[The immunological characteristics and enzymatic activity of lymphocytes from the peripheral blood of the children presenting with pharyngeal tonsillar hypertrophy]. / Immunologicheskie pokazateli i énzimaticheskaia aktivnost' limfotsitov perifericheskoi krovi u detei s gipertrofiei glotochnoi mindaliny.

The objective of the present work was to study the immunological characteristics and activity of NAD(P)-dependent dehydrogenases in peripheral blood lymphocytes in the young children presenting with pharyngeal tonsil hypertrophy (PTH). A total of 57 children at the age from 1 to 3 years with PTH were available for the examination. The control group was comprised of 35 age-matched practically healthy children. The amount of CD3+, CD4+, CD8+, CD16+/56-, and CD19+ cells in the peripheral blood was determined with the use of the cytoflowmetric technique. The activity of NAD(P)-dependent dehydrogenases in the peripheral blood lymphocytes was quantified by the method of A.A. Savchenko and co-workers ([14]. Serum IgA, IgM, and IgG levels were measured as described by G. Mancini and co-workers [12], and the levels of the circulating immune complexes (CIC) by the method of Haskova and co-workers [13]. The children presenting with pharyngeal tonsil hypertrophy were found to undergo changes in the immune-phenotypic spectrum of peripheral bloods lymphocytes, the decrease of serum IgA concentration, and the increase in the serum CIC level. The activity of riboso-5-phpsphate- and NADH-dependent reactions of the macromolecular synthesis was increased whereas the role of the malate-aspartate shunt in the cellular energy metabolism and activity of glycolysis decreased. On the contrary, the substrate flow in the tricarbonic acid cycle was rather high while glutathione reductase activity was low. The present study has shown that the children presenting with pharyngeal tonsil hypertrophy underwent changes in the immune and phenotypic spectrum of peripheral bloods lymphocytes, the decrease of serum IgA concentration, and the increase in the serum CIC level.

[Changes of the Immunophenotypic Spectrum and the Enzymatic Profile of Peripheral Blood Lymphocytes in Infants with Hypertrophy of the Pharyngeal Tonsil].

OBJECTIVE: to study immunophenotype and NAD- and NAD(P)-dependent dehydrogenase of blood lymphocytes activity indicators in children with hypertrophy of the pharyngeal tonsils (HPT). METHODS: 57 children aged 1-3 years with HPT were examined. The focus group included 35 healthy children of the similar age. The number of CD3⁺-, CD4⁺-, CD8⁺-, CD16⁺/56⁺-, CD19⁺-cells in the blood was determined by flow cytometry. The activity of NAD(P)-dependent dehydrogenase was studied by the method of A. Savchenko and coauth. (1989). RESULTS: The changes of immunophenotypic spectrum of peripheral blood lymphocytes in infants with HPT have been revealed. The increase of ribose-5-phosphate and NADN-dependent reactions of macromolecular synthesis, the reduction of malataspartat shunt role in cell energy, the reduction of anaerobic lactate dehydrogenase reaction, the compensatory increase in the glycerol-3-phosphate dehydrogenase activity, the high substrate flow of the citric acid cycle and the reduced level of glutathione have been fixed. The correlation analysis has showed increase in the number of correlations between indicators of investigated oxidoreductase activity in blood lymphocytes in children with HPT and the high level of correlation between the metabolic reactions of the mitochondrial compartment. CONCLUSION: the change of immunophenotype, enzymatic activity, correlation pattern of connection between intracellular enzymes of peripheral blood lymphocytes have been revealed in children aged 1-3 years with HPT.

[The role of environmental factors in the development of pathology of the lymphadenoid pharyngeal ring in children].

The authors present the analysis of the prevalence and structure of pathology of the lymphadenoid pharyngeal ring in children living in the districts of the city of Krasnoyarsk differing in terms of atmospheric pollution. It is shown that that pharyngeal adenoid hypertrophy occurs significantly more frequently in the children from the districts exposed to an extremely high level of atmospheric pollution than in those residing under more favourable environmental conditions.

[Immunomodulating effect of protein fractions isolated from bifidobacteria]. / Immunomoduliruiushchii éffect belkovykh fraktsii, vydelennykh iz bifidobakterii.

The screening of the immunomodulating activity (IMA) of different protein fractions isolated from bifidobacteria was carried out and the capacity of these fractions for changing the proliferative activity of immunocompetent cells was evaluated. Soluble proteins were extracted from lyophilized and sonicated bacterial mass of B. bifidum strain 1 in Na2HPO4 (pH 8) in a water bath at 65 degrees C for 30 minutes. After the formation and removal of nucleic acid sediment the resulting supernatant fluid was dialyzed, its adsorption spectra were analyzed and the fluid was fractionated in a specially proposed device for preparative electrophoresis. Protein fractions were tested for IMA on spleen cells of CBA mice in the reaction of lymphocyte blast-transformation by the level of the inclusion 3H thymidine. The analysis of IMA of protein fractions revealed that their high-molecular components produced a pronounced dose-dependent effect on the proliferative activity of spleen cells. The fractions containing low-molecular components were either inactive (fraction 4) or active only in the maximum dose (fraction 5).

[The unknown functions of heterologous antigens of different origins]. / Neizvestnye funktsii geterogennykh antigenov raznogo proiskhozhdeniia.

After a single injection of 125I-labeled erythrocytic antigen to mice, the protein determinants of this antigen were found in nuclear RNP of B lymphocytes of the immune animals. On day 2 after the injection of Shigella sonnei phase I LPS (also containing protein) to mice hybridization between total RNA isolated from B lymphocytes and 32P-labeled plasmid pSS 120, controlling the synthesis of the S. sonnei phase I antigen, was observed. On day 1 after immunization no hybridization between cell RNA and plasmid pSS 120 occurred. Besides, no hybridization of plasmid DNA with the RNA of nonimmune animals or with RNA, obtained on day 3 after the injection of Salmonella kentucky LPS to mice, was observed. The signal indicating the occurrence of hybridization between the RNA of immune B lymphocytes and plasmid pSS 120 appeared on day 2 and increased on days 3 and 5 after the injection of the antigen. This was seemingly indicative of the accumulation of new nucleotide sequences, complementary to the gene controlling the synthesis of the heterologous antigen which caused immune response, in immune B lymphocytes.

[Lipopolysaccharide changes in smooth-type avirulent Shigella in comparison with initial virulent strains]. / Izmeneniia lipopolisakharidov u avirulentnykh shigell gladkogo tipa po sravneniiu s iskhodnymi virulentnymi shtammami.

The comparative study of the lipopolysaccharides (LPS) of virulent and avirulent strains of S. sonnei, phase I (smooth colonies), has been made. Electrophoresis of LPS and subsequent densitometry of electrophoregrams have revealed the increase of the fraction of long 0-chains with a considerable number of recurring elements in 2 out of 3 LPS preparations obtained from avirulent shigellae. In mice immunized with these LPS preparations a considerably greater number of antibody-producing cells can be detected in Jerne's test on sheep red blood cells (SRBC) sensitized with the LPS of a virulent strain than on those sensitized with the above LPS preparations. Long 0-specific chains supposedly inhibit the fixation of individual complement components on the corresponding sensitized SRBC. The LPS of the third avirulent strain of S. sonnei, phase I, with transposon integrated into its genome, which has led to the formation of the avirulent variant of a previously virulent strain, seems to contain fine structural differences from the initial virulent strain. The immunogenicity of the LPS of this avirulent strain is greatly (3-4 times) decreased, which is manifested by the number of antibody-producing cells detected in Jerne's test on SRBC sensitized with LPS preparations obtained from these two strains.

[Changes in antigenic properties of lipopolysaccharides of Shigella sonnei phase I having lost the virulence due to transposon integration into the invasiveness plasmid PSS120]. / Izmenenie antigennykh svoistv lipopolisakharida shigell Zonne I fazy, utrativshikh virulentnost' v sviazi s integratsiei transpozona v plazmidu invazivnosti PSS120.

A nonvirulent strain of Shigella sonnei phase I has been obtained by integration of the transposon Tn5 into the invasiveness plasmid pSS120 in the virulent strain and designated NR18. The presence of the plasmid pSS120 in both strains results in the similar morphology and bacterial ability to agglutinate in the presence of antiserum to Shigella sonnei phase I antigen. The lipopolysaccharide preparations from the virulent and nonvirulent strains give the similar reactions with the antiserum in the reaction of hemagglutination. However, in the reaction of passive local hemolysis in the gel (Jerne reaction) the significant difference is revealed in the immunogenicity of the preparations, with the preparations from the virulent strain being 4-5 fold more immunogenic. In crossreaction, the antibodies secreted by the mouse spleen cells immunized by LPS from the virulent strain show a weak reaction with the ram erythrocytes sensitized by the LPS of the nonvirulent strain. Thus, the biological changes in the LPS of the nonvirulent strains that are, evidently, the consequence of the structural changes, are identified only by the most sensitive immunological techniques.

[Protein synthesizing structures and protein synthesis in mouse lymphocytes]. / Beloksinteziruiushchie struktury i sintez belka v limfotsitakh myshi.

The properties of cytoplasmic mRNP particles from spleen lymphocytes (sedimentation coefficients and buoyant density) and the kinetics of incorporation of impulse amino acid label at definite labelling periods (5, 15, 30, 60 sec) were studied. After 5 or 15 sec labelling periods the labelled amino acid was detected in the polysomes (sedimentation coefficient 130-240S, buoyant density 1.55 g/cm3) and in small-sized particles (sedimentation coefficient 80S and less, buoyant density 1.34, 1.50 and 1.60 g/cm3). After 30 sec of labelling the label is incorporated into the medium-sized particles (sedimentation coefficient 80-120 S, buoyant density 1.47 g/cm3); no increase of the label incorporation into the small-sized particles is observed. After 60 sec the label content in all the three sucrose gradient zones is increased. The observed succession of the amino acid label incorporation first into the small-sized particles and then into the large-sized ones having a different buoyant density and the data from impulse uridyl labelling suggest that in the lymphocytes the translation occurs already in the complexes of informosomes with single ribosomes and continues in the course of their conversion to polysomes. The short-time labelling of the cells with an impulse amino acid label allows to determine the time of the polypeptide chain translation from the time of polysome saturation with the label.

[Presence of antigenic determinants in mRNA-containing RNP particles of immune B-lymphocytes]. / Prisutstvie determinant antigena v mRNK-soderzhashchikh pibonukleoproteidnykh chastitsakh immunnykh B-limfotsitov.

An iodinated erythrocyte antigen isolated from sheep erythrocytes (125I-EAG) was injected to the BALB/c or CBA mice in a single dose of 20-25 mkg as for protein. The antigen determinants were detected by incorporation of a radioactive label into spleen B-lymphocytes on the 1sh-5th post-immunization days. It was shown that the T-cells contain no label and that 2/3 of the radioactivity of B-lymphocyte suspension of the 125I-EAG-immunized mice are detected in the cell nuclei within mRNP. During electrophoresis of the particles in 2,5% polyacrylamide gel nuclear mRNP isolated from B-lymphocytes of mice immunized with cold EAG are adsorbed by an immunoadsorbent containing antibodies against sheep erythrocytes. Fractionation of 125I-cytoplasmic extract obtained from 125I-EAG-immunized mouse B-lymphocytes in sucrose gradient revealed that the radioactive label was detected in the same two regions of the sucrose gradient, which is occupied by light RNP complexes containing mRNA and polyribosomes. Under experimental conditions allowing to detect the transitions of mRNP complexes into polyribosomes, a shift of the 125I-labelled material (probably together with the RNP particles) towards polyribosomes was observed. Thus, the antigen determinants are detected as part of nuclear and cytoplasmic mRNP of B-lymphocytes during the whole cycle of the primary immune response and are detected within the composition of cytoplasmic mRNP during the immunoglobulin polypeptide chain synthesis.

[Ribonucleoprotein mRNA-containing cytoplasmic particles from mouse lymphocytes before and after injection of antigen]. / Ribonukleoproteidnye mRNK-soderzhashchie chatitsy tsitoplazmy limfotsitov myshei do i posle vvedeniia antigena.

In mouse spleen lymphocytes mRNA was discovered not only in poliribosomes but also in lighter RNP complexes having sedimentation coefficients of 80--120S and buyoant density of 1,40--1,52 g/cm3. The incorporation of 14C-pulse aminoacid lable has shown that the most active polypeptide synthesis takes place in light RNP-complexes and not in polyribosomes. The pattern and the contents of labeled mRNA-containing particules in spleen cytoplasm changed within different periods after the antigen injection. The greatest contents of these particles have been discovered on the second day after the antign injection, 3H- and 14C-radioactivity of RNP-complexes with higher sedimentation coefficients, and that of polyribosomes was increased; the optical density of polyribosomes peak was also increased. On the 4th day after the antigen injection contents of light RNP-complexes with higher sedimentation coefficients were decreased. On the 5th day the decrease of RNP-complexes contents was more pronounced. The changes of radioactive RNP-complexes contents in spleen cytoplasm at different periods of the immune response, which can be discovered during determined period of labling, are probably due to acceleration and then to retardation of the transition of light RNP complexes into polyribosomes. It, probably, reflects the existens in spleen lymphocytes mechanisms of translation regulation, irrespective of mRNA stability which are realized during immune reaction. Periods of the translation increasing or inhibiting in immune lymphocytes don't correspond to periods of increasing or decreasing of number of antibody synthesising cells in spleen.

[Erythrocyte antigen: isolation, biochemical and immunologic properties]. / Eritrotsitarnyi antigen: vydelenie, biokhimicheskie i immunologicheskie svoistva.

The authors suggest a simple method of obtaining erythrocytic antigen in considerable amounts. This antigen is of stromal origin, contains from 10 to 20% protein, and is relatively homogenous. With the concentration of from 1 to 50 microgram by protein the preparation represents a transparent solution; with greater concentrations the antigen is white, turbid, but is well dissolved and convenient for administration to the animals. In case of a single administration without any adjuvants the antigen is highly immunogenic in low doses by protein. To the optimal immunizing dose of erythrocytes (5 X 10(8)) correspond about 100 microgram of the antigen by protein. The primary response to the antigen is similar to the response to sheep red blood cells (SRBC). It is exceedingly effective for the formation of immunological memory. The level of secondary responses in the adoptive transfer to all the EAG doses always exceeded the secondary response to SRBC. By adding EAG into agar during the local hemolysis in gel test determined the avidity of the antibodies synthesized at various periods of the immune response to SRBC.

[Isolation of preparative amounts of polyribosomes from normal rabbit and guinea pig spleen]. / V'delenie poliribosom iz selezenki normal'nogo krolika i morskikh svinok v preparativn'kh kolichestvakh

Preparative amounts of polyribosomes were isolated from normal rabbit and guinea pig spleen; up to 40 optical units of the polyribosome preparation could be obtained by centrifugation in a Spinco L-2B centrifuge with SW-27 rotor. The amount of polyribosomes isolated from spleens of immune animals was 2-3 higher than that isolated from normal animal spleens. Concentration of polyribosomal preparations by lyophylization and the storage of dried preparations do not alter the sedimentation properties of the polyribosomes. The distribution pattern of normal rabbit spleen polyribosomes in a linear sucrose gradient and the sedimentation constants of the polyribosome peaks are in good agreement with data reported by some other authors for plasmocytome polyribosomes. Using electrophoresis in agarose-polyacrylamide gel the radioactive proteins synthesized in the cell culture of normal rabbit spleen it was shown that in normal spleen the average amount of globulins makes up to 35% of total protein synthesis, as reported by some authors.

[Use of proteins, covalently bound to agar gel, for specific sorption in electrophoresis]. / Primenenie belkov, kovalentno prisoedinennykh k geliu agara, dlia spetsificheskoi sorptsii v usloviiakh elektroforeza

A method is developed for electrophoretic fractionation of serum proteins in agar gel with simultaneous specific sorption of gamma-globulin fraction, migrating during the electrophoresis through specific immunosorbents incorporated into agar gel. The reaction was carried out in a special apparatus designed for the electrophoresis in two perpendicular directions. The sorbent was separated from impurities by passing the current in the first direction. The unspecific sorbent did not alter the electrophoretic mobility of migrating proteins, but the specific sorbents immobilized completely both antibodies and antigens. The antibodies were shown to bind repeatedly to the immunoglobulins, which had been linked to the sorbent. The method facilitated distinctly the estimation of antigens and antibodies in complicated mixtures. By analogy with affinity chromatography a term "affinity electrophoresis" was used for designation of the method described.