High-Throughput Platform for B-Cell Screening Based on Fluorescent Phage-Display Technology.

A system for detection of malignantly transformed cells, including follicular lymphoma Bcells, was developed and experimentally validated. The system is based on the use of bacteriophages carrying exposed ligands for pathogenic B-cell receptors. The efficiency of binding to target cells is several times higher than in systems with chemically synthesized biotinylated peptides. The new method is proposed as a noninvasive diagnostic test for mapping B-cell lymphoma and for determining the specificity of B-cell receptors and high-throughput combinatorial selection of various repertories of B cells.

Simultaneous Percutaneous Coronary Intervention and Endovascular Closure of Atrial Septal Defect in Adults.

AIM: to assess clinical efficacy and expediency (appropriateness) of simultaneous single stage combined coronary stenting and closure of atrial septal defect. MATERIALS AND METHODS: Of total number of patients who underwent endovascular correction of atrial septal defect (ASD) (n=91), in 6 (6.6 %) the procedure of endovascular repair of secondary ASD was combined with performed at same session oronary stenting. Mean age of these patients was 63±6.4 years. Mean diameter of ASD according to transesophageal echocardiography was 13.7±3.1 мм (from 10 to 17 mm). Two patients had dysplasia of atrial septum with pronounced aneurysmal protrusion in the right atrial cavity. Estimate of coronary arteries (CA) involvement SYNTAX score was 14.5±4.9. RESULTS: At initial stage we performed coronary stenting, then ASD closure with occluder. Technical success of combined endovascular procedures was 100 %. Six ASD occluders were implanted in 6 patients. Mean occluder diameter was 21±7,3 mm. Immediately after occluder implantation complete defect closure was achieved in 5 cases, in one case small residual shunt was observed. CA stenting procedure, in one patient after successful recanalization of chronic CA occlusion, in all cases was fulfilled without complications. At control examination after 13.5±1.5 months complete closure of defects was preserved. In all cases significant reduction of right heart chambers occurred. According to echocardiography right atrial volume decreased from 48.6±5.6 to 32.6±3.3 cm3, right ventricular volume - from 45.2±5.1 to 33.4±3.8 cm3, systolic pulmonary pressure fell from 49.7±8.6 to 32.6±6.9 mm Hg. According to control coronary angiography good effect of endovascular procedures was preserved. Tolerance to exercise rose from 68.5±11.8 до 85.3±12.4 W. CONCLUSION: Same time CA stenting and endovascular ASD closure appears to be safe and effective procedure. The strategy used was not associated with additive risk for a patient and shortened duration of hospital stay.

[The role of «Youth and aging proteins¼ in essential hypertension pathogenesis.]

Essential hypertension (EG) is an age-associated disease. Often EG of elderly patients haven't good way of treatment. Thus, the search of new target molecules for EG therapy is an actual goal of gerontology and molecular medicine. It was shown, that during EG concentrations of GDF11 «youth protein¼ decreased in 3,3 times and GDF15, JAM-A/1, CCL11 «aging proteins¼ increased in 1,4-2,4 times. EG patients have abnormal microcirculation processes. It was shown as decreasing in 1,3 and 1,7 times of hemodynamic HI1 and H1-H3 indexes. EG patients have negative correlation of GDF15 concentration with arterial pressure. EG patients have no correlation of JAM-A/1 concentration with arterial pressure. Normal is positive correlation with GDF15, JAM-A/1 concentration with arterial pressure. GDF15 blood level during EG have positive correlation with HI1-HI3 and negative correlation with NEUR_HI2 and MAYER_HI3 indexes. It can show on pathogenesis mechanisms of endothelial and smooth muscles function of vessels tissues. We suppose, that the regulation of GDF11, GDF15, JAM-A/1, CCL11 «youth and aging proteins¼ can be target object of EG therapy.

Comparative Analysis of Polyethyleneimine Efficiency for Improvement of Plasmid DNA Bioavailability.

We studied the influence of the type and structure of polyethyleneimine on bioavailability and expression of plasmid DNA carrying IGF-1 gene. Polymers with different molecular weights (2.5, 10, 25, and 60 kDa) of linear and branching structure were studied. It was found that the time of polyplex circulation in the blood did not exceed 24 h and the maximum concentration of plasmid DNA was attained with complexes with a molecular weight of 60 kDa. Analysis of liver samples showed that administration of 60-kDa branched polyethyleneimine complex provides DNA protection from degradation for 4 h; in 24 h from the start of the experiment, its concentration was significantly higher than the concentration of other studied polyethyleneimines. The expression of plasmid IGF-1 DNA for this complex attained maximum in 4 h and was equal to 15.50 (7.98; 21.98) arb. units/ml. These results allow us to recommend using polyethyleneimines with branched structure and a molecular weight of 60 kDa for improving plasmid DNA protection and bioavailability.

Directed Change in TNFα Specificity to Create DR5 Antagonists.

Death receptor 5 (DR5) is a promising target for antitumor therapy due to its high expression on different tumor cells. Resistance of various tumor cells against TRAIL, a natural ligand for the death receptors, reduces its therapeutic potential and prompts the search for novel agonists at these receptors. Previous screening across the combinatorial peptide library yielded a peptide sequence KVVLTHR that specifically binds DR5. Incorporation of this sequence into TNFα resulted in binding DR5 with mutant protein TNFα-mut and appearance of cytotoxicity against lymphoma cells.

Combinatorial Screening of Peptides, Specific Ligands of Death Receptor DR5.

Death receptors, in particular DR5, are highly attractive targets of antitumor therapy. The major limitation to application of natural death receptor ligands (TRAIL) is their non-specific cytotoxicity against normal cells. Since TRAIL can also bind decoy receptors (DcR) and prevent induction of apoptosis, the search for new DR-specific ligands is a topical issue. In the present study, we used combinatorial phage display peptide libraries to select a panel of DR5-binding amino acid sequences. A comparative analysis of the selected peptides enabled identification of the consensus sequence responsible for binding to DR5. Integration of this motif into polypeptide cytotoxic agents may provide targeted elimination of malignantly transformed cells.

Design of Chemical Conjugate for Targeted Therapy of Multiple Sclerosis Based of Constant Fragment of Human Antibody Heavy Chain and Peptoid Analog of Autoantigen MOG35-55.

Elimination of B cells producing autoantibodies to neuroantigens is considered as beneficial in the treatment of multiple sclerosis. Myelin oligodendrocyte glycoprotein (MOG) is a significant autoantigen in multiple sclerosis. It was shown that MOG-like peptoid AMogP3 can bind autoantibodies produced by pathological lymphocytes. We propose a structure of an innovative drug for targeted elimination of the pool of autoreactive B cells responsible for multiple sclerosis pathogenesis; this compound is a complex of peptoid AMogP3 with Fc fragment of human immunoglobulin. The obtained Fc-PEG-AMogP3 conjugate effectively interact with autoreactive antibodies, which attests to their high therapeutic potential.

A Study of the Protective Properties of an Antibody-Based Antidote Metabolizing Organophosphorus Pesticide Paraoxon.

A catalytic antibody A17 and its mutants highly efficiently interact with organophosphorus pesticide paraoxon. In this work, we studied the protective properties of antibody A17-K47 in paraoxon poisoning using a mouse model. The optimal paraoxon dose simulating the acute toxic effect of organophosphorus compounds was 550 µg/kg. The pharmacokinetic parameters of A17-K47 antibody were t1/2distr =7.2±1.4 min, t1/2el =330±20 min. The antibody did not cause toxic effects when administered at a ten-fold calculated therapeutic dose (610 mg/kg). The drug did not reduce mortality from acute paraoxon poisoning; however, the absence of drug toxicity opens up prospects for its use in symptomatic treatment of chronic paraoxon poisoning.

Study of Fibronectin Type III-Like Domains Role in Activation of gp130 Receptor.

Chimeric gp130 receptors were produced to study the role of three fibronectin type III-like domains in activation of gp130 receptor machinery. The ligand-induced dimerization of gp130 was sufficient to trigger STAT3 signaling pathway. These findings can be used as the basis in designing novel therapeutic gp130 inhibitors.

[Endoscopic treatment of cicatricial esophageal strictures using ionized argon plasma (with commentary)]. / Endoskopicheskoe lechenie rubtsovykh striktur pishchevoda s ispol'zovaniem ionizirovannoi argonovoi plazmy (s kommentariem).

AIM: To estimate an efficacy of endoscopic treatment of benign cicatricial esophageal strictures using ionized argon plasma. MATERIAL AND METHODS: It is presented treatment of 35 patients with post-ambustial and 17 patients with peptic strictures of the esophagus. Their age ranged from 17 to 73 years. Esophageal stenosis achieved 0.1-1.0 cm. Local, tubular and prolonged stenoses were observed in 18 (51.5%), 3 (8.6%) and 14 (39.9%) patients respectively. Peptic stenoses were local in 100% of cases and localized in lower esophagus before cardia. RESULTS: Endoscopic techique resulted good and satisfactory outcomes in 53.8% and 27% of cases within short terms. The best results were obtained in case of local post-ambustial and peptic ulcers. At the end of treatment inflammation and mitotic activity were decreased in mucosa and submucosa of the narrowed segment. Also thickness of the epithelium and stratification of tissue layers were normal. Connective tissue volume was moderately decreased in subepithelial area.

CHANGES IN BURNING WOUND IN APLLICATION OF MAGNETO-PLASMATIC THERAPY.

The article analyzed treatments results of 76 patients with deep burns. It was stated that magneto-plasma therapy facilitated to the change from an inflammatory type of burn cytograms to regenerative type. These changes were expressed in reducing of neutrophil quantity in wound on the fifth day, decreasing of degenerative changed leukocytes on the tenth day, increasing of phagocytic activity of neutrophils in all terms, growing of macrophage reaction by the fifth day and multiplication of fibroblast quantity in wound on the tenth day. The neutrophil and macrophage quantities were increased in the wound after necrectomy due to influence of magneto-plasmatic therapy on the third day. There was synthesized interleukin-8. The quantity of cells producing IL-8 was reduced on the tenth day.

The Effect of Kinesitherapy Exercises on the Level of Irisin among Females with Cardio-vascular diseases depending on the body mass and hormonal status.

The observation was conducted on 41 female subjects age 32 to 69 with compensated cardio-vascular diseases. 23 of the subjects had an increased body mass index (BMI). It was established that the older the females the less of the irisin muscle hormone is found in the blood. In the subjects with a higher BMI the level of irisin in the blood is also higher. Direct correlations were found between the level of irisin and the level of female sex hormones - estrogen and progesterone. Under the effect of kinesitherapy exercises the level of irisin in females with normal BMI increases; whereas in the females with a higher BMI it generally stays the same or is decreased. The characteristics of irisin's response to the kinesitherapy exercises depends on its original level, the intensity of physical exercise and the subject's physique.

[STEM CELLS PLAY NO CONSIDERABLE ROLE IN CARDIOMYOCYTE REPOPULATION OF ADULT HUMAN HEART].

There are two viewpoints concerning cardiac regeneration. One assumes that the myocardium of an adult human heart has a weak regenerative capacity. According to another, myocardium can renew at a high rate due to the presence of resident stem cells. This study was aimed to test the role of stem cells in myocardium repopulation in adult humans of different age by examining the distribution of cardiomyocytes as to their size and ploidy. Cytofluorimetry and interferometry were used to determine the dry weight, volume and ploidy of myocytes isolated from the left ventricle of the normal heart of 12 men aged 20-30 years (n = 7) and 40-50 years (n = 5). Dry weight of cardiomyocytes made up 6906 ± 182 pg (10(-12) g) aged 20-30 years and 9126 ± 263 pg in men aged 40-50 years. There were no cells with an intermediate volume between amplifying and mature myocytes. The number of candiomyocytes in the left ventricle made up (3.18 ± 0.05) x 10(9) cells in the age group 20-30 years and (2.06 ± 0.6) x 10(9) cells in the age group 40-50 years. Most of the myocyte population was represented by mononucleate cells with tetraploid nuclei (41.3%). Proportion of myocytes of different ploidy classes did not change in the interval from 20 to 50 years. Our results strongly suggest that stem cells of the heart are not involved in the regeneration of human myocardium during aging. The function of the aging heart is mostly compensated by the hypertrophy of the remaining myocytes.

[Experimental comparative pharmacokinetics of levofloxacin, triazavirin, and related conjugate].

A comparative study of the pharmacokinetics of levofloxacin and triazavirine as well as 2-methylthio-6-nitro-1,2,4-triazolo[5,1-ñ]-1,2,4-triazine-7(4Í)-ide (3S)-(-)-9-fluoro-2,3-dihydro-3-methyl-10-(4-methylpiperazin-1-yl)-7-oxo-7H-pyrido[1,2,3-d,e]-1,4-benzoxazine-6-carboxylic acid (conjugate 2) obtained by conjugation of triazavirine and levofloxacin, representing a new class of pharmacological agents, was carried out in experiments on rats. It is established that conjugate 2 in comparison to individual levofloxacin and triazavirine has a higher relative bioavailability and lower rate of elimination, which can lead to improved effectiveness of therapy at reduced dose and frequency of drug administration.

Heat shock induces production of reactive oxygen species and increases inner mitochondrial membrane potential in winter wheat cells.

Heat shock leads to oxidative stress. Excessive ROS (reactive oxygen species) accumulation could be responsible for expression of genes of heat-shock proteins or for cell death. It is known that in isolated mammalian mitochondria high protonic potential on the inner membrane actuates the production of ROS. Changes in viability, ROS content, and mitochondrial membrane potential value have been studied in winter wheat (Triticum aestivum L.) cultured cells under heat treatment. Elevation of temperature to 37-50°C was found to induce elevated ROS generation and increased mitochondrial membrane potential, but it did not affect viability immediately after treatment. More severe heat exposure (55-60°C) was not accompanied by mitochondrial potential elevation and increased ROS production, but it led to instant cell death. A positive correlation between mitochondrial potential and ROS production was observed. Depolarization of the mitochondrial membrane by the protonophore CCCP inhibited ROS generation under the heating conditions. These data suggest that temperature elevation leads to mitochondrial membrane hyperpolarization in winter wheat cultured cells, which in turn causes the increased ROS production.

[Induction of Hsp104 synthesis in Saccharomyces cerevisiae is inhibited by the petite mutation in the stationary growth phase].

The elevation of Hsp104 (heat shock protein) content under heat shock plays a key role in yeast (Saccharomyces cerevisiae) cells. Hsp104 synthesis is increased under heat stress in the stationary growth phase. As shown, the loss of mitochondrial DNA (petite mutation) inhibited the induction of the Hsp104 synthesis under heat stress (39 degrees C) during the transition to the stationary growth phase. Also, the petite mutation suppressed the activity of antioxidant enzymes in the same phase, which led to lower thermotolerance. At the same time, the mutation inhibited production of the reactive oxygen species and prevented cell death under heat shock in the logarithmic growth phase. The results of this study suggest that disruption of the mitochondrial functional state suppresses the expression level of yeast nuclear genes upon transitioning to the stationary growth phase.

[Influence of calcium and rhizobial infections (Rhizobium leguminosarum) on the dynamics of nitric oxide (NO) content in roots of etiolated pea (Pisum sativum L.) seedlings].

The effect of exogenous calcium (Ca2+) and rhizobial infections (Rhizobium leguminosarum bv viceae) on the dynamics of the level of nitric oxide (NO) was studied in cross cuts of roots of two-day-old etiolated pea seedlings (Pisum sativum L.) using a DAF-2DA fluorescent probe. Fluctuations of the NO level, indicating the presence of a rhythm in the generation of NO in roots, were observed during the incubation of seedlings in water, a CaCl2 solution, and with rhizobial infections. Exogenous factors (Ca2+ and two rhizobial stamms) change the time dynamics of the NO level in comparison with the control (water).

Protein Tyrosine Phosphatase CD45 As an Immunity Regulator and a Potential Effector of CAR-T therapy.

The leukocyte common antigen CD45 is a receptor tyrosine phosphatase and one of the most prevalent antigens found on the surface of blood cells. CD45 plays a crucial role in the initial stages of signal transmission from receptors of various immune cell types. Immunodeficiency, autoimmune disorders, and oncological diseases are frequently caused by gene expression disorders and imbalances in CD45 isoforms. Despite extensive research into the structure and functions of CD45, the molecular mechanisms behind its role in transmitting signals from T-cell receptors and chimeric antigen receptors remain not fully understood. It is of utmost importance to comprehend the structural features of CD45 and its function in regulating immune system cell activation to study oncological diseases and the impact of CD45 on lymphocytes and T cells modified by chimeric antigen receptors.

CAR-tropic extracellular vesicles carry tumor-associated antigens and modulate CAR T cell functionality.

Tumor-derived extracellular vesicles (EVs) are active contributors in metastasis and immunosuppression in tumor microenvironment. At least some of the EVs carry tumor surface molecules such as tumor-associated antigens (TAAs) and/or checkpoint inhibitors, and potentially could interact with T cells or CAR T cells. Upon contact with T cells, EVs could alter their phenotype and functions by triggering signaling through TCR or CAR reprogramming them to escape immune response. We hypothesize that EVs that possess TAA on the surface will probably interact with CAR T cells which can recognize and bind corresponding TAA. This interaction between EVs and CAR T cells may change the outcome of CAR T-based cancer immunotherapy since it should affect CAR T cells. Also, EVs could serve as adjuvants and antigenic components of antitumor vaccines. Herein, we isolated EVs from B cell precursor leukemia cell line (pre-B ALL) Nalm-6 and demonstrated that recognition and binding of CD19+EVs with CD19-CAR T cells strongly depends on the presence of CD19 antigen. CD19+EVs induce secretion of pro-inflammatory cytokines (IL-2 and IFN-y) and upregulated transcription of activation-related genes (IFNG, IFNGR1, FASLG, IL2) in CD19-CAR T cells. Tumor necrosis factor receptor superfamily (TNFRSF4 and TNFRSF9) and T-cell exhaustion markers (CTLA4, LAG3, TIM3 and PDCD1LG2) were also upregulated in CD19-CAR T cells after incubation with CD19+EVs. Long-term cultivation of CD19+ or PD-L1+EVs with CD19-CAR T cells led to increased terminal differentiation and functional exhaustion according to elevated expression of PD-1, TIGIT, CD57. In summary, our results suggest that chronic exposure of CD19-CAR T cells to CD19+EVs mediates activation and systemic exhaustion in antigen-specific manner, and this negative effect is accompanied by the impaired cytotoxic activity in vitro.

In Vitro Antiviral Activity of a New Indol-3-carboxylic Acid Derivative Against SARS-CoV-2.

The coronavirus disease (COVID-19) pandemic has brought into sharp relief the threat posed by coronaviruses and laid the foundation for a fundamental analysis of this viral family, as well as a search for effective anti-COVID drugs. Work is underway to update existent vaccines against COVID-19, and screening for low-molecular-weight anti-COVID drug candidates for outpatient medicine continues. The opportunities and ways to accelerate the development of antiviral drugs against other pathogens are being discussed in the context of preparing for the next pandemic. In 2012-2015, Tsyshkova et al. synthesized a group of water-soluble low-molecular-weight compounds exhibiting an antiviral activity, whose chemical structure was similar to that of arbidol. Among those, there were a number of water-soluble compounds based on 5-methoxyindole-3-carboxylic acid aminoalkyl esters. Only one member of this rather extensive group of compounds, dihydrochloride of 6-bromo-5-methoxy-1-methyl-2-(1-piperidinomethyl)-3-(2-diethylaminoethoxy) carbonylindole, exhibited a reliable antiviral effect against SARS-CoV-2 in vitro. At a concentration of 52.0 µM, this compound completely inhibited the replication of the SARS-CoV-2 virus with an infectious activity of 106 TCID50/mL. The concentration curves of the analyzed compound indicate the specificity of its action. Interferon-inducing activity, as well as suppression of syncytium formation induced by the spike protein (S-glycoprotein) of SARS-CoV-2 by 89%, were also revealed. In view of its synthetic accessibility - high activity (IC50 = 1.06 µg/mL) and high selectivity index (SI = 78.6) - this compound appears to meets the requirements for the development of antiviral drugs for COVID-19 prevention and treatment.