Biomedical subjects
Publications and source records attributed to M A Nikolaeva.
Expression of testicular angiotensin-converting enzyme on the surface of human spermatozoa was studied by means of flow cytometry with monoclonal antibodies. Expression of testicular angiotensin-converting enzyme on the cell surface depended on functional and morphological characteristics of spermatozoa.
The expression of testicular isoform of angiotensin-converting enzyme on the surface of human spermatozoa was evaluated by flow cytometry with monoclonal antibodies to epitopes of C-terminal domain of human angiotensin-converting enzyme and the mean path, mean curvilinear and straight-line velocities were determined by computer analysis of spermatozoon movement. A positive correlation between the expression of testicular isoform of angiotensin-converting enzyme on the cell surface and spermatozoon movement velocity was revealed.
Generation of free radicals in ejaculate samples from infertile patients was evaluated using chemiluminescent technique. The presence of antisperm antibodies in samples increased the possibility of damages to spermatozoon plasma membranes due to excessive generation of free radicals.
Conflicting opinions about the effects of antisperm antibodies on fertilization can be due to inadequacy of experimental approaches in evaluating the antisperm immunity. Detection of antisperm antibodies bound to the surface of live spermatozoa can be associated with aggregation of surface antigen-antibody complexes followed by metabolic activation of spermatozoa and acrosomal reaction, which impair cell resistance. New concept of antisperm immunity and its influence on reproduction can be formulated only after comprehensive studies of the mechanisms of spermatozoon response to binding of antisperm antibodies. Further improvement of quantitative assays of antisperm antibodies and evaluation of their effect on spermatozoon function should be aimed at selection of experimental conditions preventing changes in spermatozoa coated with antisperm antibodies during in vitro manipulations.
Flow cytometry (FCM) analysis of live antibody-coated spermatozoa subjected to immunofluorescence staining (FCM test) is considered an objective method for the quantitative detection of antisperm antibodies (ASA). But the cross-linking of cell surface antigen (Ag) with bivalent antibodies and/or antigen-antibody (Ag-Ab) complexes with second antibodies may induce the reorganization of surface components (patching and capping) and result in their shedding from the sperm surface. The present study estimates the relationship between aggregation of Ag-Ab complexes on the sperm surface and the results of indirect FCM test. Swim-up spermatozoa of normozoospermic men were incubated with ASA-positive sera from infertile patients and with second antibodies fluorescein isothiocyanate (FITC)-labelled goat anti-human IgG polyclonal antiserum under different conditions and then analysed by FCM and fluorescence microscopy. It was shown that low temperature, cytochalasin B, excess or lack of the primary and/or secondary antibodies and sperm fixation by paraformaldehyde may inhibit aggregation and shedding of Ag-Ab complexes and dramatically increase ASA quantity determined on the sperm surface. However, inhibition of aggregation on the live sperm surface was observed only in a minority of ASA-positive samples and was poorly reproducible using semen of different donors. A high probability of Ag-Ab complex shedding from the sperm surface during experimental manipulation limits the use of indirect FCM test for quantitative ASA determination.
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Acrosome status in human spermatozoa from 20 normozoospermic men was evaluated by flow cytometry following the induction of the acrosome reaction with the ionophore A23187. Dual fluorescence staining of methanol fixed spermatozoa incubated with and without (control) the ionophore A23187 was performed with probes which targeted the outer acrosomal membrane (OAM) (rhodamine-labelled Arachis hypogaea agglutinin) or constituents of the acrosomal vesicle (fluorescein-labelled Pisum sativum agglutinin). Flow cytometry analysis revealed two major subpopulations of cells: acrosome-intact and acrosome-reacted spermatozoa after induction of the acrosome reaction. The intensity of green and red fluorescence in acrosome-reacted spermatozoa was significantly lower than that of the acrosome-intact control spermatozoa (P < 0.0001). The intensity of green fluorescence in the acrosome-intact subpopulation of spermatozoa was significantly higher than that of the control population (P < 0.002). Exposure of spermatozoa to the ionophore A23187 resulted in reliable enhancement of the number of spermatozoa with very high intensity of green and/or red fluorescence compared with the control (P < 0.03). An inverse correlation between the number of acrosome-reacted spermatozoa and spermatozoa with a very high intensity of green and/or red fluorescence was demonstrated (r = -0.631, P < 0.01). This method provides an objective and efficient procedure for quantitative estimation of the acrosomal status of human spermatozoa.
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OBJECTIVE: To investigate the influence of sera and peritoneal fluids (PFs) from fertile and infertile women on the binding of antisperm antibodies to the surface of spermatozoa. DESIGN: The immunoglobulin (Ig) G antisperm antibodies binding to the surface of liver spermatozoa was evaluated after their incubation in antisperm antibodies-positive serum from an infertile male in the presence and absence of female sera or PFs. SETTING: Russian Scientific Center for Obstetrics, Gynaecology, and Perinatology. PATIENT(S): Serum and PF samples from fertile and infertile women; antisperm antibodies-positive serum from infertile men; high-quality fresh semen from healthy donors. INTERVENTION(S): Serum samples were obtained from fertile and infertile women and from infertile men. Peritoneal fluids were collected during routine laparoscopy. MAIN OUTCOME MEASURE(S): The proportion of spermatozoa positive for IgG antibodies and the quantity of antisperm antibodies on the sperm surface measured by flow cytometry (FCM). RESULT(S): The addition of sera from fertile or infertile women with endometriosis or pelvic adhesion disease to an IgG antisperm antibodies-positive male serum resulted in significant inhibition of the antisperm antibodies binding to the sperm surface. CONCLUSION(S): Sera of fertile as well as infertile woman contain factors that block IgG antisperm antibodies binding to the surface of live spermatozoa.
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OBJECTIVE: To assess the use of flow cytometry (FCM) analysis of living spermatozoa subjected to indirect immunofluorescence staining with the mouse anti-human immunoglobulin (Ig)G monoclonal antibodies (FCM test) as an objective method for the detection of antisperm antibodies in semen and serum. DESIGN: The comparative studies were conducted with the direct and indirect mixed antiglobulin reaction (MAR) and FCM test performed by FCM analyzer FACSCAN (Becton Dickinson Immunocytometry Systems, Mountain View, CA). SETTING: Clinical infertile service. PATIENTS: Seventy-six semen and 20 sera samples of infertile patients were analyzed. RESULTS: For IgG in semen there was a highly significant correlation between the direct MAR and FCM test. Six of 12 IgG-positive samples revealed also IgA, and just one sample, which showed negative MAR, was positive for IgM in FCM test. Flow cytometry test and indirect MAR showed also a highly significant correlation of the results of IgG detection in sera. CONCLUSIONS: Flow cytometry test may be used to verify that IgG antisperm antibodies are on the surface of living spermatozoa either retrieved directly from an ejaculate or after exposure to serum and to determine proportion of antisperm antibodies-positive spermatozoa, titer, and quantity of antibodies bound to the cell surface.
Distribution of surface lymphocyte markers was assessed in foetus lung by cytofluorimetry using monoclonal antibodies produced by "Ortho Diagnostic systems" and "Becton Dickinson". Seven lymphocyte subpopulations were detected in developing lungs even at the pseudoglandular stage (8-15 weeks). There was a significant increase in these lymphocyte subpopulations in canalicular stage. Quantity of T3+ cells increased 8-fold and B-cells 10-fold. There was a tendency to increased number of cortical thymocytes (T6+), and to decreased ratio T6+/T3+ lymphocytes. The ratio of immunoregulatory lymphocyte subpopulations (T4+/T8+) resembled that of these subpopulations in adult human lung rather than in cord blood. In canalicular stage the share of lung lymphoid cells in S-phase was decreased.
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The proposed article provides data on labour conditions assessment in radio items processing and characteristics of major unfavourable factors, e.i. air contamination, degrees of illumination, noise, temperature and labour intensity. Expounded are different degrees of the involvement of all these factors depending on the automatization level. Health related labour conditions improvement measures are substantiated and given detailed descriptions.
Detection of antisperm antibodies is an important step of the diagnosis of infertility. In present study we conducted comparative studies with the detection of antisperm antibodies in IgG, IgA and IgM classes on the surface of living spermatozoa by flow cytometry and with Mixed Agglutination Reaction and Sperm immobilization. Our data show that flow cytometry may be used to verify that immunoglobulins are on the surface of living spermatozoa, either retrieved directly from an ejaculate or following exposure to cervical mucus or serum and to determine their isotype, proportion of sperm which bound antibody and quantity of immunoglobulins bound to cell surface.
Endothelial cells (EC) were harvested by 0.1% collagenase treatment for adult human thoracic aortas obtained 1-3 h after sudden death. At least 35-70% of EC were removed from the intimal surface of aorta, 90-95% of them being viable. Plating efficiency was 70-80%. Monolayer formation was achieved at a seeding density of 5-8 X 10(2) cells/mm2. The cells were identified as endothelium by the presence of Factor VIII antigen, Weigel-Palade bodies and typically endothelial morphology at confluence. Unlike endothelial cultures derived from human umbilical veins and infant aortas, primary cultures obtained from human adult aortas contain multinuclear EC with Factor VIII antigen and Weibel-Palade bodies. The number of multinuclear EC in cultures isolated from aortas affected by atherosclerosis was 2-fold higher (P less than 0.05) than in cultures obtained from grossly normal aortas taken from donors of the same age. EC with numerous lipid inclusions revealed by oil-red-O staining were present in all the EC primary cultures derived from aortas affected by atherosclerosis. No oil-red-O-positive cells were detected among the EC cultured from infant aorta, aorta of young donors, and umbilical vein. An electron microscopic examination of EC from atherosclerotic aorta in culture and in situ failed to reveal any ultrastructural peculiarities distinguishing multinuclear EC from the mononuclear EC.