RESUMEN
Commensal microbiota-specific T helper type 17 (Th17) cells are enriched in the intestines, which can convert into T follicular helper (Tfh) in Peyer's patches, and are crucial for production of intestinal immunoglobulin A (IgA) against microbiota; however, the role of Th17 and Tfh cytokines in regulating the mucosal IgA response to enteric microbiota is still not completely known. In this study, we found that intestinal IgA was impaired in mice deficient in interleukin (IL)-17 or IL-21 signaling. IL-21, but not IL-17, is able to augment B-cell differentiation to IgA(+) cells as mediated by transforming growth factor ß1 (TGFß1) and accelerate IgA class switch recombination (CSR). IL-21 and retinoic acid (RA) induce IgA(+) B-cell development and IgA production and drives autocrine TGFß1 production to initiate IgA CSR. Repletion of T-cell-deficient TCRßxδ(-/-) mice with Th17 cells specific for commensal bacterial antigen increased the levels of IgA(+) B cells and IgA production in the intestine, which was blocked by neutralizing IL-21. Thus IL-21 functions to strongly augment IgA production under intestinal environment. Furthermore, IL-21 promotes intestinal B-cell homing through α4ß7 expression, alone or with TGFß and RA. Together, IL-21 from microbiota-specific Th17 and/or Tfh cells contributes to robust intestinal IgA levels by enhancing IgA(+) CSR, IgA production and B-cell trafficking into the intestine.
Asunto(s)
Microbioma Gastrointestinal/inmunología , Inmunidad Mucosa/fisiología , Inmunoglobulina A/inmunología , Interleucinas/inmunología , Mucosa Intestinal/inmunología , Animales , Linfocitos B/inmunología , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Movimiento Celular/genética , Movimiento Celular/inmunología , Inmunoglobulina A/genética , Interleucina-17/genética , Interleucina-17/inmunología , Interleucinas/genética , Mucosa Intestinal/microbiología , Ratones , Ratones Noqueados , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Células Th17/inmunología , Factor de Crecimiento Transformador beta1/genética , Factor de Crecimiento Transformador beta1/inmunologíaRESUMEN
The purpose of the present study was to investigate properties and mechanism of action of the synthetic adrenocorticotropin (ACTH)-like peptide VKKPGSSVKV, corresponding to the sequence 11-20 of the variable part of human immunoglobulin G1 (IgG1) heavy chain. The ACTH-like peptide was shown to act as an immunosuppressive agent in vitro: it inhibits the blast transformation of mouse thymocytes and reduces the spontaneous motility of mouse peritoneal macrophages as well as their bactericidal activity against Salmonella typhimurium 415 virulent strain bacteria. High affinity receptors for the ACTH-like peptide were found on thymocytes and macrophages and shown to be at the same time the receptors for ACTH. The kinetic characteristics of the ACTH-like peptide and 125I-labeled ACTH (13-24) (ACTH 'address segment') specific binding to the receptors were determined. It was found that the ACTH-like peptide binding to the receptors on target cells is accompanied by an increase in both adenylate cyclase activity and intracellular cAMP content.
Asunto(s)
Hormona Adrenocorticotrópica/farmacología , Inmunoglobulina G/farmacología , Activación de Linfocitos/inmunología , Activación de Macrófagos/inmunología , Macrófagos Peritoneales/inmunología , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/farmacología , Linfocitos T/inmunología , Hormona Adrenocorticotrópica/inmunología , Animales , Humanos , Inmunoglobulina G/inmunología , Activación de Linfocitos/efectos de los fármacos , Activación de Macrófagos/efectos de los fármacos , Macrófagos Peritoneales/efectos de los fármacos , Ratones , Ratones Endogámicos CBA , Fragmentos de Péptidos/síntesis química , Biblioteca de Péptidos , Linfocitos T/efectos de los fármacosRESUMEN
Influence of the ACTH-like peptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH corresponding to the sequence 11-20 of the variable part of human immunoglobulin G1 (IgG1) heavy chain on growth of MT-4 human T-lymphoblastoid cell line was investigated. It was found that the ACTH-like peptide at concentration range 10(-11) -10(-7) M inhibits the proliferation of MT-4 cells. Labeled ACTH 'address segment' [(125)I]ACTH (13-24) was used to establish that MT-4 cells express specific receptors for ACTH (K(d) = 97 pM). The ACTH-like peptide and human ACTH (but not IgG1 heavy chain) were shown to compete with [(125)I]ACTH (13-24) for binding to these receptors (K(i1) = 0.38 nM and K(i2) = 0.34 nM).
Asunto(s)
Hormona Adrenocorticotrópica/química , División Celular/efectos de los fármacos , Inmunoglobulina G/química , Inmunoglobulina G/farmacología , Fragmentos de Péptidos/farmacología , Hormona Adrenocorticotrópica/farmacocinética , Secuencia de Aminoácidos , Unión Competitiva , Línea Celular , Humanos , Cadenas Pesadas de Inmunoglobulina/química , Cadenas Pesadas de Inmunoglobulina/farmacología , Cinética , Fragmentos de Péptidos/química , Fragmentos de Péptidos/farmacocinética , Receptores de Corticotropina/fisiología , Linfocitos T/efectos de los fármacosRESUMEN
The antiproliferative and immunosuppressive in vitro effects of immunocortin, a synthetic adrenocorticotropin-like (ACTH-like) decapeptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH, whose sequence corresponds to segment 11-20 of the variable part of the human IgG1 heavy chain, were studied. At concentrations of 10(-11)-10(-7) M, immunocortin was found to inhibit the growth of the human MT-4 T-lymphoblastoid cell line, to suppress the blast transformation of thymocytes, and to decrease the spontaneous mobility of peritoneal macrophages and their bactericidal action toward the virulent strain Salmonella typhimurium 415. By using a 125I-labeled "addressing" fragment of ACTH ¿[125I]ACTH-(13-24)¿, we showed that MT-4 cells express specific receptors for ACTH (Kd 97 pM). Immunocortin and human ACTH (but not the heavy chain of IgG1) competitively inhibited the binding of [125I]ACTH-(13-24) to these receptors with Ki1 of 0.38 and Ki2 of 0.34 nM, respectively. Specific receptors for ACTH (Kd 5.8 nM) on mouse thymocytes were detected and characterized. The unlabeled immunocortin was shown to complete with labeled ACTH-(13-24) for binding to these receptors (Ki = 1.8 nM) and this binding of immunocortin to receptors on thymocytes activates adenylate cyclase from these cells and increases the intracellular concentration of cAMP.
Asunto(s)
Hormona Adrenocorticotrópica/genética , Inmunoglobulina G/genética , Inmunoglobulina G/metabolismo , Oligopéptidos , Hormona Adrenocorticotrópica/metabolismo , Animales , Humanos , Activación de Macrófagos , Macrófagos/metabolismo , Ratones , Oligopéptidos/síntesis química , Oligopéptidos/química , Oligopéptidos/genética , Oligopéptidos/metabolismo , Receptores de Corticotropina/metabolismo , Salmonella typhimuriumRESUMEN
The synthetic ACTH-like decapeptide H-Val-Lys-Lys-Pro-Gly- Ser-Ser-Val-Lys-Val-OH, corresponding to amino acid residues 11-20 of the variable part of the human IgG1 heavy chain (referred to as immunocortin) was found to have an immunosuppressive effect on cells in vitro: it inhibits blast transformation of mouse thymocytes and reduces spontaneous motility of mouse peritoneal macrophages as well as their bactericidal activity against the virulent bacterial strain Salmonella typhimurium 415. Tritium-labeled immunocortin binds with high affinity to ACTH receptors on thymocytes and macrophages (Kd 2. 1 and 2.5 nM, respectively) and activates adenylate cyclase in these cells. Thus, the interaction of immunocortin with the target cell includes the following main steps: binding to the receptor, activation of adenylate cyclase, and elevation of the intracellular content of cAMP.
Asunto(s)
Hormona Adrenocorticotrópica/química , Inmunoglobulina G/química , Inmunoglobulina G/farmacología , Cadenas Pesadas de Inmunoglobulina/farmacología , Inmunosupresores/farmacología , Fragmentos de Péptidos/farmacología , Adenilil Ciclasas/metabolismo , Secuencia de Aminoácidos , Animales , Células Cultivadas , AMP Cíclico/metabolismo , Humanos , Cadenas Pesadas de Inmunoglobulina/química , Inmunosupresores/química , Activación de Linfocitos/efectos de los fármacos , Macrófagos Peritoneales/efectos de los fármacos , Macrófagos Peritoneales/inmunología , Ratones , Ratones Endogámicos CBA , Fragmentos de Péptidos/química , Ratas , Timo/citología , Timo/efectos de los fármacos , Timo/inmunologíaRESUMEN
The effects of L-glutamic acid on the differentiation of HL-60 and K-562 cell lines and on the expression level of mRNAs encoding IL-1 beta, IL-6, and TNF alpha were studied. It was shown that TNF alpha actively induces the differentiation of these cell lines, whereas L-glutamic acid enhances its effect. Our results indicated that L-glutamic acid modulates the physiological state of the myeloid cell line in blood, in particular, by affecting both the reception and expression of cytokines functionally important for these cells.
Asunto(s)
Células de la Médula Ósea/efectos de los fármacos , Ácido Glutámico/farmacología , Células de la Médula Ósea/metabolismo , Diferenciación Celular/efectos de los fármacos , Células HL-60 , Humanos , Interleucina-1/genética , Interleucina-6/genética , Células K562 , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
A specific interaction of L-glutamic acid with promyelocytic leukemia HL-60 cells completely differentiated by all-E-retinoic acid (Kd = 0.5 microM) and by plasma membrane fraction from the same cells (Kd = 1 microM) was detected through radioligand analysis and characterized. Quisqualate, a nonlabeled structural analogue of glutamic acid, was found to inhibit competitively the specific binding of L-[3H]glutamic acid to the membranes (Ki = 0.24 microM). The stereospecificity of the binding was demonstrated. These data suggest that specific glutamate receptors appear on the surface of HL-60 cells during their differentiation.
Asunto(s)
Receptores de Glutamato/biosíntesis , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Fraccionamiento Celular , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Ácido Glutámico/metabolismo , Células HL-60 , Humanos , Ensayo de Unión Radioligante , Receptores de Glutamato/metabolismo , Tretinoina/farmacologíaRESUMEN
To elucidate the mechanism of differentiating activity on L-Glu to HL-60 cells, its influence on binding of human recombinant interleukine-1beta (rHuIL-1beta), tumour necrosis factor-alpha (rHuTNF-alpha) and interleukine-6 (IL-6) by HL-60 cells was studied. It was established, that L-Glu converted the high affinity binding of [125I]rHuIL-1beta (Kd = 0.32 nM) and [125I]rHuIL-6 (Kd = 0.075 nM) to HL-60 cells into low affinity (corresponding Kd values -13.3 and 2.1 nM) at concentration 0.1 microM. The preincubation for an hour of HL-60 cells with 0.1 microM L-Glu was shown to result in an increase of affinity and number of [125I]rHuTNF-alpha binding sites. Thus, L-Glu decreases the sensitivity of the HL-60 cells to IL-1beta and IL-6 and increases TNF-alpha binding at concentration 0.1 microM.
Asunto(s)
Ácido Glutámico/metabolismo , Interleucina-1/metabolismo , Interleucina-6/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Ácido Glutámico/farmacología , Células HL-60 , Humanos , Interleucina-1/farmacología , Interleucina-6/farmacología , Radioisótopos de Yodo , Marcaje Isotópico , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacología , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
L-Glutamic acid at a concentration of 0.1 microM was found to induce differentiation of the cell line of HL-60 promyelocytic leukemia into granulocytes or neutrophils. The HL-60 cells have no specific glutamate-binding sites, but L-glutamic acid influences the reception of several cytokines by these cells. At a concentration of 0.1 microM, L-glutamic acid completely inhibits the high-affinity binding of 125I-labeled human recombinant interleukin-1 beta (Kd = 0.32 nM) to the HL-60 cells, but does not affect their low-affinity binding (Kd = 13.3 nM) and does not change the total number of the IL-1 beta-binding sites. Preincubation of the HL-60 cells with 0.1 microM of L-glutamic acid increases 2.5 times the number of receptors for 125I-labeled human recombinant tumor necrosis factor beta. These results suggest that L-glutamic acid plays an important role in the differentiation of the blood myeloid cells.
Asunto(s)
Ácido Glutámico/farmacología , Granulocitos/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Receptores de Citocinas/metabolismo , Unión Competitiva , Diferenciación Celular/efectos de los fármacos , Ácido Glutámico/metabolismo , Granulocitos/citología , Células HL-60 , Humanos , Interleucina-1/metabolismo , Radioisótopos de Yodo , Linfotoxina-alfa/metabolismo , Neutrófilos/citología , Receptores de Citocinas/efectos de los fármacos , Proteínas Recombinantes/metabolismo , Espectrometría de Masa Bombardeada por Átomos VelocesRESUMEN
The presence of specific glutamate receptors on T lymphocytes separated from normal human blood was investigated. Upon study of T lymphocytes interaction with [3H]glutamate the last one was found to bind specifically to these cells, Kd = 2.36 x 10(-7) M. Thirteen unlabelled compounds were tested as potential glutamate competitors. It was shown that only quisqualate (a structural analogue of glutamate) and dipeptides Ala-Glu, Glu-Ala, Glu-Glu competed with [3H]glutamate in binding to a common binding site. The experiments on binding of labelled and unlabelled glutamate bulky conjugates with dextran have demonstrated that both conjugated and free glutamate interacted with the common binding sites on the outer cell membrane. By this means it was shown that T lymphocytes exposed quisqualate-sensitive glutamate receptors on the outer surface of the plasma membrane.
Asunto(s)
Ácido Glutámico/metabolismo , Receptores de Glutamato/metabolismo , Linfocitos T/metabolismo , Unión Competitiva , Dextranos , Humanos , TripsinaRESUMEN
A specific interaction of [3H]Glu with T lymphocytes from the blood of healthy donors (Kd = 0.236 microM) was revealed and described. It was found that unlabeled quisqualate, a structural analogue of L-glutamic acid, and unlabeled dipeptides Ala-Glu, Glu-Ala, and Glu-Glu competitively inhibit the specific binding of [3H]Glu to T lymphocytes (with Ki 0.19, 2.4, 3.4, and 1.2 microM, respectively). Binding experiments with conjugates of labeled and unlabeled glutamic acid with dextran showed that the receptors of [3H]Glu are localized on the outer surface of the plasma membrane of T lymphocytes.
Asunto(s)
Agonistas de Aminoácidos Excitadores/farmacología , Ácido Glutámico/metabolismo , Ácido Quiscuálico/farmacología , Receptores de Glutamato/metabolismo , Linfocitos T/metabolismo , Unión Competitiva/efectos de los fármacos , Membrana Celular/metabolismo , Dipéptidos/farmacología , Ácido Glutámico/sangre , Humanos , Estereoisomerismo , Linfocitos T/efectos de los fármacosRESUMEN
The mechanisms of action of total saponins from Panax Ginseng C.A. Meyer on the elements of intracellular signalling system in Ehrlich ascites tumor cells were studied. The action of total saponins was compared with the effect of ATP, a classical activator of these cells. Saponins at concentrations of 10(-6)-10(-3)% increased [Ca2+]i, mobilized Ca2+ ions from the endoplasmic reticulum (ER) and activated the influx of Ca2+ to cells. Like ATP, saponins activated the Na+/H+ exchange and Ca(2+)-dependent K+ channels. Of all the parameters, only the activation of Ca2+ influx in cell is directly affected by saponins. The changes in other parameters are connected with nonspecific activation of purinoreceptors. The analysis of the kinetic data suggests that, as distinct from ATP-dependent activation of purinoreceptor, saponins first activate the Ca2+ influx to cells and only then induce the mobilization of Ca2+ from ER.
Asunto(s)
Calcio/metabolismo , Carcinoma de Ehrlich/metabolismo , Panax/química , Plantas Medicinales , Agonistas Purinérgicos , Saponinas/farmacología , Transducción de Señal/efectos de los fármacos , Animales , Agonistas de los Canales de Calcio/farmacología , Carcinoma de Ehrlich/patología , Retículo Endoplásmico/efectos de los fármacos , Ratones , Intercambiadores de Sodio-Hidrógeno/metabolismo , Células Tumorales CultivadasRESUMEN
The influence of the dammarane triterpene glycosides Rb1, Rb2, Rg1 and Rf and their aglycones 20(S) protopanaxatriol and 20(S) protopanaxadiol on fluctuation of the K+ and H+ flows in erythrocytes induced by ionophore A23187 and Ca2+ was studied. It was shown that glycoside Rb1 (10 to 50 micrograms/ml) and 20(S) protopanaxatriol (30 micrograms/ml) had a property of inducing fluctuation of the K+ and H+ flows in erythrocytes. In case of Rb1 the system entirely reversed to the initial state whereas in case of the protopanaxatriol use after the fluctuation the system did not reverse to the initial state. Glycosides Rb2 (10 to 30 micrograms/ml), Rg1 (5 to 20 micrograms/ml) and Rf (10 to 100 micrograms/ml) and 20(S) protopanaxadiol (30 micrograms/ml) in the concentrations used did not initiate the fluctuations. The above substances increased the extracellular concentration of K+ and pH without the system reversion to the initial state.
Asunto(s)
Medicamentos Herbarios Chinos , Eritrocitos/efectos de los fármacos , Glicósidos/farmacología , Sapogeninas/farmacología , Triterpenos/farmacología , Animales , Calcimicina/farmacología , Eritrocitos/metabolismo , Ginsenósidos , Canales Iónicos/efectos de los fármacos , Ionóforos/farmacología , Panax/química , Plantas Medicinales , Canales de Potasio/efectos de los fármacos , Protones , Ratas , Ratas Wistar , Saponinas/farmacologíaRESUMEN
Molecular mechanisms of action of the total fraction of saponins from Red Korean Ginseng on the elements of the intracellular signalling system of the cells of ascitic Ehrlich carcinoma (AEC) were studied. The action of the total fraction of the saponins on the AEC cells was compared with that of the classic activator of such cells i.e. ATP. It was shown that the action of the total fraction of the saponins was similar to that of ATP. In concentrations of 10(-6) to 10(-3) per cent saponin induced an increase of [Ca2+]j mobilizing Ca2+ from the endoplasmic reticulum (ER) and activating the Ca2+ inlet to the cells. The same as in case with the use of ATP the Ca2+ mobilization from ER was reversible. In comparison to ATP saponin induced higher activation of the Ca2+ inlet to ER and the cells. The same as ATP saponin activated the Na+/H+ exchange and the Ca2+ - dependent K+-channels. Out of all the mention-ed parameters only the activation of the Ca2+ inlet to the cells was probably the direct result of the saponin action. The changes in the other parameters were mediated by nonspecific activation of the purine receptor. The analysis of the kinetic data demonstrated that unlike the ATP-dependent activation of the purine receptor the saponins first of all activated the inlet of Ca2+ to the cells and only after that the mobilization of the latter from ER.