RESUMEN
The secreted frizzled-related proteins (sfrp) and smoothened (smo) genes and their possible role in the regeneration of internal organs in the holothurian Eupentacta fraudatrix were studied. In this species, two sfrp genes were identified: sfrp1/2/5, sfrp3/4 and one smo gene. Their expression was analysed during regeneration of the aquapharyngeal bulb (AB) and intestine, and these genes were knock down by RNA interference. It has been shown that the expression of these genes is extremely important for the formation of AB. In all animals subjected to knockdown, at 7 days after evisceration, a full-sized AB rudiment was not formed. As a result of sfrp1/2/5 knockdown, the process of extracellular matrix remodelling in AB is interrupted, that leading to clusters of dense connective tissue formation, which slows down cell migration. When sfrp3/4 is knockdown, the connective tissue of AB anlage is completely disrupted and its symmetry is broken. The effect of smo knockdown was expressed in a significant impairment of AB regeneration, when connections between ambulacras were not formed after evisceration. However, despite severe disturbances in AB regeneration, a normal-sized gut anlage developed in all cases, which suggests that the regeneration of the digestive tube and AB occur independently of each other.
Asunto(s)
Pepinos de Mar , Proteínas Relacionadas con Frizzled Secretadas , Animales , Cicatrización de Heridas/genéticaRESUMEN
Several genes of the Wnt and Frizzled families in the holothurian Eupentacta fraudatrix are characterized, and the complete coding sequences of wntA, wnt4, wnt6, wnt16, frizzled1/2/7, frizzled4, and frizzled5/8 are obtained. The dynamics of expression of these genes during regeneration of internal organs after evisceration are studied. Evisceration and the associated damages supposedly induce the expression of wnt16 on third day after evisceration. Genes wntA, wnt4, wnt6, and frizzled1/2/7 up-regulate during the period of active morphogenesis (5-7 days after evisceration) and might participate in regulation of tissue and organ formation. The signaling induced via Frizzled5/8 is could be necessary for formation of the anterior (ectodermal) part of the digestive system and development of the calcareous ring on 10th day after evisceration. Our data suggest that the Wnt signaling pathway plays a significant role in the regulation of regeneration of internal organs in holothurians.
Asunto(s)
Receptores Frizzled/genética , Regulación de la Expresión Génica , Holothuria/fisiología , ARN/genética , Regeneración/genética , Proteínas Wnt/genética , Animales , Receptores Frizzled/biosíntesis , Modelos Animales , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Wnt/biosíntesis , Vía de Señalización WntRESUMEN
Traumatic brain injuries (TBI) of varying severity are becoming more frequent all over the world. The process of neuroinflammation, in which macrophages and microglia are key players, underlies all types of brain damage. The present study focuses on evaluating the therapeutic potential of N-docosahexaenoylethanolamine (DHEA, synaptamide), which is an endogenous metabolite of docosahexaenoic acid in traumatic brain injury. Previously, several in vitro and in vivo models have shown significant anti-neuroinflammatory and synaptogenic activity of synaptamide. The results of the present study show that synaptamide by subcutaneous administration (10 mg/kg/day, 7 days) exerts anti-inflammatory and anti-apoptotic effects in the thalamus and cerebral cortex of experimental animals (male C57BL/6 mice). Were analyzed the dynamics of changes in the activity of Iba-1- and CD68-positive microglia/macrophages, the level of production of pro-inflammatory cytokines (IL1ß, IL6, TNFα) and pro-apoptotic proteins (Bad, Bax), the expression of pro- and anti-inflammatory markers (CD68, CD206, arg-1). ATF3 transcription factor distribution and neuronal state in the thalamus and cerebral cortex of animals with craniotomy, traumatic brain injury, and therapy are quantitatively assessed. The obtained data showed that synaptamide: (1) has no effect on the total pool of microglia/macrophages; (2) inhibits the activity of pro-inflammatory microglia/macrophages and cytokines they produce; (3) increases the expression of CD206 but not arg-1; (4) has anti-apoptotic effect and (5) improves the morphological state of neurons. The results obtained confirm the high therapeutic potential of synaptamide in the therapy of traumatic brain injury.
Asunto(s)
Apoptosis , Lesiones Traumáticas del Encéfalo , Corteza Cerebral , Ratones Endogámicos C57BL , Microglía , Neuronas , Animales , Lesiones Traumáticas del Encéfalo/tratamiento farmacológico , Lesiones Traumáticas del Encéfalo/patología , Lesiones Traumáticas del Encéfalo/metabolismo , Masculino , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Apoptosis/efectos de los fármacos , Ratones , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Corteza Cerebral/patología , Microglía/efectos de los fármacos , Microglía/metabolismo , Microglía/patología , Antiinflamatorios/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Tálamo/efectos de los fármacos , Tálamo/metabolismo , Tálamo/patología , Citocinas/metabolismo , Etanolaminas/farmacología , Enfermedades Neuroinflamatorias/tratamiento farmacológico , Enfermedades Neuroinflamatorias/metabolismoRESUMEN
Mesodermal cells of holothurian Eupentacta fraudatrix can transdifferentiate into enterocytes during the regeneration of the digestive system. In this study, we investigated the expression of several genes involved in gut regeneration in E. fraudatrix. Moreover, the localization of progenitor cells of coelomocytes, juvenile cells, and their participation in the formation of the luminal epithelium of the digestive tube were studied. It was shown that Piwi-positive cells were not involved in the formation of the luminal epithelium of the digestive tube. Ef-72 kDa type IV collagenase and Ef-MMP16 had an individual expression profile and possibly different functions. The Ef-tensilin3 gene exhibited the highest expression and indicates its potential role in regeneration. Ef-Sox9/10 and Ef-Sox17 in E. fraudatrix may participate in the mechanism of transdifferentiation of coelomic epithelial cells. Their transcripts mark the cells that plunge into the connective tissue of the gut anlage and give rise to enterocytes. Ef-Sox9/10 probably controls the switching of mesodermal cells to the enterocyte phenotype, while Ef-Sox17 may be involved in the regulation of the initial stages of transdifferentiation.
Asunto(s)
Sistema Digestivo/crecimiento & desarrollo , Tracto Gastrointestinal/crecimiento & desarrollo , Regeneración/genética , Pepinos de Mar/genética , Animales , Transdiferenciación Celular/genética , Sistema Digestivo/metabolismo , Células Epiteliales/metabolismo , Tracto Gastrointestinal/metabolismo , Regulación del Desarrollo de la Expresión Génica/genética , Metaloproteinasas de la Matriz/genética , Mesodermo/crecimiento & desarrollo , Mesodermo/metabolismo , ARN Interferente Pequeño/genética , Factores de Transcripción SOX/genética , Pepinos de Mar/crecimiento & desarrollo , Inhibidores Tisulares de Metaloproteinasas/genéticaRESUMEN
The holothurian Eupentacta fraudatrix is a unique organism for studying regeneration mechanisms. Moreover, E. fraudatrix can quickly restore parts of its body and entire organ systems, yet at the moment, there is no data on the participation of stem cells in the process. To the contrary, it has been repeatedly confirmed that this process is only due to the transformation of terminally differentiated cells. In this study, we examine changes in gene expression during gut regeneration of the holothurian E. fraudatrix. Transcriptomes of intestinal anlage of the three stages of regeneration, as well as the normal gut, were sequenced with an Illumina sequencer (San Diego, CA, USA). We identified 14,617 sea urchin protein homologs, of which 308 were transcription factors. After analysing the dynamics of gene expression during regeneration and the map of biological processes in which they participate, we identified 11 factors: Ef-EGR1, Ef-ELF, Ef-GATA3, Ef-ID2, Ef-KLF1/2/4, Ef-MSC, Ef-PCGF2, Ef-PRDM9, Ef-SNAI2, Ef-TBX20, and Ef-TCF24. With the exception of TCF24, they are all involved in the regeneration, development, epithelial-mesenchymal transition, and immune response in other animals. We suggest that these transcription factors may also be involved in the transdifferentiation of coelomic epithelial cells into enterocytes in holothurians.
Asunto(s)
Transdiferenciación Celular/genética , Transdiferenciación Celular/fisiología , Pepinos de Mar/genética , Animales , Células Epiteliales , Regeneración/fisiología , Pepinos de Mar/metabolismo , TranscriptomaRESUMEN
The transcriptome of the holothurian Apostichopus japonicus was sequenced at four developmental stages-blastula, gastrula, auricularia, pentactula-on an Illumina sequencer. Based on our RNA-seq data and the paired-end reads from 16 libraries obtained by other researchers earlier, we have achieved the currently most complete transcriptome assembly for A. japonicus with the best basic statistical parameters. An analysis of the obtained transcriptome has revealed 174 differentially expressed transcription factors, as well as stage-specific transcription factors that are most promising for further study. In addition, a total of 1,174,999 high-quality single nucleotide polymorphisms have been identified, including 58,932 indels. A GO enrichment analysis of contigs containing polymorphic loci shows the predominance of GO terms associated with immune response. The data obtained by us provide an additional basis for a deeper study of the mechanisms of the planktotrophic-type development in holothurians and can be used in commercial sea cucumber breeding programs.