RESUMEN
Concanavalin A (Con A)-induced hepatitis is a mouse model of acute autoimmune hepatitis. The aim of this study was to investigate the role of hepatic dendritic cells (DC) in the immune modulation of tissue damage. Almost all hepatic DC were plasmacytoid DC (CD11c+ I-A(low) B220+); however, conventional DC were CD11c+ I-A(high) B220(-). At an early stage (3-6 h) after Con A administration, the number of DC in both the liver and spleen decreased, increasing thereafter (12-24 h) in parallel with hepatic failure. The hepatic CD11c+ DC population contained many CD11b(-) cells, while the majority of splenic CD11c+ DC were CD11b+. After Con A administration, the proportion of I-A+ and CD11b+ cells within the CD11c+ DC population tended to increase in the liver, but not in the spleen. Similarly, expression of the activation markers CD80, CD86 and CD40 by CD11c+ DC increased in the liver, but not in the spleen. Next, adoptive transfer of DC isolated from the liver and spleen was performed 3 h after Con A administration to examine the immunomodulatory function of DC. Only hepatic DC had the ability to suppress hepatic failure. Analysis of cytokine production and subsequent identification of the effector cells showed that hepatic DC achieved this by suppressing the production of interleukin (IL)-12 and IL-2, rather than modulating effector cell function.
Asunto(s)
Células Dendríticas/inmunología , Hepatitis Autoinmune/inmunología , Traslado Adoptivo , Animales , Antígeno B7-1/biosíntesis , Antígeno B7-2/biosíntesis , Antígeno CD11b/biosíntesis , Antígeno CD11b/inmunología , Antígeno CD11c/biosíntesis , Antígeno CD11c/inmunología , Antígenos CD40/biosíntesis , Concanavalina A/farmacología , Células Dendríticas/metabolismo , Citometría de Flujo , Hepatitis Autoinmune/metabolismo , Hepatitis Autoinmune/patología , Interleucina-12/biosíntesis , Interleucina-2/biosíntesis , Hígado/inmunología , Hígado/patología , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Bazo/inmunología , Linfocitos T Reguladores/inmunologíaRESUMEN
Primitive blood cells differentiate from the ventral mesoderm blood islands in Xenopus embryos. In order to determine the tissue interactions that propagate blood formation in early embryogenesis, we used embryos that had the ventral cytoplasm removed. These embryos gastrulated normally, formed a mesodermal layer and lacked axial structures, but displayed a marked enhancement of alpha-globin expression. Early ventral markers, such as msx-1, vent-1 and vent-2 were highly expressed at the gastrula stage, while a dorsal marker, goosecoid, was diminished. Several lines of experimental evidence demonstrate the critical role of animal pole-derived ectoderm in blood cell formation: 1) Mesoderm derived from dorsal blastomeres injected with beta-galactosidase mRNA (as a lineage tracer) expressed alpha-globin when interfaced with an animal pole-derived ectodermal layer; 2) Embryos in which the animal pole tissue had been removed by dissection at the blastula stage failed to express alpha-globin; 3) Exogastrulated embryos that lacked an interaction between the mesodermal and ectodermal layers failed to form blood cells, while muscle cells were observed in these embryos. Using dominant-negative forms of the BMP-4 and ALK-4 receptors, we showed that activin and BMP-4 signaling is necessary for blood cell differentiation in ventral marginal zone explants, while FGF signaling is not essential. In ventralized embryos, inactivation of the BMP-4 signal within a localized area of the ectoderm led to suppression of globin expression in the adjacent mesoderm layer, but inactivation of the activin signal did not have this effect. These observations suggest that mesodermal cells, derived from a default pathway that is induced by the activin signal, need an additional BMP-4-dependent factor from the overlying ectoderm for further differentiation into a blood cell lineage.
Asunto(s)
Diferenciación Celular , Eritrocitos/citología , Eritropoyesis/fisiología , Estratos Germinativos/metabolismo , Transducción de Señal , Xenopus laevis/embriología , Receptores de Activinas Tipo I , Activinas , Animales , Northern Blotting , Proteína Morfogenética Ósea 4 , Proteínas Morfogenéticas Óseas/metabolismo , Embrión no Mamífero , Eritrocitos/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Globinas/metabolismo , Hibridación in Situ , Inhibinas/metabolismo , Morfogénesis , ARN Mensajero/análisis , Receptores de Factores de Crecimiento/metabolismo , Proteínas de Xenopus , Xenopus laevis/metabolismoRESUMEN
Pan-cadherin antibodies recognize the conserved C-terminal region of the family of cell-cell adhesion molecules, cadherins, and have a broad spectrum of reactivity to the molecules. In the present study, by immunohistochemistry using an anti-pan cadherin monoclonal antibody (mAb), expression dynamics of cadherins in epidermal tissues were analyzed during metamorphosis of Xenopus laevis. At early stages of development, the anti-pan cadherin mAb detected signals at cell-cell boundaries and in the cytoplasm of both trunk and tail epidermal cells. During metamorphosis, the immunoreactivity decreased in the trunk skin tissue but remained in the tail. At the climax stage, immunoreactivity was observed only in the regressing tail epidermis. The signals disappeared completely from the trunk epidermis, which had already transformed into adult-type tissue. This observation was confirmed by western blot analysis. A specific band was detected in the larval skin, but not in the adult lysate, at approximately 135 kDa in molecular size, corresponding to the molecular mass of cadherins. This different immunoreactivity in larvae and adults was observed in the epidermis of the skin, but not in any other tissues examined, that is, brain, kidney and liver. The immunoreactivity seen in larval epidermal cells was drastically downregulated by thyroid hormone treatment in vitro. These changes of immunoreactivity were specific for the C-terminal region of cadherins, suggesting intracellular alteration of the molecules during metamorphosis, and the anti-pan cadherin mAb can be a marker for larval-type epidermal cells that is applicable to analysis of Xenopus metamorphosis.
Asunto(s)
Anticuerpos Monoclonales/metabolismo , Cadherinas/inmunología , Epidermis/metabolismo , Metamorfosis Biológica/fisiología , Xenopus laevis/metabolismo , Animales , Western Blotting , Células Cultivadas , Regulación hacia Abajo , Células Epidérmicas , Técnica del Anticuerpo Fluorescente , Inmunohistoquímica , Larva/fisiologíaRESUMEN
During anuran metamorphosis, larval cells of the tadpole are completely eliminated and replaced by adult cells in the corresponding tissues of the frog for the adaptation to terrestrial life from an aquatic life. Before the metamorphic climax, most of the cells have already transformed from larval cells into adult-type cells, but the tail cells remain as larval cells even at the climax stages of metamorphosis. In our previous works, we demonstrated that larval skin grafts are rejected by an inbred strain of adult Xenopus and that the larval cells are recognized and made apoptotic by splenocytes obtained from adults and/or metamorphosing tadpoles in vitro (Y. Izutsu and K. Yoshizato, 1993, J. Exp. Zool. 266, 163-167; Y. Izutsu et al., 1996, Differentiation 60, 277-286). In the present study, it was found that there were two types of larval epidermal cells, classified according to the presence of major histocompatibility complex (MHC); one is the apical cell expressing both MHC classes I and II, and the other is the skein cell, which expresses no MHC. By a Percoll gradient, we were able to separate these two types of cells and examined the proliferative response of adult T cells to each of them. It was revealed that the apical cells (MHC-positive) were recognized directly by adult splenic T cells, whereas the skein cells (MHC-negative) were recognized by the T cells via the antigen presentation by adult splenocytes. Both of these proliferative responses were restricted to MHC class II. This is the first report showing how the larval-specific antigens present in different forms in epidermal cells are recognized as immunological targets by syngeneic adult T lymphocytes.
Asunto(s)
Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Bazo/inmunología , Linfocitos T/inmunología , Xenopus laevis/genética , Xenopus laevis/inmunología , Animales , Cruzamientos Genéticos , Epidermis/inmunología , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase II/genética , Endogamia , Larva/inmunología , Activación de Linfocitos , Complejo Mayor de Histocompatibilidad , Metamorfosis Biológica , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Xenopus/genética , Xenopus laevis/crecimiento & desarrolloRESUMEN
Larval cells of the anuran tadpole are replaced by adult cells in the corresponding tissues of the frog during metamorphosis; tissues of the tail, which have no counterpart in the adult, are completely eliminated during metamorphosis. We have previously demonstrated that young adults of the major histocompatibility complex (MHC) homozygous inbred J strain of Xenopus laevis reject skin grafts from larvae of the same strain, showing that there is histoincompatibility between larval and adult skin tissues [19]. Thus, we postulated that some immunological recognition might be involved in this specific elimination of the tail tissue and set out to test the idea. Using in vitro assays, we detected a significant increase in proliferation of splenocytes derived from adult and metamorphic climax animals co-cultured with larval tail tissue. This response was shown to be thymus-dependent. However, the degeneration of larval tissues was not observed in such co-cultures under our standard culture conditions. To detect the possible cytotoxicity of splenocytes, the culture conditions were modified by supplementing with 10% heat-inactivated adult Xenopus serum instead of 10% fetal calf serum (FCS). After this modification, the degeneration of larval tissues was observable macroscopically and microscopically with co-cultured adult splenocytes, but not tadpole ones. The nuclear fragmentation of the epidermal cells was seen by light and electron microscopy. Apoptosis was evidenced by the demonstration of the "ladder pattern" upon electrophoresis of genomic DNA obtained from the degenerating larval tissues. Surprisingly, this response was thymus independent. Moreover, it was shown that this response was not observed when the larval tissues were cultured with adult thymocytes or adult epidermal cells. In vivo, migration of T cells into the epidermis of tail tissues at the late climax of metamorphosis was demonstrated immunohistochemically using a monoclonal antibody against Xenopus T cells, even in the early thymectomized tadpoles. Considering these results, we propose that populations of adult-type non-T leukocytes might participate in the specific detection and elimination of larval type cells during metamorphosis.
Asunto(s)
Larva/citología , Complejo Mayor de Histocompatibilidad/fisiología , Xenopus laevis/fisiología , Factores de Edad , Animales , Apoptosis/fisiología , División Celular , Núcleo Celular , Células Cultivadas , Técnicas de Cocultivo , Técnicas de Cultivo/métodos , ADN/fisiología , Células Epidérmicas , Reacción Injerto-Huésped , Leucocitos/citología , Metamorfosis Biológica , Bazo/citología , Cola (estructura animal)/citología , Timo/citología , Xenopus laevis/crecimiento & desarrollo , Xenopus laevis/inmunologíaRESUMEN
Larval cells in tissues of the anuran tadpole are replaced by adult cells in the corresponding tissue of the frog during metamorphosis. As an extreme example of such replacement, tissues of the tail, which have no counterpart in the adult, are completely eliminated during metamorphosis. We postulated that some immunological recognition mechanism might be involved in this specific elimination of the tail tissue. Our working hypothesis was tested by applying the skin transplantation technique to individuals of the inbred J strain of Xenopus laevis, which accept homografts. We demonstrated that young adults reject skin grafts from larvae. This rejection was immunological in nature because the secondary response of rejection was observed. There was a clear difference in graft rejection between grafts from the tail and those from the body. Grafts of tail skin were rejected irrespective of the metamorphic stages of donors. By contrast, grafts of body skin became acceptable as donors metamorphosed. The mean survival time of the larval skin was much longer than that of major histocompatibility complex (MHC)-disparate skin grafts reported by other investigators, suggesting that the rejection described in the present study is due to disparity in minor histocompatible (minor-H) antigens. We propose the involvement of the immunological recognition mechanism in the process of specific detection and elimination of larval cells in the tail.
Asunto(s)
Tolerancia Inmunológica , Metamorfosis Biológica/fisiología , Piel/embriología , Animales , Embrión no Mamífero/inmunología , Rechazo de Injerto , Piel/inmunología , Trasplante de Piel , Xenopus laevisRESUMEN
The City University colour vision test (CUCVT) was used for the examination of 158 subjects suffering from congenital colour vision defects (36 protanopes, 122 deutanopes) and its results were compared with that of an anomaloscope and of the panel D-15. 23% of the subjects classified as protanopes and 98% of the subjects classified as deuteranopes by means of the anomaloscope were also classified as such by means of the CUCVT, while 93% of the subjects classified as protanomalous and 90% of the subjects classified as deuteranomalous by means of the anomaloscope gave normal answers at the CUCVT. The results of the CUCVT were almost the same as with panel D-15 except protanopia. The colour spots of each plate of the CUCVT were plotted on a CIE chromaticity diagram and the results of this study are also reported.