[High Heterogeneity of Virus-Neutralizing and RBD-Binding Activities of COVID-19 Convalescent Sera].

The parameters of the humoral response are an important immunological characteristic of donors who recovered from COVID-19 and vaccinated individuals. Analysis of the level of virus-binding antibodies has become widespread. The most accurate predictor of effective immune protection against symptomatic SARS-CoV-2 infection is the activity of virus-neutralizing antibodies. We determined virus-neutralizing activities in plasma samples of individuals (n = 111) who had COVID-19 from April to September 2020. Three independent methods were used: conventional with live virus, with virus-like particles pseudotyped with spike protein, and a surrogate virus-neutralization test (cVNT, pVNT and sVNT, respectively). For comparison, the levels of IgG, IgA and IgM antibodies against the receptor-binding domain of the SARS-CoV-2 spike protein were also evaluated. The levels of virus-binding as well as virus-neutralizing antibodies in cVNT and pVNT showed high heterogeneity. A comparison of cVNT and pVNT results showed a high correlation, sVNT results also correlated well with both cVNt and pVNT. To the greatest extent, the level of IgG antibodies correlated with the results of cVNT, pVNT and sVNT. These results can be used in the selection of plasmas that are best suited for transfusion and treatment of acute COVID-19. In addition, data on the virus-neutralizing activity of plasma are important for the selection of potential donors, for the isolation of SARS-CoV-2-specific B-lymphocytes, in order to further generate monoclonal virus-neutralizing antibodies.

High Heterogeneity of Virus-Neutralizing and RBD-Binding Activities of COVID-19 Convalescent Sera.

The parameters of the humoral response are an important immunological characteristic of donors who recovered from COVID-19 and vaccinated individuals. Analysis of the level of virus-binding antibodies has become widespread. The most accurate predictor of effective immune protection against symptomatic SARS-CoV-2 infection is the activity of virus-neutralizing antibodies. We determined virus-neutralizing activities in plasma samples of individuals (n = 111) who had COVID-19 from April to September 2020. Three independent methods were used: conventional with live virus, with virus-like particles pseudotyped with spike protein, and a surrogate virus-neutralization test (cVNT, pVNT and sVNT, respectively). For comparison, the levels of IgG, IgA and IgM antibodies against the receptor-binding domain of the SARS-CoV-2 spike protein were also evaluated. The levels of virus-binding as well as virus-neutralizing antibodies in cVNT and pVNT showed high heterogeneity. A comparison of cVNT and pVNT results showed a high correlation, sVNT results also correlated well with both cVNT and pVNT. To the greatest extent, the level of IgG antibodies correlated with the results of cVNT, pVNT and sVNT. These results can be used in the selection of plasmas that are best suited for transfusion and treatment of acute COVID-19. In addition, data on the virus-neutralizing activity of plasma are important for the selection of potential donors, for the isolation of SARS-CoV-2-specific B-lymphocytes, in order to further generate monoclonal virus-neutralizing antibodies.

CD44-Associated Tn Antigen as a New Biomarker of Tumor Cells with Aberrant Glycosylation.

Tn antigen is a tumor-associated antigen that appears on cancer cells as a result of aberrant O-glycosylation. The most studied form of Tn antigen is found in mucins, in particular, in mucin 1 (MUC1). Antibodies against this form of Tn antigen are used to diagnose tumors, as well as to generate T-killers with a chimeric receptor. Some carcinomas do not carry MUC1 and antibodies of a different specificity are required to detect Tn antigen on these cells. In our work, we searched for anti-Tn antibodies without preliminary assumptions about the proteins that may be carriers of the Tn antigen. For this purpose, we obtained several pairs of isogenic cell lines with the wild type and knockout of the Cosmc gene, which is essential for correct protein O-glycosylation. Using the created lines as immunogens, we generated a monoclonal antibody AKC3, which reacted with the Cosmc-deficient A549 lung adenocarcinoma cells and did not bind to the wild-type cells. Using mass spectrometry, as well as co-immunoprecipitation, it was shown that the AKC3 antibody recognized the Tn antigen in the context of CD44 protein - a protein important for tumor growth. The AKC3 antibody can be used for tumor diagnosis, and to generate T cells with a chimeric receptor for treatment of tumors that do not express mucins.

[Identification of the Molecular Partners of Lymphocyte Phosphatase-Associated Phosphoprotein (LPAP) That Are Involved in Human Lymphocyte Activation].

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a small transmembrane protein that is found in lymphocytes and is tightly associated with the phosphatase CD45. The function of LPAP is still unknown. Studies of the LPAP interactome may reveal new details of how C45 and lymphocyte signaling in general are regulated. LPAP binding partners were sought using coimmunoprecipitation coupled with mass spectrometry, stabilization of protein complexes with chemical crosslinkers, and Blue Native electrophoresis. In addition to CD45, several proteins were identified as LPAP partners, including CD71, CD98, cytoskeletal proteins, the amino acid transporter SLC1A4, and the cell signaling component HS1. It was confirmed that more than 70% of LPAP molecules were bound with CD45 in a 1 : 1 complex. The effect of CD45 on LPAP was studied in CEM and Jurkat cells with a CD45 knockout. The LPAP levels in the cells were 10% of the level in wild-type cells. In the absence of CD45, LPAP phosphorylation at Ser-153 and Ser-163 was not affected, whereas phosphorylation at Ser-99 and Ser-172 decreased significantly. Based on the results, CD45 was assumed to play a role in regulating LPAP expression and phosphorylation status.

[HIV Restriction Factors and Their Ambiguous Role during Infection].

Currently, more than 37 million individuals worldwide are infected with the human immunodeficiency virus (HIV). Antiretroviral therapy may control the viral infection but is incapable of eradicating it. It is important to understand how cells respond to HIV-1 infection and what cellular factors are involved in this process to develop novel classes of antiviral drugs. This review summarizes the current understanding of the HIV restriction mechanism. We discuss the ambiguous role of HIV restriction factors in viral infection and counteraction mediated by HIV-1 accessory proteins.

Lymphocyte Phosphatase-Associated Phosphoprotein Is a Substrate of Protein Kinase CK2.

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a molecular partner of CD45 phosphatase that plays a key role in the regulation of antigen-specific activation of lymphocytes. The functions of LPAP still remain unknown. We believe that studying LPAP phosphorylation pathways could shed light on its functions. In this work, we studied the phosphorylation of LPAP ectopically expressed in non-lymphoid cells in order to determine the effect of LPAP interaction partners on its phosphorylation. We found that phosphorylation at Ser153 and Ser163 in non-hematopoietic HEK293 cells was conserved, while phosphorylation at Ser99 and Ser172 was almost absent. The pattern of LPAP phosphorylation in K562 erythroid and U937 myeloid cells expressing endogenous CD45 protein was similar to that observed in T and B lymphocytes. We demonstrated for the first time that LPAP is a substrate for protein kinase CK2 that phosphorylates it at Ser153, presumably ensuring LPAP resistance to degradation.

[A case of solid pseudopapillary tumor of the pancreas: features of the course, difficulty in diagnosis]. / Nabliudenie solidno-psevdopapilliarnoi opukholi podzheludochnoi zhelezy: osobennosti techeniia, slozhnosti v diagnostike.

Solid pseudopapillary tumor is a low-grade malignant neoplasm of the pancreas. The clinical manifestations are variable, ranging from an asymptomatic course to cases with severe symptoms that dramatically impair the patients' status. The paper describes the rare case of a solid pseudopapillary tumor in a 34-year-old woman, which was accompanied by difficulties in the interpretation of clinical data and morphological patterns.

[Next-Generation Techniques for Discovering Human Monoclonal Antibodies].

Monoclonal antibodies have found wide applications in the treatment of cancer, as well as of autoimmune, infectious, and other diseases. Several dozen new antibodies are currently undergoing different stages of clinical trials, and some of them will soon be added to the list of immunotherapeutic drugs. Most of these antibodies have been generated using hybridoma technology or a phage display. In recent years, new methods of obtaining human monoclonal antibodies have been actively developing. These methods rely on sequencing immunoglobulin genes from B lymphocytes, as well as on the creation of antibody-secreting stable B-cell lines. The term next-generation antibody-discovery platforms has already been established in the literature to refer to these approaches. Our review focuses on describing the results obtained by these methods.

Next-Generation Techniques for Discovering Human Monoclonal Antibodies.

Monoclonal antibodies have found wide applications in the treatment of cancer, as well as of autoimmune, infectious, and other diseases. Several dozen new antibodies are currently undergoing different stages of clinical trials, and some of them will soon be added to the list of immunotherapeutic drugs. Most of these antibodies have been generated using hybridoma technology or a phage display. In recent years, new methods of obtaining human monoclonal antibodies have been actively developing. These methods rely on sequencing immunoglobulin genes from B lymphocytes, as well as on the creation of antibody-secreting stable B-cell lines. The term next-generation antibody-discovery platforms has already been established in the literature to refer to these approaches. Our review focuses on describing the results obtained by these methods.

Epitope mapping of lymphocyte phosphatase-associated phosphoprotein.

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a transmembrane protein with unknown function. The available data on its close association with phosphatase CD45 and its phosphorylation depending on cell activation suggest that LPAP can play a significant role in the antigenic stimulation of lymphocytes. We have localized three antigenic epitopes of the LPAP molecule that can be detected using monoclonal antibodies prepared earlier. Experiments on reactions of antibodies with point mutants and shortened forms of the LPAP protein revealed regions of the amino acid sequence that correspond to the epitopes recognized by the antibodies.

Profiling of the CD4 receptor complex proteins.

Intermolecular complexes produced by the CD4 molecule were studied. To preserve the integrity of weak protein-protein interactions of the CD4 antigen, cells were lysed in a mild nonionic detergent Brij97. Protein constituents of the complex were identified by our previously proposed fluorescence immunoprecipitation assay with subsequent mass spectrometry. In total, 26 proteins associated with CD4 were identified on CEM cells. The CD4 complex included the following major components: tyrosine phosphatase CD45, transferrin receptor CD71, tyrosine kinase Lck, and a lymphocyte phosphatase-associated phosphoprotein LPAP. The CD4 complex also contained some components of cytoskeleton and heat shock proteins. The association between CD4, CD71, and CD45 molecules was confirmed by immunoblotting. The CD4 complexes were not detected on the U937 myeloid cells lacking Lck and LPAP. We attempted to quantitatively characterize the CD4 complex composition.

Peculiarities of active avoidance conditioning in rats with various forms of inherited arterial hypertension.

Peculiarities of active avoidance conditioning were studied in NISAG rats (hereditary stress-induced arterial hypertension) and spontaneously hypertensive SHR rats. Conditioning was successful in 90% normotensive Wistar rats and in only 9.1% NISAG rats. Hypertensive SHR rats were intermediate between Wistar and NISAG rats by their learning capacity (66.7%). Our results suggest that differences in learning capacity of hypertensive rats are determined by genetic characteristics of animal behavior and emotional state, rather than blood pressure elevation.

Complement-mediated lysis by anti-CD20 mAb correlates with segregation into lipid rafts.

Despite the clinical success of anti-CD20 monoclonal antibody (mAb) in the treatment of lymphoma, there remains considerable uncertainty about its mechanism of action. Here we show that the ability of mAbs to translocate CD20 into low-density, detergent-insoluble membrane rafts appears to control how effectively they mediate complement lysis of lymphoma cells. In vitro studies using a panel of anti-B-cell mAbs revealed that the anti-CD20 mAbs, with one exception (B1), are unusually effective at recruiting human complement. Differences in complement recruitment could not be explained by the level of mAb binding or isotype but did correlate with the redistribution of CD20 in the cell membrane following mAb ligation. Membrane fractionation confirmed that B1, unlike 1F5 and rituximab, was unable to translocate CD20 into lipid rafts. In addition, we were able to drive B1 and a range of other anti-B-cell mAbs into a detergent-insoluble fraction of the cell by hyper-cross-linking with an F(ab')(2) anti-Ig Ab, a treatment that also conferred the ability to activate lytic complement. Thus, we have shown that an important mAb effector function appears to be controlled by movement of the target molecule into membrane rafts, either because a raft location favors complement activation by mAbs or because rafts are more sensitive to complement penetration.

[Soft-tissue contour plastic repair of the maxillofacial area using a biologically compatible polyacrylamide gel]. / Konturnaia plastika miagkikh tkanei cheliustno-litsevoi oblasti biologicheski sovmestimym poliakrilamidnym gelem.

Polyacrylamide hydrogel was used in contour plastic repair of maxillofacial soft tissues in 161 female patients. The remote results (up to 2 years after the manipulation) demonstrate the advantages of this method over traditional surgery. Biogel preserves the natural appearance of soft tissues of the face; the clinical effect is stable.

[The surgical treatment of mandibular prognathism after the elimination of congenital clefts of the upper lip and palate]. / Khirurgicheskoe lechenie prognatii nizhnei cheliusti posle ustraneniia vrozhdennykh rasshchelin verkhnei guby i neba.

In 4.47% of observation in patients after liquidation of the cleft lip and the palate the lower prognathia is diagnosed. From 77 patients being operated for lower prognathia such kind of patients were 7. The subsequence of the treatment measures is the following: correction of the upper jaw on the basis of compactostetomy and it's subsequent widening with orthodontic devices. In case of impossibility of sufficient widening of the dental arch the removable dental prosthesis is made for application on the upper jaw with doubling dental row accounting achievement of the desired bite. At the second stage the osteotomy of the upper jaw brunch and sagittal removement of the central section back are carried out up to achievement of the neutral bite. In dependence on jaw sagittal removement degree operative treatment may be performed by the intravial (I degree) or extravial method (II-III progemy degrees). Quite satisfactory remote results are presented. The paper is illustrated by 4 figures. The list of literature is given.

[Immunologic characterization of patients with insulin-dependent diabetes mellitus with varying duration of disease]. / Immunologicheskie kharakteristiki bol'nykh insulinzavisimym sakharnym diabetom s razlichnoi dlitel'nost'iu zavolevaniia.

The paper presents data on the cellular and humoral immunity in different periods of insulin-dependent diabetes mellitus (with the disease standing of 0.5 +/- 0.4 years, group A; 3 +/- 1.8 years, group B; and 15 +/- 4 years, group C). Group A patients presented with the immunity system activation: increased counts of T cells, B lymphocytes, T helpers and T inductors, increased share of active T cells (that is, DR positive ones), elevated content of IgM, IgG, IgA (214 +/- 51 mg%, 1200 +/- 124 mg%, 250 +/- 34 mg%, respectively) as against the reference group (156 +/- 74, 914 +/- 387, 189 +/- 49 mg%, respectively) (p < 0.01). In group B patients, who suffered a longer disease, the immunity parameters were within the normal range, and in group C patients, in whom the disease standing was the longest, these shifts were contrary-wise as against those in group A, that is, T and B cell counts were lowered, as were the counts of T-helpers-inductors, Ig levels, and the phagocytosis index was 65 +/- 5 vs. 85 +/- 10% in the controls (p < 0.05), the phagocytosis level being 4 +/- 2 vs. 10 +/- 2 in the controls (p < 0.05). The authors analyze the association of the HLA system characteristics with the immunity shifts. Patients with the HLA A9, B8, B15, B18, DR3, DR4 presented with significant shifts in the immunity status as against those with the HLA A1, A10, B5, B12, B16, B27, DR5, DR7.(ABSTRACT TRUNCATED AT 250 WORDS)

[The subpopulation structure of human T-lymphocytes studied by using a monoclonal antibody (MCA) to CD27]. / Izuchenie subpopuliatsionnoi struktury T-limfotsitov cheloveka s pomoshch'iu monoklonal'nogo antitela (MKAT) CD27.

The obtained murine mAb LT27 (IgG2a) assigned to the cluster of differentiation CD27 was used to study the distribution of antigen CD27 among human lymphocytes scbpopulations in normal state and immunopathology. In normal donors the antigen CD27 was found to be expressed most frequently on CD4+ cells (90 +/- 8% of which coexpressed antigen CD27) and to the lesser extent on- CD8+ cells (only 77 +/- 28% of CD8+ cells carried antigen CD27). 79 +/- 12% of double negative lymphocytes (CD3+CD4-CD8-) expressed antigen CD27. In patients with hypogammaglobulinemia the proportion of CD4+CD27+/CD4+ and CD8+CD27+/CD8+ was significantly reduced to 80 +/- 11% (p < 0.01) and 45 +/- 19% (p < 0.001), respectively. The ratio CD4+CD27+/CD4+ varied insignificantly with the increase of CD4+ population, but the increase of the CD8+ population was accompanied by the definite tendency to a decrease of the ratio CD8+CD27+/CD8+. The distribution of CD27 antigen inside CD4+ and CD8+ subpopulations was found to be different from the distribution of CD29 and CD45RA antigens.

Two-colour flow cytometry study of lymphocyte subpopulations in patients with primary immunodeficiencies.

Immunofluorescent flow cytometric examination of one hundred and eighty-five children with different primary immunodeficiency syndromes and sixty-nine control patients revealed twenty-six cases with a bimodal distribution of antigens CD5 and CD7. Such abnormalities were most frequently found in patients with total antibody deficiency, namely those with common variable hypogammaglobulinaemia (10/24 patients) and congenital agammaglobulinaemia with lack of B cells (10/40), but were never seen in normal controls. Two-colour flow immunofluorescence demonstrated that antigen CD4 was expressed only on intensely fluorescent CD5+ cells, irrespective of the immunodeficiency state. Antigen CD4 was detected on cells with both high and low expression of antigen CD7, but a small percentage (2%-5%) of CD4+ lymphocytes did not belong to the CD7+ population. Antigen CD8 was found equally on intensely and weakly fluorescent CD5+ and CD7+ cells. In some immunodeficient patients suffering from ataxia-telangiectasia (12/36) and in some with Wiskott-Aldrich syndrome (2/6) there was a significant excess (greater than 20%) of CD7+ over CD5+ cells. In these patients a considerable number of the CD8+ cells were not part of the CD5+ population, but were always part of the CD7+ population. Cell populations with the phenotype CD5-, CD7+ consisted mainly of lymphocytes showing weak expression of antigen CD8.

[Study of human lymphocyte subpopulations using a panel of monoclonal antibodies]. / Issledovanie subpopuliatsionnogo sostava limfotsitov cheloveka s pomoshch'iu paneli monoklonal'nykh antitel.

Characteristics of newly produced monoclonal antibodies (MCA) against differentiation antigens of human T- and B-lymphocytes and monocytes have been presented. The features of new MCA were compared with those of MCA with predefined specificity. According to two-color immunofluorescence and radioimmunoprecipitation data some of the former MCA were assigned to established clusters and the other were considered to recognize previously noncharacterized antigens. Potential value of MCA against different antigens in immunological diagnosis has been discussed.

[The expression of an antigen interacting with monoclonal antibody IB4 on the surface membranes of normal human leukocytes and in lymphoproliferative diseases]. / Izuchenie ékspressii antigena, vzaimodeistvuiushchego s monoklonal'nym antitelom IB4, na poverkhnostnykh membranakh leikotsitov cheloveka v norme i pri limfoproliferativnykh zabolevaniiakh.

Monoclonal antibody (MAb) 1B4 of IgG3 isotype reacting with 55 +/- 10% peripheral blood lymphocytes was obtained. MAb 1B4 is bound with 100% of B and NK cells and 50-70% of T cells in blood. The most part of CD8+ cells (60-100%) and approximately 50% of CD4+ cells belong to 1B4+ population. T-ALL cells do not express 1B4 antigen. Analysis of reactivity of MAb 1B4 with transformed malignant cells has shown that 1B4 antigen appeared in the process of differentiation of B lymphocytes at the state of pre-B cells and disappeared with activation of B lymphocytes and transition to plasma cells.