicity.Materials and methods. The strain of SARS-CoV-2 alpha variant virus was used in the work. The experiments were carried out on linear and transgenic mice, Syrian hamsters, guinea pigs, ferrets and two types of primates: rhesus macaques and green monkey. The pathomorphological examination was performed by optical microscopy of histological lung preparations using a computerized microscope with digital microphotography.Results and discussion. A comparative histological analysis of the lungs in six different types of laboratory animals was carried out when modeling a new coronavirus infection; similar morphometric signs of the severity of the disease caused by the SARS-CoV-2 virus in sensitive animals were determined, and a dose-dependent correlation of pathological changes in lung tissues with intranasal administration of various infectious doses was revealed. The features of pathomorphological changes in six different animal species in the simulation of a new coronavirus infection have been characterized, and their dose-dependent nature determined. The presented research results can be used to select a model animal for the purpose of in-depth study of the pathogenesis of COVID-19 caused by newly isolated coronavirus variants, the dynamics of immune reactions of the body during the development of the disease, as well as in vivo studies of the protective effect of promising therapeutic drugs and vaccines.

Tumor necrosis factor alpha (TNF-α) is a cytokine that is responsible for many processes associated with immune response and inflammation. It is involved in the development of an antiviral response to many virus infections. This factor was shown to be activated in influenza A virus infection, which enhances production of other cytokines. The overexpression of these cytokines can lead to a cytokine storm. To study the role of TNF-α in the development of pathologies associated with viral infection, we generated a Tnfa knockout mouse strain. We demonstrated that these mice were characterized by a significant increase in the number of viral genomes compared to that in the parental strain, but the amount of live virus did not differ. A histopathology of the lungs in the genetically modified animals was significantly lower in terms of interalveolar septal infiltration. The generated model may be used to further study pathological processes in viral infections.

Introduction. Since early May 2022, more than 90,000 cases of monkeypox virus infection have been reported in more than 70 countries around the World. This is the largest outbreak of monkeypox ever recorded outside of Africa. The aim of the study is to confirm the first case of monkeypox in Russia, to isolate and sequence a new strain of monkeypox virus (MPXV), and to assess its sensitivity to the 7-[N-(4-trifluoromethylbenzoyl)-hydrazinocarbonyl]-tricyclo-[3.2.2.0^2,4]non-8-en-6-carboxylic acid (NIOCH-14) antipox drug. Materials and methods. The biological materials obtained from the affected area of the skin (contents of vesicles), a nasopharyngeal smear, sputum and venous blood from a patient with suspected monkeypox were used. The disease was confirmed by PCR followed by determination of the nucleotide sequence of viral DNA by sequencing. Isolation of the new MPXV strain from clinical samples was carried out in Vero E6 cells. The antiviral effectiveness of NIOCH-14 against the new MPXV strain was assessed using an adapted spectrophotometric method. Results. A diagnostic study of the biological samples of a patient who returned from a tourist trip to European countries with complaints of skin rashes all over the body revealed MPXV DNA. A new strain of MPXV was isolated from vesicles in Vero E6 cells, and the genomic sequence MPXV-pustule S45 was assembled using high-throughput parallel sequencing (NGS). Discussion. The effectiveness of the finished dosage form of NIOCH-14 against the new strain of MPXV based on the results of determining the 50% virus inhibitory concentration (IC50) was 0.02 μg/mL, and the selectivity index (SI) was 15,000. Conclusion. In this study, the pathogen of monkeypox was detected and identified using real-time PCR, NGS and electron microscopy, and the first imported case of this disease in Russia was confirmed. It has been proven that the drug NIOCH-14 exhibits high antiviral activity in vitro against the new MPXV strain.

We present a series of articles proving the existence of a previously unknown mechanism of interaction between hematopoietic stem cells and extracellular double-stranded DNA (and, in particular, double-stranded DNA of the peripheral bloodstream), which explains the possibility of emergence and fixation of genetic information contained in double-stranded DNA of extracellular origin in hematopoietic stem cells. The concept of the possibility of stochastic or targeted changes in the genome of hematopoietic stem cells is formulated based on the discovery of new, previously unknown biological properties of poorly differentiated hematopoietic precursors. The main provisions of the concept are as follows. The hematopoietic stem cell takes up and internalizes fragments of extracellular double-stranded DNA via a natural mechanism. Specific groups of glycocalyx factors, including glycoproteins/proteoglycans, glycosylphosphatidylinositol-anchored proteins and scavenger receptors, take part in the internalization event. The binding sites for DNA fragments are heparin-binding domains and clusters of positively charged amino acid residues that are parts of protein molecules of these factors. Extracellular fragments delivered to the internal compartments of hematopoietic stem cells initiate terminal differentiation, colony formation, and proliferation of hematopoietic precursors. The molecular manifestation of these processes is the emergence and repair of pangenomic single-strand breaks. The occurrence of pangenomic single-strand breaks and restoration of genome (genomic DNA) integrity are associated with activation of a “recombinogenic situation” in the cell; during its active phase, stochastic homologous recombination or other recombination events between extracellular fragments localized in the nucleus and chromosomal DNA are possible. As a result, genetic material of initially extracellular localization either integrates into the recipient genome with the replacement of homologous chromosomal segments, or is transitively present in the nucleus and can manifest itself as a new genetic trait. It is assumed that as a result of stochastic acts of homologous exchange, chromosome loci are corrected in hematopoietic stem cells that have acquired mutations during the existence of the organism, which are the cause of clonal hematopoiesis associated with old age. In this regard, there is a fundamental possibility of changing the hematopoietic status of hematopoietic stem cells in the direction of polyclonality and the original diversity of clones. Such events can form the basis for the rejuvenation of the blood-forming cell system. The results of the laboratory’s work indicate that other stem cells in the body capture extracellular DNA fragments too. This fact creates a paradigm for the overall rejuvenation of the body.

In this part of the study, the first component of the concept of “natural genome reconstruction” is being proven. It was shown with mouse and human model organisms that CD34+ hematopoietic bone marrow progenitors take up fragments of extracellular double-stranded DNA through a natural mechanism. It is known that the process of internalization of extracellular DNA fragments involves glycocalyx structures, which include glycoproteins/protein glycans, glycosylphosphatidylinositol-anchored proteins and scavenger receptors. The bioinformatic analysis conducted indicates that the main surface marker proteins of hematopoietic stem cells belong to the indicated groups of factors and contain specific DNA binding sites, including a heparin-binding domain and clusters of positively charged amino acid residues. A direct interaction of CD34 and CD84 (SLAMF5) glycoproteins, markers of hematopoietic stem cells, with double-stranded DNA fragments was demonstrated using an electrophoretic mobility shift assay system. In cells negative for CD34, which also internalize fragments, concatemerization of the fragments delivered into the cell occurs. In this case, up to five oligonucleotide monomers containing 9 telomeric TTAGGG repeats are stitched together into one structure. Extracellular fragments delivered to hematopoietic stem cells initiate division of the original hematopoietic stem cell in such a way that one of the daughter cells becomes committed to terminal differentiation, and the second retains its low-differentiated status. After treatment of bone marrow cells with hDNAgr, the number of CD34+ cells in the colonies increases to 3 % (humans as the model organism). At the same time, treatment with hDNAgr induces proliferation of blood stem cells and their immediate descendants and stimulates colony formation (mouse, rat and humans as the model organisms). Most often, the granulocyte-macrophage lineage of hematopoiesis is activated as a result of processing extracellular double-stranded DNA. The commitment process is manifested by the appearance and repair of pangenomic single-strand breaks. The transition time in the direction of differentiation (the time it takes for pangenomic single-strand breaks to appear and to be repaired) is about 7 days. It is assumed that at the moment of initiation of pangenomic single-strand breaks, a “recombinogenic situation” ensues in the cell and molecular repair and recombination mechanisms are activated. In all experiments with individual molecules, recombinant human angiogenin was used as a comparison factor. In all other experiments, one of the experimental groups consisted of hematopoietic stem cells treated with angiogenin.

INTRODUCTION:Intranasal vaccination using live vector vaccines based on non-pathogenic or slightly pathogenic viruses is the one of the most convenient, safe and effective ways to prevent respiratory infections, including COVID-19. Sendai virus is the best suited for this purpose, since it is respiratory virus and is capable of limited replication in human bronchial epithelial cells without causing disease. The aim of the work is to design and study the vaccine properties of recombinant Sendai virus, Moscow strain, expressing secreted receptor-binding domain of SARS-CoV-2 Delta strain S protein (RBDdelta) during a single intranasal immunization.MATERIALS AND METHODS:Recombinant Sendai virus carrying insertion of RBDdelta transgene between P and M genes was constructed using reverse genetics and synthetic biology methods. Expression of RBDdelta was analyzed by Western blot. Vaccine properties were studied in two models: Syrian hamsters and BALB/c mice. Immunogenicity was evaluated by ELISA and virus-neutralization assays. Protectiveness was assessed by quantitation of SARS-CoV-2 RNA in RT-PCR and histological analysis of the lungs.RESULTS:Based on Sendai virus Moscow strain, a recombinant Sen-RBDdelta(M) was constructed that expressed a secreted RBDdelta immunologically identical to natural SARS-CoV-2 protein. A single intranasal administration of Sen-RBDdelta(M) to hamsters and mice significantly, by 15 and 107 times, respectively, reduced replicative activity of SARS-CoV-2 in lungs of animals, preventing the development of pneumonia. An effective induction of virus-neutralizing antibodies has also been demonstrated in mice.CONCLUSION:Sen-RBDdelta(M) is a promising vaccine construct against SARS-CoV-2 infection and has a protective properties even after a single intranasal introduction.

It is well-established that double-stranded RNA (dsRNA) exhibits noticeable radioprotective and radiotherapeutic effects. The experiments conducted in this study directly demonstrated that dsRNA was delivered into the cell in its native form and that it induced hematopoietic progenitor proliferation. The 68 bp synthetic dsRNA labeled with 6-carboxyfluorescein (FAM) was internalized into mouse hematopoietic progenitors, c-Kit+ (a marker of long-term hematopoietic stem cells) cells and CD34+ (a marker of short-term hematopoietic stem cells and multipotent progenitors) cells. Treating bone marrow cells with dsRNA stimulated the growth of colonies, mainly cells of the granulocyte–macrophage lineage. A total of 0.8% of Krebs-2 cells internalized FAM-dsRNA and were simultaneously CD34+ cells. dsRNA in its native state was delivered into the cell, where it was present without any signs of processing. dsRNA binding to a cell was independent of cell charge. dsRNA internalization was related to the receptor-mediated process that requires energy from ATP. Synthetic dsRNA did not degrade in the bloodstream for at least 2 h. Hematopoietic precursors that had captured dsRNA reinfused into the bloodstream and populated the bone marrow and spleen. This study, for the first time, directly proved that synthetic dsRNA is internalized into a eukaryotic cell via a natural mechanism.

Introduction. The COVID-19 pandemic combined with seasonal epidemics of respiratory viral diseases requires targeted antiviral prophylaxis with restorative and immunostimulant drugs. The compounds of natural origin are low-toxic, but active against several viruses at the same time. One of the most famous compounds is Inonotus obliquus aqueous extract. The fruit body of basidial fungus I. obliquus is called Chaga mushroom. The aim of the work ‒ was to study the antiviral activity of I. obliquus aqueous extract against the SARS-CoV-2 virus in vivo. Materials and methods. Antiviral activity of I. obliquus aqueous extract sample (#20-17) was analyzed against strain of SARS-CoV-2 Omicron ВА.5.2 virus. The experiments were carried out in BALB/c inbred mice. The SARS-CoV-2 viral load was measured using quantitative real-time PCR combined with reverse transcription. The severity of lung tissue damage was assessed by histological methods. Results. The peak values of the viral load in murine lung tissues were determined 72 hours after intranasal inoculation at dose of 2,85 lg TCID50. The quantitative real-time PCR testing has shown a significant decrease in the viral load compared to the control group by 4,65 lg copies/ml and 5,72 lg copies/ml in the lung tissue and nasal cavity samples, respectively. Histological methods revealed that the decrease in the number and frequency of observed pathomorphological changes in murine lung tissues depended on the introduction of the compound under study. Conclusion. The results obtained indicate the possibility of using basidial fungus Inonotus obliquus aqueous extract as a preventive agent against circulating variants of SARS-CoV-2 virus.

INTRODUCTION:The COVID-19 pandemic combined with seasonal epidemics of respiratory viral diseases requires targeted antiviral prophylaxis with restorative and immunostimulant drugs. The compounds of natural origin are low-toxic, but active against several viruses at the same time. One of the most famous compounds is Inonotus obliquus aqueous extract. The fruit body of basidial fungus I. obliquus is called Chaga mushroom. The aim of the work ‒ was to study the antiviral activity of I. obliquus aqueous extract against the SARS-CoV-2 virus in vivo. MATERIALS AND METHODS:Antiviral activity of I. obliquus aqueous extract sample (#20-17) was analyzed against strain of SARS-CoV-2 Omicron ВА.5.2 virus. The experiments were carried out in BALB/c inbred mice. The SARS-CoV-2 viral load was measured using quantitative real-time PCR combined with reverse transcription. The severity of lung tissue damage was assessed by histological methods. RESULTS:The peak values of the viral load in murine lung tissues were determined 72 hours after intranasal inoculation at dose of 2,85 lg TCID50. The quantitative real-time PCR testing has shown a significant decrease in the viral load compared to the control group by 4,65 lg copies/ml and 5,72 lg copies/ml in the lung tissue and nasal cavity samples, respectively. Histological methods revealed that the decrease in the number and frequency of observed pathomorphological changes in murine lung tissues depended on the introduction of the compound under study. CONCLUSION:The results obtained indicate the possibility of using basidial fungus Inonotus obliquus aqueous extract as a preventive agent against circulating variants of SARS-CoV-2 virus.