RESUMEN
Estrogenic endocrine disrupting chemicals (EEDC) have been suspected to impact offspring in a transgenerational manner via modifications of the germline epigenome in the directly exposed generations. A holistic assessment of the concentration/ exposure duration-response, threshold level, and critical exposure windows (parental gametogenesis and embryogenesis) for the transgenerational evaluation of reproduction and immune compromise concomitantly will inform the overall EEDC exposure risk. We conducted a multigenerational study using the environmental estrogen, 17α-ethinylestradiol (EE2), and the marine laboratory model fish Oryzias melastigma (adult, F0) and their offspring (F1-F4) to identify transgenerationally altered offspring generations and phenotype persistence. Three exposure scenarios were used: short parental exposure, long parental exposure, and a combined parental and embryonic exposure using two concentrations of EE2 (33ng/L, 113ng/L). The reproductive fitness of fish was evaluated by assessing fecundity, fertilization rate, hatching success, and sex ratio. Immune competence was assessed in adults via a host-resistance assay. EE2 exposure during both parental gametogenesis and embryogenesis was found to induce concentration/ exposure duration-dependent transgenerational reproductive effects in the unexposed F4 offspring. Furthermore, embryonic exposure to 113 ng/L EE2 induced feminization of the directly exposed F1 generation, followed by subsequent masculinization of the F2 and F3 generations. A sex difference was found in the transgenerationally impaired reproductive output with F4 females being sensitive to the lowest concentration of EE2 (33 ng/L) upon long-term ancestral parent exposure (21 days). Conversely, F4 males were affected by ancestral embryonic EE2 exposure. No definitive transgenerational impacts on immune competence were identified in male or female offspring. In combination, these results indicate that EEDCs can be transgenerational toxicants that may negatively impact the reproductive success and population sustainability of fish populations.
Asunto(s)
Oryzias , Contaminantes Químicos del Agua , Animales , Femenino , Masculino , Oryzias/fisiología , Aptitud Genética , Contaminantes Químicos del Agua/toxicidad , Reproducción , Fertilidad , Etinilestradiol/toxicidadRESUMEN
Germ plasm, a cytoplasmic factor of germline cell differentiation, is suggested to be a perspective tool for in vitro meiotic differentiation. To discriminate between the: (1) germ plasm-related structures (GPRS) involved in meiosis triggering; and (2) GPRS involved in the germ plasm storage phase, we investigated gametogenesis in the marine medaka Oryzias melastigma. The GPRS of the mitosis-to-meiosis period are similar in males and females. In both sexes, five events typically occur: (1) turning of the primary Vasa-positive germ plasm granules into the Vasa-positive intermitochondrial cement (IMC); (2) aggregation of some mitochondria by IMC followed by arising of mitochondrial clusters; (3) intramitochondrial localization of IMC-originated Vasa; followed by (4) mitochondrial cluster degradation; and (5) intranuclear localization of Vasa followed by this protein entering the nuclei (gonial cells) and synaptonemal complexes (zygotene-pachytene meiotic cells). In post-zygotene/pachytene gametogenesis, the GPRS are sex specific; the Vasa-positive chromatoid bodies are found during spermatogenesis, but oogenesis is characterized by secondary arising of Vasa-positive germ plasm granules followed by secondary formation and degradation of mitochondrial clusters. A complex type of germ plasm generation, 'the follicle cell assigned germ plasm formation', was found in late oogenesis. The mechanisms discovered are recommended to be taken into account for possible reconstruction of those under in vitro conditions.
Asunto(s)
Gránulos Citoplasmáticos/fisiología , ARN Helicasas DEAD-box/metabolismo , Células Germinativas/citología , Oocitos/citología , Oogénesis , Oryzias/crecimiento & desarrollo , Espermatocitos/citología , Espermatogénesis , Animales , Núcleo Celular , Gránulos Citoplasmáticos/ultraestructura , Femenino , Proteínas de Peces/metabolismo , Células Germinativas/metabolismo , Células Germinativas/ultraestructura , Masculino , Oocitos/metabolismo , Espermatocitos/metabolismoRESUMEN
Lamin is an intermediate protein underlying the nuclear envelope and it plays a key role in maintaining the integrity of the nucleus. A defect in the processing of its precursor by a metalloprotease, ZMPSTE24, results in the accumulation of farnesylated prelamin in the nucleus and causes various diseases, including Hutchinson-Gilford progeria syndrome (HGPS). However, the role of lamin processing is unclear in fish species. Here, we generated zmpste24-deficient medaka and evaluated their phenotype. Unlike humans and mice, homozygous mutants did not show growth defects or lifespan shortening, despite lamin precursor accumulation. Gonadosomatic indices, blood glucose levels, and regenerative capacity of fins were similar in 1-year-old mutants and their wild-type (WT) siblings. Histological examination showed that the muscles, subcutaneous fat tissues, and gonads were normal in the mutants at the age of 1â¯year. However, the mutants showed hypersensitivity to X-ray irradiation, although p53target genes, p21 and mdm2, were induced 6 h after irradiation. Immunostaining of primary cultured cells from caudal fins and visualization of nuclei using H2B-GFP fusion proteins revealed an abnormal nuclear shape in the mutants both in vitro and in vivo. The telomere lengths were significantly shorter in the mutants compared to WT. Taken together, these results suggest that zmpste24-deficient medaka phenocopied HGPS only partially and that abnormal nuclear morphology and lifespan shortening are two independent events in vertebrates.
Asunto(s)
Núcleo Celular/patología , Modelos Animales de Enfermedad , Proteínas de Peces/deficiencia , Proteínas de la Membrana/deficiencia , Metaloendopeptidasas/deficiencia , Oryzias/genética , Progeria/patología , Aletas de Animales/enzimología , Aletas de Animales/patología , Aletas de Animales/efectos de la radiación , Animales , Animales Modificados Genéticamente , Núcleo Celular/enzimología , Núcleo Celular/efectos de la radiación , Forma del Núcleo Celular/efectos de la radiación , Células Cultivadas , Codón sin Sentido , Femenino , Proteínas de Peces/química , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Técnicas de Inactivación de Genes , Proteínas Fluorescentes Verdes/química , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Heterocigoto , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Metaloendopeptidasas/genética , Metaloendopeptidasas/metabolismo , Oryzias/metabolismo , Progeria/enzimología , Progeria/genética , Tolerancia a Radiación , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Análisis de Supervivencia , Acortamiento del Telómero/efectos de la radiaciónRESUMEN
Marine medaka Oryzias melastigma at 4months (young), 8months (middle-aged) and 12months old (senior) were employed to determine age-associated change of sex ratios, sex hormones, telomere length (TL), telomerase activity (TA), telomerase transcription (omTERT) and oxidative damage in the liver. Overall, O. melastigma exhibited gradual senescence, sex differences in longevity (F>M), TL (F>M) and oxidative damage (F