Background: Torque teno virus (TTV) is a circular, single-stranded DNA virus that chronically infects healthy individuals of all ages worldwide. There is a lot of data on the prevalence and genetic heterogeneity of TTV in healthy populations and in patients with various diseases now available. However, little is known about TTV load among healthy human population. In this study we analyzed TTV load in the group of 512 Russian elite athletes, who are supposed to be, by some standards, the healthiest part of the human population.

Background: First vaccines for prevention of Coronavirus disease 2019 (COVID-19) are becoming available but there is a huge and unmet need for specific forms of treatment. In this study we aimed to evaluate the anti-SARS-CoV-2 effect of siRNA both in vitro and in vivo.Methods: To identify the most effective molecule out of a panel of 15 in silico designed siRNAs, an in vitro screening system based on vectors expressing SARS-CoV-2 genes fused with the firefly luciferase reporter gene and SARS-CoV-2-infected cells was used. The most potent siRNA, siR-7, was modified by Locked nucleic acids (LNAs) to obtain siR-7-EM with increased stability and was formulated with the peptide dendrimer KK-46 for enhancing cellular uptake to allow topical application by inhalation of the final formulation -siR-7-EM/KK-46. Using the Syrian Hamster model for SARS-CoV-2 infection the antiviral capacity of siR-7-EM/KK-46 complex was evaluated. Results:We identified the siRNA, siR-7, targeting SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) as the most efficient siRNA inhibiting viral replication in vitro.Moreover, we showed that LNA-modification and complexation with the designed peptide dendrimer enhanced the antiviral capacity of siR-7 in vitro. We demonstrated significant reduction of virus titer and lung inflammation in animals exposed to inhalation of siR-7-EM/KK-46 in vivo.Conclusions: Thus, we developed a therapeutic strategy for COVID-19 based on inhalation of a modified siRNA-peptide dendrimer formulation. The developed medication is intended for inhalation treatment of COVID-19 patients. | 2841KHAITOV eT Al.

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The effects of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in women on the gestation course and the health of the fetus, particularly in the first and second trimesters, remain very poorly explored. This report describes a case in which the normal development of pregnancy was complicated immediately after the patient had experienced Coronavirus disease 2019 (COVID-19) at the 21st week of gestation. Specific conditions included critical blood flow in the fetal umbilical artery, fetal growth restriction (1st percentile), right ventricular hypertrophy, hydropericardium, echo-characteristics of hypoxic-ischemic brain injury (leukomalacia in periventricular area) and intraventricular hemorrhage at the 25th week of gestation. Premature male neonate delivered at the 26th week of gestation died after 1 day 18 h due to asystole. The results of independent polymerase chain reaction (PCR), mass spectrometry and immunohistochemistry analyses of placenta tissue, umbilical cord blood and child blood jointly indicated vertical transmission of SARS–CoV-2 from mother to the fetus, which we conclude to be the major cause for the development of maternal vascular malperfusion in the studied case.

α-Actinin-3 (ACTN3) has been proposed to regulate skeletal muscle differentiation and hypertrophy through its interaction with the signalling protein calcineurin. Since the inhibition of calcineurin potentiates the production of testosterone, we hypothesized that α-actinin-3 deficiency (predicted from the ACTN3 XX genotype) may influence serum levels of testosterone of athletes. Objective: To investigate the association of ACTN3 gene R577X polymorphism with resting testosterone levels in athletes. Methods: A total of 209 elite Russian athletes from different sports (119 males, 90 females) were genotyped for ACTN3 gene R577X polymorphism by real-time PCR. Resting testosterone was examined in serum of athletes using enzyme immunoassay. Results: The mean testosterone levels were significantly higher in both males and females with the ACTN3 R allele than in XX homozygotes (males: RR: 24.9 (5.7), RX: 21.8 (5.5), XX: 18.6 (4.9) ng · mL-1, P = 0.0071; females: RR: 1.43 (0.6), RX: 1.21 (0.71), XX: 0.79 (0.66) ng · mL-1, P = 0.0167). Conclusions: We found that the ACTN3 R allele was associated with high levels of testosterone in athletes, and this may explain, in part, the association between the ACTN3 RR genotype, skeletal muscle hypertrophy and power athlete status.

BackgroundPreviously we demonstrated that the resection of primary 4T1 tumors only slightly prolongs mouse survival, but importantly, creates a “window of opportunity” with attenuated suppressor cell and increased activated T cell populations. This suggests that additional activation of the immune system by immunostimulatory agents during this period may enhance anti-tumor immunity and potentially eradicate micro-metastatic disease in this stringent model.We hypothesized that the immunostimulator Immunomax®, which is comprised of a plant-derived polysaccharide, is non-toxic in humans and stimulates immune defense during the infectious diseases treatment, may have also anti-tumor activity and be beneficial in the adjuvant setting when endogenous anti-tumor responses are present and during the “window of opportunity” in post-resection metastatic breast cancer model. Here we provide the initial report that Immunomax® demonstrates the capacity to eliminate micro-metastatic disease in the post-resection, 4T1 mouse model of breast cancer.MethodsThe efficacy of Immunomax® was evaluated by analyzing survival rate and the number of spontaneous clonogenic tumor cells in the lung homogenates of mice. The frequencies of activated NK, CD4+ and CD8+ cells as well as myeloid-derived suppressor cells and Treg cells were evaluated using flow cytometry. Highly purified mouse and human dendritic and NK cells were sorted and the effect of Immunomax® on activation status of these cells was assessed by flow cytometry. The property of Immunomax® as TLR-4 agonist was determined by NF-κB/SEAP reporter gene assay, WB, RT-PCR.ResultsImmunomax® injections significantly prolonged overall survival and cured 31% of mice. This immunostimulator activates DCs via the TLR-4, which in turn stimulates tumoricidal NK cells and in vitro, completely inhibits growth of 4T1 cells. Incubation of PBMC from healthy donors with Immunomax® activates NK cells via activation of plasmacytoid DC leading significantly higher efficacy in killing of human NK-target cells K562 compared with non-treated cells.ConclusionThis is the first demonstration that Immunomax® is a TLR-4 agonist and the first report of a documented role for this pharmaceutical grade immunostimulator in augmenting anti-tumor activity, suggesting that incorporation of Immunomax® into developing breast cancer therapeutic strategies may be beneficial and with less potential toxicity than checkpoint inhibitors.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-014-0322-y) contains supplementary material, which is available to authorized users.

Immuno-PCR (iPCR) is one of the methods used for the detection of a wide range of analytes and features the high sensitivity of the polymerase chain reaction (PCR) method. iPCR uses antibodies coupled to DNA, followed by the amplification of the attached DNA using RT-PCR. Two major types of antibody-DNA conjugates are currently used, which are obtained as a result of non-covalent (biotin-streptavidin) or covalent interactions. Using a strain-promoted azide-alkyne cycloaddition (SPAAC), we synthesized covalent DNA-antibody conjugates, optimized the reaction conditions, and developed an efficient protocol for the purification of conjugates, with which all unreacted antibodies and oligonucleotides are separated. Covalent DNA-antibody conjugates were tested with iPCR assays that were previously developed for the detection of IgE and IgM antibodies with the use of the supramolecular complex of 5'- and 3'-biotinylated DNA and streptavidin. The results show that the modification of antibodies with amino groups did not allow us to obtain monolabeled antibodies or antibodies with a strictly defined number of DNA-labels. The degree of labeling determined by the dyes introduced through the azido group reflects the actual labeling degree statistically. If the average labeling degree for azido groups is 1.1, the conjugates contain 25% mono-labeled antibodies, 50% double-labeled antibodies, and 25% unlabeled ones. The specificity of the monoclonal antibody to human IgE (BE5) changed after conjugation with the oligonucleotide. The sensitivity of iPCR in the detection of IgM antibodies produced against the LeC disaccharide using a covalent conjugate was similar to that of a supramolecular complex of 5'- and 3'-biotinylated DNA and streptavidin, but the new procedure is two steps shorter.

Exercise-induced oxidative stress is a state that primarily occurs in athletes involved in high-intensity sports when pro-oxidants overwhelm the antioxidant defense system to oxidize proteins, lipids, and nucleic acids. During exercise, oxidative stress is linked to muscle metabolism and muscle damage, because exercise increases free radical production. The T allele of the Ala16Val (rs4880 C/T) polymorphism in the mitochondrial superoxide dismutase 2 (SOD2) gene has been reported to reduce SOD2 efficiency against oxidative stress. In the present study we tested the hypothesis that the SOD2 TT genotype would be underrepresented in elite athletes involved in high-intensity sports and associated with increased values of muscle and liver damage biomarkers. The study involved 2664 Caucasian (2262 Russian and 402 Polish) athletes. SOD2 genotype and allele frequencies were compared to 917 controls. Muscle and liver damage markers [creatine kinase (CK), creatinine, alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP)] were examined in serum from 1444 Russian athletes. The frequency of the SOD2 TT genotype (18.6%) was significantly lower in power/strength athletes (n = 524) compared to controls (25.0%, p = 0.0076) or athletes involved in low-intensity sports (n = 180; 33.9%, p < 0.0001). Furthermore, the SOD2 T allele was significantly associated with increased activity of CK (females: p = 0.0144) and creatinine level (females: p = 0.0276; males: p = 0.0135) in athletes. Our data show that the SOD2 TT genotype might be unfavorable for high-intensity athletic events.