RESUMO
Adenosines, endogenous purine nucleosides, appear in the extracellular space under metabolically stressful conditions associated with ischemia, inflammation, and cell damage. Their activity on innate immunity is prevalently inhibitory and can develop both in infectious and neoplastic diseases. During cancer development, tumor cells that release high concentrations of adenosines can impair the function of tumor-infiltrating lymphocytes and assist tumor growth by neo-angiogenesis. We evaluated the influence of A(2) adenosine receptor (A(2)AR) agonist on cytotoxic-cell response comparing human with other mammalian species (rodents, pigs, goats), both in healthy and in cancer conditions. The A(2)AR agonist developed dose-dependent inhibition of the cytotoxic activity of immune effector cells in all studied species. However, variability of the response was observed in relation to the species and the target cells that were used. Altogether, our data indicate that the A(2)AR plays a central role in adenosine-mediated inhibition of immune response to tumors.
Assuntos
Agonistas do Receptor A2 de Adenosina , Adenosina/análogos & derivados , Citotoxicidade Imunológica/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Adenosina/imunologia , Adenosina/farmacologia , Animais , Células Cultivadas , Feminino , Cabras , Humanos , Células Matadoras Naturais/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/imunologia , Ratos , Ratos Wistar , Receptores A2 de Adenosina/imunologia , Roedores , SuínosRESUMO
The aim of this work was to evaluate ontogeny of reactive nitrogen species (RNS) production by peripheral blood phagocytes in pig. Pig fetuses (55 and 92 days of gestation) and postnatal piglets (1, 3, 8, 17, 31 and 41 days after birth) were used. RNS production was measured by fluorescent probes diaminofluorescein-diacetate (DAF-FMDA) and dichloro-fluorescein-diacetate (H2DCFDA). Levels of nitration of cell proteins were established by immunofluorescent detection of nitrotyrosine. Levels of plasma nitrites/nitrates were detected spectrophotometrically by Griess reaction. Nitric oxide production measured by DAF-FMDA in neutrophils decreased during postnatal life. Spontaneous RNS measured by H2DCFDA decreased from 55th day of gestation to the 41st day of life. Phorbol-12-myristate-13-acetate activated production decreased during postnatal life. Production of NO measured by DAF-FMDA in macrophages decreased from the first to 41st day after birth. RNS production measured by H2DCFDA in monocytes did not show any significant changes during ontogeny. The level of nitrotyrosine significantly decreased from the third to 17th day. Levels of plasma nitrites/nitrates gradually decreased from the 55th day of gestation to the 41st day after birth. A temporary increase in all parameters occurred after weaning, but without any significance. In conclusion, RNS production has a decreasing trend during ontogeny and is transiently upregulated after weaning.
Assuntos
Envelhecimento/sangue , Fagócitos/metabolismo , Espécies Reativas de Nitrogênio/metabolismo , Sus scrofa/crescimento & desenvolvimento , Animais , Animais Recém-Nascidos , Desenvolvimento Fetal , Estudos Longitudinais , Óxido Nítrico/metabolismo , Sus scrofa/sangue , Sus scrofa/embriologia , DesmameRESUMO
Depression is a complex disorder related to chronic inflammatory processes, chronic stress changes and a hippocampal response. There is a increasing knowledge about the role of glial cells in nutrient supply to neurons, maintenance of synaptic contacts and tissue homeostasis within the CNS. Glial cells, viewed in the past as passive elements with a limited influence on neuronal function, are becoming recognized as active partners of neurons and are starting to be discussed as a possible therapeutic target. Their role in the pathogenesis of depressive disorders is also being reconsidered. Attention is devoted to studies of the different types of antidepressants and their effects on transmembrane signaling, including levels of alpha subunits of G proteins in C6 glioma cells in vitro as a model of postsynaptic changes in vivo. These models indicate similarities in antidepressant effects on G proteins of brain cells and effector cells of natural immunity, natural killers and granulocytes. Thus, an antidepressant response can exhibit certain common characteristics in functionally different systems which also participate in disease pathogenesis. There are, however, differences in the astrocyte G-protein responses to antidepressant treatment, indicating that antidepressants differ in their effect on glial signalization. Today mainstream approach to neurobiological basis of depressive disorders and other mood illnesses is linked to abnormalities in transmembrane signal transduction via G-protein coupled receptors. Intracellular signalization cascade modulation results in the activation of transcription factors with subsequent increased production of a wide array of products including growth factors and to changes in cellular activity and reactivity.
Assuntos
Antidepressivos/farmacologia , Transtorno Depressivo/fisiopatologia , Neuroglia/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Adaptação Psicológica/efeitos dos fármacos , Animais , Transtorno Depressivo/tratamento farmacológico , Transtorno Depressivo/etiologia , Humanos , Transdução de Sinais/fisiologia , Estresse Psicológico/complicações , Estresse Psicológico/fisiopatologia , Transmissão Sináptica/efeitos dos fármacosRESUMO
The decapeptide QLNLKEYNLV corresponding to the C-terminus of Gq/G11alpha guanine nucleotide-binding proteins (G-proteins) was synthesized by the solid-phase method and conjugated to keyhole limpet hemocyanin. The rabbits were immunized with these conjugates and an antiserum that reacted specifically with the alpha subunit of Gq/G11 proteins was used in this study. The antiserum exhibited no cross-reactivity with the alpha subunits of stimulatory (Gs) or inhibitory (Gi) G-proteins associated with adenylate cyclase. Immunoblots with the antiserum showed that it specifically recognized the Gq/G11 alpha-proteins in cholate extracts of adipose tissue membranes of goats. Treatment of young castrated male goats with bST had no effect on the quantity of Gq/G11 alpha subunits in adipose tissue and the results thus obtained did not support the idea that the bST signal in adipose tissue is transmitted via Gq/G11 alpha-proteins.
Assuntos
Tecido Adiposo/química , Tecido Adiposo/efeitos dos fármacos , Cabras/metabolismo , Hormônio do Crescimento/farmacologia , Proteínas Heterotriméricas de Ligação ao GTP/análise , Tecido Adiposo/metabolismo , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Western Blotting , Proteínas Heterotriméricas de Ligação ao GTP/imunologia , Proteínas Heterotriméricas de Ligação ao GTP/metabolismo , Soros Imunes/biossíntese , Soros Imunes/imunologia , Masculino , Orquiectomia , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , Coelhos , Fosfolipases Tipo C/metabolismoRESUMO
Ergot alkaloids (EAs), products of Claviceps spp., are widely used in various fields of clinical medicine (neurology, psychiatry, endocrinology). In the present work we studied the neuroimmunomodulative effect of EAs on activation of NK cells and their signalling pathways. Furthermore, the killing capability of rat NK cells in vitro was examined in the presence of glycosidic derivatives of elymoclavine, agroclavine, and liposome-encapsulated EAs. The engagement of appropriate NK cell membrane receptors by EAs cause an indirect enhancement of adenylyl cyclase system through inhibition of G-protein al,2-subunit (up to 50 % of control values). All of the tested EAs enhanced the rat NK cell-mediated cytotoxic activity in vitro, particularly against target cells of astrocyte origin (C-6 glioma). The present results argue for a possible EA immunomodulatory role of cell-mediated immunity in tumour regression processes.
Assuntos
Alcaloides de Claviceps/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Neuroimunomodulação/efeitos dos fármacos , Animais , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Claviceps , Citotoxicidade Imunológica/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Ergolinas/administração & dosagem , Ergolinas/farmacologia , Alcaloides de Claviceps/administração & dosagem , Humanos , Lipossomos , Camundongos , Monócitos/citologia , Monócitos/efeitos dos fármacos , Oligossacarídeos , Ratos , Transdução de Sinais/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
An allogeneic reaction among brain cortex cells (mixed reaction) was demonstrated previously by H-2 alloantigen-induced uncoupling of oxidative metabolism (Kováru Med. Biol. 58: 273, 1980). In the present study we have demonstrated that alloantigen already increased cell surface Na+,K+-ATPase activity after 100 min when the enzyme activation was highest at Mg2+/ATP ratio 4: 1. The allogeneic cell reaction was accompanied by an elevation of membrane lipid fluidity and probably also by a thermotropic lipid phase transition which might influence the membrane lipid-dependent Na+,K+-ATPase activity, while Mg2+-ATPase remained unaffected. Furthermore, the effects of proteins and peptides released into the supernatant during the allogeneic reaction were analyzed in brain cortex cells. One of the isolated active peptide fractions, FA (m.w. lower than 2.5 kD), was able to enhance Na+,K+-ATPase activity as well as to block K+-evoked O2 uptake by brain cortex cells. Thus the FA fraction simulated primary allorecognition events. The data indicate that various brain cell surface domains were influenced by a regulatory peptide fraction of the cytokine type during the early phase of allogeneic reaction. Allorecognition among brain cortex cells is directed against functionally important metabolic reactions.
Assuntos
Córtex Cerebral/imunologia , Isoantígenos/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Células Cultivadas , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/enzimologia , Magnésio/metabolismo , Masculino , Fluidez de Membrana , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos CBA , Consumo de Oxigênio/efeitos dos fármacos , ATPase Trocadora de Sódio-Potássio/metabolismoRESUMO
The aim was to study the role of major histocompatibility complex (MHC), in mice named H-2, during early allogeneic reactions (AR) of brain cortex cells or lymphocytes. We used neuronal and glial enriched perikarya, spleen and thymus lymphocytes or their subpopulations. Rat AR was also assayed between C-6 astrocytoma cells and spleen lymphocytes. We demonstrated that: 1) H-2 dependent stimulation of Na+,K+-ATPase and ouabain-sensitive K+-dependent p-nitrophenylphosphatase (K+-pNPPase) activities represented specific response in both AR of unseparated brain cells or lymphocytes. On the other hand, non-specific AR-induced stimulation of Ca2+-ATPase activity was observed. 2) Allogeneic enriched glial fractions reacted similarly by the same enzyme activation in contrast to no change in AR between enriched neuronal fractions. Allorecognition ability of glial cells was confirmed by AR between C-6 astrocytoma cells and lymphocytes. 3) Mature thymus lymphocytes exerted alloreactivity by specific activation of Na+,K+-ATPase or K+-pNPPase, in contrast to no change in AR between immature lymphocyte subpopulations. 4) MHC Class II monoclonal antibody inhibited Na+,K+-ATPase and K+-pNPPase activities in brain cells as well as in thymus and spleen lymphocytes in a dose-dependent manner. Results support former studies about alloantigen-induced uncoupling in brain oxidative cortex metabolism (Kováru Med. Biol. 58: 273, 1980) via Na+,K+-ATPase and K+-pNPPase inhibition by mechanism which can mimic MHC restriction.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Córtex Cerebral/enzimologia , Córtex Cerebral/imunologia , Antígenos H-2/imunologia , Linfócitos/enzimologia , Linfócitos/imunologia , 4-Nitrofenilfosfatase/imunologia , 4-Nitrofenilfosfatase/metabolismo , Animais , Anticorpos Monoclonais/imunologia , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Ratos , ATPase Trocadora de Sódio-Potássio/imunologia , ATPase Trocadora de Sódio-Potássio/metabolismo , Propriedades de Superfície , Células Tumorais CultivadasRESUMO
This study describes the effects of cytokine peptides released into the supernatant during an early allogeneic reaction (AR) of mouse spleen lymphocytes or brain cortex cells which differ in their major histocompatibility complex (MHC). The peptides were isolated by ultrafiltration, liquid chromatography and HPLC. We found that both peptides stimulated the cell surface Na+,K+-ATPase and Ca2+-ATPase activities of quiescent spleen lymphocytes in vitro and mimicked early allogeneic cell interactions. Both brain and spleen AR peptides inhibited Concanavalin A-stimulated spleen lymphocyte proliferation, whereas 3H-TdR incorporation into DNA of the E7 neuroblastoma cell line was stimulated by these peptides. The peptide isolated from the supernatant of the allogeneic brain cell reaction inhibited phagocytosis in phorbol myristate-stimulated LA5-9/8 mouse macrophage cell line. Immunosuppressive activity of spleen AR peptide is supported by inhibition of spontaneous E rosette formation by lymphocytes. The immunosuppressive effect of isolated peptide cytokines on lectin-activated lymphocytes was comparable with the serum thymic factor (FTS, Lenfant et al. 1983). These changes demonstrate the pleiotropic cytokine actions mediated by plasma membrane of immune system and brain cells.
Assuntos
Córtex Cerebral/metabolismo , Citocinas/metabolismo , Sistema Imunitário/metabolismo , Peptídeos/metabolismo , Baço/metabolismo , Animais , Bradicinina/metabolismo , ATPases Transportadoras de Cálcio/metabolismo , Divisão Celular , Concanavalina A/farmacologia , DNA/metabolismo , Sistema Imunitário/imunologia , Linfócitos/efeitos dos fármacos , Linfócitos/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C3H , Neuroblastoma/imunologia , Neuroblastoma/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Baço/imunologia , Propriedades de Superfície , Fator Tímico Circulante/metabolismo , Timidina/metabolismo , Tuftsina/metabolismo , Células Tumorais CultivadasRESUMO
Gel electrophoresis of DNA was used for estimation of DNA changes caused in C6 glioma cells by treatment with psychotropic drugs (imipramine, amitryptiline and fluoxetine). Some discrete bands containing a population of short DNA fragments appeared after 1 and 5 days of cultivation. Apoptotic DNA breaks were verified at single cell level using the TUNEL test in cells treated with fluoxetine.
Assuntos
Antidepressivos/farmacologia , Apoptose , Glioma/patologia , Imipramina/farmacologia , Amitriptilina/farmacologia , Fragmentação do DNA/efeitos dos fármacos , Fluoxetina/farmacologia , Humanos , Células Tumorais CultivadasRESUMO
Ouabain enhances a modulation effect of lipopolysaccharide (LPS) on the expression of MHC class II antigens. A similar but lesser effect than that of ouabain, which increases intracellular concentration of sodium ions, has been observed when treating cells with ionophores quercetin which increases [Ca2+]i or valinomycin which increases [K+]i. These findings confirm the role of ion flux in cellular activation and differentiation. Optimal concentration of ouabain is 10(-7) M with 10 micrograms of LPS/ml for human and pig peripheral blood cells. The increase in radioactivity of mononuclear human blood cells after the binding of 125I-anti-MHC class II monoclonal antibody is at the fourth hour of cultivation with LPS and ouabain, the number of immunofluorescently positive cells rises at the twelfth hour. A similar modulation has been observed on pig kidney cell line. It can be concluded that the expression of class II antigens on the plasma membrane can be induced by cumulative effect of LPS and agents alterating ion transport.
Assuntos
Antígenos de Histocompatibilidade Classe II/biossíntese , Ouabaína/farmacologia , Animais , Anticorpos Monoclonais/imunologia , Células Cultivadas , Imunofluorescência , Antígenos HLA-DP/biossíntese , Antígenos HLA-DR/biossíntese , Humanos , Ativação Linfocitária/efeitos dos fármacos , SuínosRESUMO
Alpha-fetoprotein (AFP) was found to suppress the phagocytic activity of the blood monocytes and neutrophils in vitro. The amounts of AFP detectable by immunofluorescence in the livers of nu/nu, nu/+ and +/+ mice were quite comparable, and thus could not have been responsible for the alterations in phagocytosis found in leukocytes of athymic nude mice during their ontogenetic development.
Assuntos
Fagocitose , alfa-Fetoproteínas/imunologia , Fatores Etários , Animais , Técnicas In Vitro , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Nus , Fagócitos/efeitos dos fármacos , Fagócitos/imunologia , Fagocitose/efeitos dos fármacos , alfa-Fetoproteínas/metabolismo , alfa-Fetoproteínas/farmacologiaRESUMO
Surface markers were demonstrated on pig lymphocytes using anti-T cell-IgG and anti-Helix pomatia (HP) IgG during prenatal and postnatal development. A fluorescence-activated cell sorter analysis of T-cell surface markers was accompanied by an image analysis to prove the association of T antigenic determinants with the plasma membrane only. We found development-dependent changes in both anti-T cell and HP surface markers in both primary and secondary lymphatic organs. The number of T-positive (T+) cells estimated by anti-T cell-IgG was very similar to the results obtained by spontaneous E-rosette forming tests. At all selected age intervals, changes in the number of T+ cells were not significant in the thymus, but a marked increase in T+ cells was found in both spleen and lymph nodes. The image analysis confirmed the expression of T cell markers on the cell surface. The distribution of T cell markers was uneven, i.e. various degree of fluorescence intensity on whole ring-pattern projection of the cell surface image was estimated. In second lymphatic organs especially, fluorescence intensity of cells, i.e. total number of T cell markers estimated by anti-T cell-IgG, increased with age. On fetal day 73, T cell markers were slightly expressed, but very high fluorescence intensity and heterogeneous distribution of T cell markers on lymphocytes were found on fetal day 107 and postnatal day 56. The results indicate the possibility of functional maturation of various T cell markers on T cell subsets, furthermore a different degree of expression of T cell markers on various T cell subsets can be suggested. The number of HP+ cells increased with age in both primary and secondary lymphatic organs. In the prenatal period, the expression of HP receptors was very weak in both primary and secondary organs in contrast to the marked increase in HP+ cells during the postnatal interval. Differences in fluorescence intensity of cells were found, representing the increase by 22% in thymus cells comparing to cells of secondary lymphatic organs. Heterogeneity of HP+ cell populations in thymus was shown by the Scatchard plot, indicating at least two subpopulations of HP+ cells with different avidity to HP. Cells with low HP avidity could include a subset with cytolytic activity.
Assuntos
Linfócitos/imunologia , Suínos/imunologia , Fatores Etários , Animais , Antígenos de Superfície/análise , Separação Celular/métodos , Citometria de Fluxo , Coelhos , Formação de Roseta , Ovinos , Baço/citologia , Baço/imunologia , Timo/citologia , Timo/embriologiaRESUMO
Alpha-fetoprotein (AFP) has been screened in the brain of pig embryos by immunocytofluorescence, crossed immunoelectrophoresis and the immunoblotting technique. Immunocytochemistry showed that the cell and tissue localization of AFP changes with age of the embryos, and/or maturation of the brain. At the earliest stages studied (embryonic days (ED) 26 to 28), the AFP in the brain prevails near the cerebral ventricles, while later (ED 35-46) AFP also appears in higher concentrations in the differentiating parenchyma of some regions of the brain (especially in deep layers of the cortical plate or the region of the differentiating hippocampus) and spinal cord. Its intracellular presence was well evident in the large neuroblasts of these regions. During the 3rd month of embryonic life AFP starts to disappear from the brain parenchyma. At all ages studied, bright fluorescence in the choroid plexus and the meningovascular apparatus is evident. In the choroid plexus as well as ependymal cells a mosaic-like pattern of fluorescence appeared, especially near its invagination into the lateral ventricles; the latter suggests transependymal transport of AFP by some cells. Single and crossed immunoelectrophoresis revealed one reacting band in the water extract of homogenate of the brain of 35-day-old pig embryos. Similarly, only one band has been stained in SDS electrophoretograms prepared from such extracts and stained by the immunoblotting technique; its molecular weight corresponded to the pig AFP. In contrast to the brain the AFP band in the liver was also found in the membrane fraction. It is concluded that maximal tissue concentrations of AFP in the pig brain are attained during the second embryonic month and that, in addition to extracellular fluids, it is also transitionally present in the cytoplasmic compartment of brain cells, especially those more advanced in maturation.
Assuntos
Encéfalo/embriologia , alfa-Fetoproteínas/análise , Animais , Encéfalo/citologia , Feminino , Imunofluorescência , Imunoeletroforese , Imunoeletroforese Bidimensional , Gravidez , SuínosRESUMO
Alpha-fetoprotein (AFP) and some other serum proteins have been studied in the developing brain of rats and pigs using the indirect immunofluorescence technique. AFP is shown to be present in the ventricular ependyma, meningeal envelopes, the choroid plexus, blood vessel walls and in a wide scale of differentiating parenchymal cells ever since early embryonic ages of both species. In brain parenchyma the content of AFP is low in immature germinative cells; in both species it starts increasing in postmigratory neuroblasts and reaches a peak at the time of accelerated nerve cell differentiation. In rats, the amount of AFP is highest towards the end of the first postnatal week; then it starts decreasing and disappears towards the end of the 3rd week. In both species AFP is localized in the cytoplasm of nerve cell perikarya and their differentiating processes. Higher concentration of this protein has often been observed at the axonal pole of the cytoplasm of differentiating pyramidal neurons. Immunoglobulin G has been found in non-parenchymal structures, and small amounts also in parenchymal cells of embryonic and early postnatal rats following a pattern of cell-and-tissue distribution similar to that of AFP. In pigs, a low amount of albumin has been shown in differentiating leptomeninges. These data suggest uptake of AFP, and some other serum proteins, from the cerebrospinal fluid into cells of the immature rat and pig brain and its increase (or higher binding) in differentiating neurons.
Assuntos
Encéfalo/embriologia , Ratos Endogâmicos/embriologia , Suínos/embriologia , alfa-Fetoproteínas/análise , Animais , Encéfalo/metabolismo , Diferenciação Celular , Feminino , Imunofluorescência , Imunoglobulina G/análise , Masculino , Neurônios/análise , Neurônios/fisiologia , Gravidez , RatosRESUMO
An immunomodulatory effect of pig alpha-fetoprotein (AFP) on early phytohaemagglutinin (PHA)-stimulated intact thymocytes from piglets was analyzed. The activity of surface-localized Na+, K+-adenosinetriphosphatase was estimated, as the early reaction of mitogen-stimulated T-lymphocytes. A high inhibition of PHA-stimulated enzyme activity was found in the presence of AFP. The reaction was dependent of PHA concentration when using a constant AFP concentration. The highest AFP-induced decrease in PHA-induced Na+, K+-ATPase activity was 72%. AFP was isolated by a new technique based on salting out in ammonium sulphate and acetate buffer solutions. A gentle, simple and rapid procedure makes it possible to obtain AFP in a high degree of purity with preserved biological activity. The presented results can contribute to understanding the molecular aspects of AFP immunoregulation of lymphocyte reactivity in vitro.
Assuntos
Ativação Linfocitária/efeitos dos fármacos , ATPase Trocadora de Sódio-Potássio/metabolismo , Linfócitos T/enzimologia , alfa-Fetoproteínas/farmacologia , Animais , Fito-Hemaglutininas/farmacologia , SuínosRESUMO
Stimulation of oxygen uptake by brain cortex cells with high K+ concentration is one of model reactions characteristic for the functional activity in vitro. The present study was undertaken to clarify the initial findings of the near abolition of K+-induced stimulation of oxygen uptake by mixed brain cortex cells which originate from genetically different mice. Increase in K+-stimulated oxygen consumption by mixed brain cortex cells from H-2 compatible mice (C3H/Cbi/Bom and CBA/J strains) reached 150%. K+-induced oxygen uptake by mixed cells from H-2 incompatible mice (A/Ph and C3H/Cbi/Bom) reached 104%. K+-stimulated respiration by allogeneic cells from congenic lines with H-2 differential loci was also lowered. K+-induced breakdown of ATP and creatine phosphate was similar in both allogeneic cells (to 72% and 64%, respectively) and syngeneic controls (to 72% and 61%, respectively). Factors involved in the demonstration of allogeneic interactions of brain cortex cells were tested. K+-mediated metabolic activity in allogeneic brain cortex cells was dependent on strong antigenic differences, cell-cell contacts and optimal amount of interacting cells. It, therefore, seems that genetic differences such as in H-2 complex linked alloantigens may participate in alterations of some metabolic processes of the brain function.
Assuntos
Córtex Cerebral/metabolismo , Camundongos Endogâmicos/genética , Cloreto de Potássio/farmacologia , Trifosfato de Adenosina/metabolismo , Animais , Células Cultivadas , Córtex Cerebral/citologia , Meios de Cultura , Complexo Principal de Histocompatibilidade , Camundongos , Consumo de Oxigênio/efeitos dos fármacos , Fosfocreatina/metabolismoRESUMO
Male (A/Ph and C3H/Cbi/BOM) mice were used in our experiments. The cell-free supernatants of spleen mixed leucocyte cultures and allogeneic brain cortex cells were collected upon short-term incubation and tested in a comparative study. Spleen and brain low molecular factors were fractionated (ultrafiltration and gel column chromatography) and assayed for their activity in mitogen-induced 3H-thymidine incorporation into spleen lymphocytes. The specific release of the allogeneic spleen fraction (ASF-2) has been observed. A non-specific stimulating effect of ASF-2 on PHA- and LPS-activated spleen lymphocytes was shown. In contrast, a non-specific suppressive effect of brain fraction on PHA- and LPS-stimulated lymphocytes was demonstrated. The opposite effects of spleen and brain factors can be connected with their different composition. The lymphocyte cultures were not affected by spleen or brain factor, when cultured without the addition of mitogen. Positive and negative regulation or modulation of triggering events in lymphocyte activation by these factors can be suggested.
Assuntos
Córtex Cerebral/citologia , Ativação Linfocitária , Teste de Cultura Mista de Linfócitos , Animais , Cromatografia em Gel , DNA/biossíntese , Lipopolissacarídeos/farmacologia , Linfócitos/metabolismo , Masculino , Camundongos , Peso Molecular , Fito-Hemaglutininas/farmacologiaRESUMO
Cell suspensions were prepared form mouse brain cortices. Cells were incubated in a medium also containing 10 mmol.l-1 glucose and either 5 mmol l-1 K+ or 50 mmol.l-1 K+ concentration. In order to standardize individual experiments, the biuret reaction was modified for rapid determination of the protein in cell suspensions. The cellular reserves of energy-rich phosphates were determined in the course of 60 min of cell incubation with 5 mmol.l-1 K+ and following 30 min incubation of cells with 50 mmol.l-1 K+. The level of ATP was significantly elevated after 10-60 min of incubation with low K+, from 0.58 to 0.78 micromoles per 100 mg protein; the creatine phosphate content during the same interval was in the range 1.27-1.44 micromoles per 100 mg protein. A significant decrease of energy reserves in cells was observed if the extracellular concentration of K+ was increased. After 10 and 30 min of incubation, a decrease by 36.2% and 38.5% for creatine phosphate and 34.6% and 44.9% for ATP was found, respectively. Na+ K+-ATPase activity of cells incubated for 60 min with 5 mmol.l-1 K+ was expressed as 4.99 micromoles of liberated Pi per 100 mg protein.1 h. Enzyme activity was stimulated with 50 mmol.l-1 K+ by 24.3% and 25.7% after 10 and 30 min of cell incubation respectively. Stimulation of Na+ K+-ATPase activity of brain cortex cells was directly dependent on the actual presence of stimulating 50 mmol.l-1 K+ concentration.