Biomedical subjects
Publications and source records attributed to L A Zakharova.
Morphofunctional immune disorders were revealed in vasopressin-deficient Brattleboro rats with diabetes insipidus during ontogeny. We observed a permanent decrease in the number of blood lymphocytes, increase in neutrophil count, reduced activity of macrophages, early involution of the thymus and spleen, and suppression of antibody production. These changes reflect impaired general resistance of these animals.
The role of the neuroendocrine system in the development of cell-mediated immunity has been studied in fetal rats. The spontaneous and mitogen-induced proliferation of liver lymphocytes and thymocytes was evaluated in vitro in rats at the 22nd prenatal day following surgical ablation of the forebrain (encephalectomy) or of the entire brain and pituitary (decapitation) in rat fetuses in utero at the 18th day. Non-operated and sham-operated fetuses served as controls. The ablation of the entire brain and pituitary in rat fetuses resulted in an increase (40-60%) of spontaneous proliferation of liver and thymic cells in comparison with sham-operated fetuses. The ablation of the forebrain including the hypothalamus caused a decrease in the mitogenic proliferative response of thymocytes and liver lymphocytes for 40 and 20%, respectively. The ablation of the entire brain including the hypothalamus and pituitary resulted in a 80% decrease of the proliferative response of thymocytes and in the full suppression of proliferation of liver lymphocytes. The immune proliferative response was restored by the LHRH administration either systemically to operated fetuses (0.2 microg/fetus) or to the cell culture (10(-9) and 10(-7) M). It was concluded that the central nervous system was important for maturation of the immune system in rats during the prenatal period. In particular, neuroendocrine system are likely to play a major role as LHRH treatment in vitro and in vivo appeared to contribute to this regulation.
The effect of endogenous luteinizing hormone-releasing hormone (LHRH) on the proliferation induced by concanavalin A (Con A) in rat fetal thymocytes was studied. A selective antagonist (2 microg per fetus) or antibodies to LHRH (20 microl per fetus) were injected in utero into 20-day-old rat fetuses, and this resulted in a two- or fivefold decrease in the Con A-induced proliferation of thymocytes, respectively. In combined culture of the antagonist (10-5-10-6 M) with fetal thymocytes, the proliferative response was not decreased. The concentration of LHRH was determined by radioimmunoassay in tissues of immunocompetent organs and in blood serum of 18- and 21-day-old fetuses, and the hormone was found in the hypothalamus, thymus, and peripheral blood. The initially low level of LHRH in the thymus increased by 65 and 40%, respectively, on the first day after birth and became similar to the level in the hypothalamus. In the fetal blood serum, the LHRH level was significantly higher than in the thymus and hypothalamus of fetuses of the same age. The hormone concentration was greatest in the 18-day-old fetuses, and it decreased twofold by the 21st day. The findings indicate that LHRH is involved in regulation of T-cell immunity even during prenatal ontogenesis.
In this study the influence of mu-, delta-, and kappa-selective opioid agonists (DAMGO, DSLET, and dynorphin A (1-13)) on cytoplasmic free Ca2+ ([Ca2+]i) level in normal and concanavalin-A (Con A)-activated mouse lymphocytes was investigated. [Ca2+]i was measured using the fluorescent dye FURA-2AM. The opioid peptides at 10-12-10-7 M induced some increase in [Ca2+]i in non-activated lymphocytes. However, DAMGO and DSLET (10-13-10-7 M) considerably inhibited a Con A-induced increase in [Ca2+]i. The inhibiting effect of both peptides was higher after 20-min preincubation compare to 2-h preincubation. The effect of the kappa-agonist dynorphin A (1-13) was significantly different depending on the duration of cell pretreatment and the concentration of the peptide used. After preincubation for 20 min at low concentrations (10-12-10-11 M) it slightly stimulated, while at higher (10-10-10-7 M) concentrations it inhibited lymphocyte response to Con A. After preincubation for 2 h, pronounced stimulation of mitogen-induced Ca2+ flux was observed at peptide concentration 10-9 M. The effects of opioids were antagonized by naloxone. These data indicate that functionally active opioid receptors expressed on lymphocytes could be involved in early stages of mitogen activation.
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The naloxone-dependent modulating effect of synthetic analogs of myelopeptides, MP1 and MP2, on pain sensitivity in mice was determined at doses 10(-13) and 10(-8) g/animal. Binding of the peptides to opioid receptors of mouse brain membranes was studied by the radioligand assay. For MP1, the displacement of [3H]DAGO (selective mu-agonist) by IC50 = 7.3 x 10(-5) M and [3H]DSLET (selective delta-agonist) with IC50 = 7.0 x 10(-5) M. The data obtained suggest that the hypoalgesic effect of the peptides cannot be due to their direct interaction with opioid receptors. However, the naloxone dependence provides a possibility of mediated involvement of the opioidergic system in the realization of this effect.
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Met-enkephalin (ME) in the range of concentrations 10(-15) M to 10(-9) M has exhibited an immunomodulating effect on concanavalin A (Con A) induced proliferation of mice lymphocytes from lymph nodes in vitro. The effect of ME was shown to vary with the stage of lymphocyte activation and to depend on the mitogen dose. In the case that ME and Con A were injected simultaneously, at the zero time of proliferation, maximum inhibition was observed on the first and fourth days. In an other set of experiments when ME was added at intervals of 4 hours before measuring the proliferation response value, both inhibition (after 24 hours of proliferation) and stimulation (after 28 and 96 hours) were observed. Opioid ligands of various classes were found to act in the same manner as ME. Naloxone was shown to block the immunomodulating effect of opioids.
The alteration of opioid receptor total binding level as well as ae- and mu-, delta-type distribution has been investigated on the every day of primary and secondary immune response on bovine gamma-globulin. It has been shown that a level of opioid binding measured with [3H]naloxone increased by 6 times during the primary, and by 1.5 times during the secondary immune response on the antigen. A maximal value of opioid receptor activity has been observed at the 7-th and the 5-th days correspondingly, i.e. at the peak of the response. Receptor activity has been measured by displacement of [3H]naloxone with corresponding selective opioid ligand. Growth of the quantity of ae-receptors and decrease of that of mu- and delta-receptor types has been found to take place starting with zero time till 7-th and 5-th days of response development. A reverse picture has been detected during the immune response falling down. Selective ae-ligand stimulated the secretion of specific antibodies to bovine gamma-globulin, whereas mu- and delta-ligands suppressed the antibody production. Functional role of different types of opioid peptides taking part in immune regulation is analyzed.
The influence of Met-enkephalin on mitogenic stimulation of mouse splenocytes was investigated. Met-enkephalin (ME) was shown to suppress proliferation induced by Concanavalin A and activate proliferation induced by Staphylococcus enterotoxin A. Both effects were revealed at low (down to 10(-14) M) concentration of pentapeptide. Naloxone reversed ME influence on cell activation. The number of receptors for naloxone was shown to increase up to 2.5-fold during mitogenic activation. The difference in expression of various types of opioid receptors at mitogenic stimulation was demonstrated by ligand displacement experiments.
The influence of met-enkephalin on specific antibody production by lymphocytes from mouse lymph nodes was studied in vitro in productive phase of immune response. It was shown that the peptide did not influence secretion of IgM-antibody to T-independent antigen-trinitrobenzensulfoacidic group, but suppressed secretion of IgG-antibody to T-dependent antigens both during primary and secondary response. The efficiency of superlow concentrations of the peptide (10(-15)-10(-14) M) for the response to ovalbumin, but not for the response to bovine gamma-globulin was shown. All effects of met-enkephalin were naloxone-reversible. The existence of individual distribution in dose-dependences of peptide action on antibody secretion in response to ovalbumin was demonstrated.
The binding of the opioid antagonist [3H]-naloxone to immunocompetent cells of the mouse, F1(CBA x C57B1/6), in medium 199 has been studied. The binding was reversible and reached a maximum during 15-20 min at 37 degrees C. The stereospecificity profile was proven to correspond to mu-type receptors. The binding curve was characterized by high positive cooperativity (nH = 2.3, IC50 = 5 nM). Mitogenic stimulation by Con A, SEA, and ML caused an increase in the number of receptors. Besides, stimulation by an antigen (ovalbumin) changed the binding parameters. The distribution of binding sites for naloxone on various immunocompetent cells was investigated. The maximal number of sites was found on lymphocytes of lymph nodes and bone marrow cells. A conclusion is drawn that both T- and non-T-cells play a role in naloxone binding.
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