Plasticity of Human THP-1 Cell Phagocytic Activity during Macrophagic Differentiation.

Studies of the role of macrophages in phagocytosis are of great theoretical and practical importance for understanding how these cells are involved in the organism's defense response and in the development of various pathologies. Here we investigated phagocytic plasticity of THP-1 (acute monocytic human leukemia) cells at different stages (days 1, 3, and 7) of phorbol ester (PMA)-induced macrophage differentiation. Analysis of cytokine profiles showed that PMA at a concentration of 100 nM induced development of the proinflammatory macrophage population. The functional activity of macrophages was assessed on days 3 and 7 of differentiation using unlabeled latex beads and latex beads conjugated with ligands (gelatin, mannan, and IgG Fc fragment) that bind to the corresponding specific receptors. The general phagocytic activity increased significantly (1.5-2.0-fold) in the course of differentiation; phagocytosis occurred mostly through the Fc receptors, as shown previously for M1 macrophages. On day 7, the levels of phagocytosis of gelatin- and Fc-covered beads were high; however, the intensity of ingestion of mannan-conjugated beads via mannose receptors increased 2.5-3.0-fold as well, which indicated formation of cells with an alternative phenotype similar to that of M2 macrophages. Thus, the type and the plasticity of phagocytic activity at certain stages of macrophage differentiation can be associated with the formation of functionally mature morphological phenotype. This allows macrophages to exhibit their phagocytic potential in response to specific ligands. These data are of fundamental importance and can be used to develop therapeutic methods for correcting the M1/M2 macrophage ratio in an organism.

Structural and functional state of heart left ventricle depending on polymorphism rs966221 phosphodiesterase 4D gene in emergency workers of the Chornobyl NPP suffering from coronary heart disease. / STRUKTURNO FUNKTsIONAL'NYY̆ STAN MIOKARDA LIVOGO ShLUNOChKA SERTsIa V UChASNYKIV LIKVIDATsIÏ NASLIDKIV AVARIÏ NA ChORNOBYL'S'KIY̆ AES Z IShEMIChNOIu KhVOROBOIu SERTsIa ZALEZhNO VID POLIMORFIZMU rs966221 GENA FOSFODIESTERAZY 4D.

OBJECTIVE: This study consisted in examination the features of structural and functional state of the cardiovascular system in emergency workers (EW) of the Chernobyl nuclear power plant (ChNPP) who suffered from coronary heart disease (CHD) and having different genotypes due to polymorphism rs966221 phosphodiesterase 4D (PDE4D) gene. MATERIALS AND METHODS: The study involved 121 EW and 63 non irradiated patients with CHD. Standardized survey included echo doppler cardiography (EchoCG) that was done by Diagnostic Ultrasound System DS N3 (Mindray). Polymorphism rs966221 PDE4D determined by polymerase chain reaction followed by restriction reaction products. RESULTS: The distribution of genotypes PDE4D in EW was as follows: CC - 42, CT - 49 and TT - 30 patients. In the con trol group, carriers of the same genotypes were 27, 21 and 15 persons respectively. All echocardiographic parame ters in EW workers and non irradiated patients did not differ significantly. Amongst TT genotype carriers of both groups the proportion of patients with increased myocardial mass index was the highest (82.9%) compared to CC genotype (78.4%) and CT (71.4%). The concentric type of left ventricular (LV) hypertrophy was found in 54.9% of patients with CC genotype, in 51.8% with CT genotype and 45.7% with TT genotype, while the eccentric type in 23.5, 21.4 and 37.1% respectively. The relative number of people with high LV end diastolic volume (EDV) normalized by body surface area (BSA) was 27.5% in CC genotype carriers, 26.8% in CT genotype and 40% in TT genotype carriers (p > 0.05). The increase of BSA indexed LV end systolic volume (ESV) was found in 27.5, 30.4 and 28.6%, and the ejection fraction in 15.7, 23.2 and 22.9% respectively. The largest number of CHD patients with inadequate dias tolic function was in carriers of TT genotype (75%) compared with the data in CC (66.7%) and CT genotypes (42.9%) carriers. CONCLUSIONS: In patients with the same genotype, both EW and non irradiated persons there were virtually no dif ferences in indicators of the structural and functional status of LV. The analysis of changes of LV structure the fol lowing feature was revealed: eccentric type of LV hypertrophy was more common for patients with TT genotype, but concentric type for CC genotype carriers. In one third of patients with CC and CT genotypes and in 40% of TT geno type carriers it was observed LV systolic function disorders. Diastolic dysfunction manifested as often in patients with TT genotype compared with CC and CT genotypes carriers.

[Urinary bladder substitution using combined membrane based on secretions of human mesenchymal stem cells and type I collagen].

AIM: Despite the widespread use of intestinal cystoplasty, urinary bladder substitution remains a challenging problem due to the complexity of operations and the potentially high risk of complications. A promising alternative may be bio-engineered collagen-based matrices containing stem cells or their secretions. MATERIAL AND METHODS: To evaluate the effectiveness of this bladder substitution modality, an experiment was conducted on 14 male rabbits. The animals underwent resection of urinary bladder, and the formed defect was substituted with a membrane of type I collagen (series 1, 5 rabbits) or a membrane of the same composition containing a conditioned medium with secretion of mesenchymal stem/stromal cells derived from human adipose tissue (series 2, 5 rabbits). In the comparison group (4 rabbits) resection of the bladder and the closure of the defect was carried out without bladder substitution (series 3). RESULTS: At 1 month after surgery, there was a complete epithelization of the inner surface of the implant, and body tissues replaced the collagen matrix. In series 1, the collagen implant was replaced mainly by connective tissue ingrown with occasional solitary smooth muscle cells. In series 2, the newly formed bladder wall contained numerous smooth muscle cells, growing into the collagen matrix and forming the muscular coat. In series 3, the muscular layer regeneration at the scar site was also noted, but it was less intense, which was confirmed by morphometry. In series 2, more active vascularization of the collagen implant occurred due to neo-angiogenesis, which was more intense than that in series 3, and especially in series 1. Functional studies revealed a reduced bladder functional capacity in series 1 and 3, while in series 2 it was close to normal. During filling cystometry, changes in intra-vesical pressure profile in series 2 were close to normal, while in series 1 and 3 infusion of a small volume of saline resulted in a marked increase in intra-vesical pressure, showing a reduced compliance of the reconstructed bladder. Discussion The study findings show that implants based on type I collagen can be effectively used to substitute a part of the urinary bladder wall, but bio-engineered collagen matrix grafts containing cell regeneration stimulants secreted by stem cells in their culture medium seem to be more promising.

DNA hydrolysis by monoclonal autoantibody BV 04-01.

Monoclonal anti-DNA autoantibody BV 04-01 catalyzed hydrolysis of DNA in the presence of Mg2+. Catalysis was associated with BV 04-01 IgG, Fab, and single-chain-antibody (SCA) proteins. Cleavage of both ss and dsDNA was observed with efficient hydrolysis of the C-rich region of A7C7ATATAGCGCGT2, as well as a preference for cleaving within CG-rich regions of dsDNA. Data on specificity of ssDNA hydrolysis and kinetic data obtained from wild-type SCA, and two SCA mutants were used to model the catalytically active antibody site using the previously resolved X-ray structure of BV 04-01. The resulting model suggested that the target phosphodiester bond is activated by induction of conformational strain. In addition, the antibody-DNA complex contained a Mg2+ coordination site composed of the L32Tyr and L27dHis side chains and a DNA 3'-phosphodiester group. Induction of strain along with the metal coordination could be part of the mechanism by which this antibody catalyzes DNA hydrolysis. Sequence data for BV 04-01 V(H) and V(L) genes suggested that the proposed catalytic-antibody active site was germline-encoded. This observation suggests that catalytic activity might represent an important-rarely examined-function for some antibody molecules.

DNA hydrolysis by monoclonal anti-ssDNA autoantibody BV 04-01: origins of catalytic activity.

Monoclonal anti-DNA autoantibody BV 04-01 catalyzed hydrolysis of DNA in the presence of Mg2+ ions. DNA hydrolyzing activity was associated with BV 04-01 IgG, Fab, and SCA 04-01 proteins. Pronounced cleavage specificity for both ss and dsDNA was observed with efficient hydrolysis of the C-rich region of the oligonucleotide A7C7ATATAGCGCGT7 as well as preference for cleavage within CG-rich regions of double-stranded DNA. Data on specificity of ssDNA hydrolysis and kinetic data obtained from wild-type SCA 04-01 and two SCA 04-01 mutants (L32Phe and L27dHis) were used to model the catalytically active antibody site utilizing the previously resolved X-ray structure of (dT)3 liganded Fab 04-01. The resulting model suggested that BV 04-01 activates the target phosphodiester bond by induction of conformational strain. In addition, the antibody-DNA complex contained a potential Mg2+ ion coordination site composed of the L32Tyr and L27dHis amino acid side chains and a DNA 3'-phosphodiester group. Induction of strain and metal coordination could be constituents of a mechanism by which this antibody catalyzed DNA hydrolysis. Sequence data for BV 04-01 VH and VL genes suggested that the proposed catalytic antibody active site was germ-line encoded. This observation suggests the hypothesis that catalytic activity might represent an important but unspecified function of some antibody molecules.

DNA-hydrolyzing autoantibodies.

Catalysis by antibodies could be a frequent phenomenon if the immune system generates a sufficiently diverse number of antibody-active sites, some of which may possess catalytic activity. A catalytic antibody can be expected to do more damage than one that simply binds antigen. The best biochemical marker of systemic lupus erythematosus (SLE) is presence of autoantibodies to DNA. In the present article, we describe the DNA-hydrolyzing activity of DNA-binding autoantibodies purified from SLE patients. The substrates employed were supercoiled plasmid, radiolabeled plasmid fragments, and oligonucleotides. Hydrolysis of DNA by the antibodies was indicated by the appearance of fragments visualized by ethidium bromide staining of agarose gels or autoradiography of polyacrylamide gels. Changes in linear dichroism values were also indicative of DNA hydrolysis. The antibody activity was purified by protein A-sepharose chromatography, high-performance liquid chromatography gel filtration, and DNA-affinity chromatography. Scrupulous control studies were done to demonstrate that DNA-hydrolyzing activity really belongs to the antibodies. Purified Fab fragments showed hydrolyzing activity, whereas the Fc fragment was inactive. The specificity of DNA cleavage was investigated, and the rate parameters of hydrolysis by antibodies and conventional nucleases were compared.

The family of human Na,K-ATPase genes. ATP1AL1 gene is transcriptionally competent and probably encodes the related ion transport ATPase.

The multigene family of human Na,K-ATPase is composed of 5 alpha-subunit genes, 3 of which were shown to encode the functionally active alpha 1, alpha 2 and alpha 3 isoforms of the catalytic subunits. This report describes the isolation, mapping and partial sequencing of the fourth gene (ATP1AL1) that was demonstrated here to be functionally active and expressed in human brain and kidney. Limited DNA sequencing of the ATP1AL1 exons allowed one to suggest that the gene probably encodes a new ion transport ATPase rather than an isoform of the Na,K-ATPase or the closely related H,K-ATPase.

[Study of expression of Na+,K+-ATPase subunit genes in the human brain using cDNA libraries]. / Izuchenie ékspressii genov alpha-sub''edinitsy Na+,K+-ATP-azy v mozge cheloveka metodom analiza kDNK-bankov.

Human brain cDNA libraries were screened with cDNA inserts corresponding to the mRNA for the Na+,K(+)-ATPase alpha-subunit from pig kidney. The results obtained demonstrate the existence of two highly homologous mRNAs encoding the alpha- and alpha III-isoforms of the Na+,K(+)-ATPase catalytic subunit.

[Tolerance for frustration as a reliability factor in the work of the human operator]. / Tolerantnost' k frustratsii kak faktor nadezhnosti raboty cheloveka-operatora.

Frustration tolerance is a personality trait that contributes to the reliable performance of an air traffic controller. This paper presents the results of a psychological examination of air traffic controllers using the Rosenzweig frustration test and emphasizes a correlation between the predominant behavior type in frustrating circumstances and professional success. The paper contains examples of realistic observations over air traffic controllers which confirm experimental data.