RESUMO
c-Jun N-terminal kinase (JNK) is one of the mitogen-activated protein kinases. Previous studies showed that the JNK is involved in signaling pathways initiating cell cycle, and eventually, causing apoptosis through persistent activation in mammals. In this article, it is further revealed that the jnk1 gene is closely related with the embryonic development and organogenesis in zebrafish. RT-PCR and Western blot analysis show that there were distinct expression patterns of JNK at the different developmental stages as well as in the various tissues in zebrafish. Knockdown of jnk1 by RNA interference (RNAi) resulted in high lethal, serious retardation and malformations of embryos in zebrafish. SP600125, a JNK-specific inhibitor, gives rise to high mortality in zebrafish, similar to that caused by the jnk1 RNA interference. SP600125 is also responsible for the severe abnormality of organs, especially the skeletal system, such as skull, mandible deficiency, and cyrtosis heterauxesis. The results also indicate that the inhibition of JNK by SP600125 suppresses the ovarian differentiation during the embryo development in zebrafish. Overall, our study demonstrates that the jnk1 gene is required for ovary differentiation and development in the zebrafish, and down-regulated JNK directly inhibits ovary differentiation during early ontogenetic stages.
Assuntos
Desenvolvimento Embrionário/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Organogênese/fisiologia , Transdução de Sinais/fisiologia , Peixe-Zebra/embriologia , Animais , Antracenos/farmacologia , Western Blotting , Primers do DNA/genética , Desenvolvimento Embrionário/efeitos dos fármacos , Desenvolvimento Embrionário/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/genética , Técnicas Histológicas , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Organogênese/efeitos dos fármacos , Organogênese/genética , Ovário/efeitos dos fármacos , Ovário/crescimento & desenvolvimento , Interferência de RNA , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genéticaRESUMO
Molecularly imprinted polymers (MIPs) were prepared using bisphenol A (BPA) as a template by precipitation polymerization. The polymer that had the highest binding selectivity and ability was used as solid-phase extraction (SPE) sorbents for direct extraction of BPA from different biological and environmental samples (human serum, pig urine, tap water and shrimp). The extraction protocol was optimized and the optimum conditions were as follows: conditioning with 5 mL methanol-acetic acid (3:1), 5 mL methanol, 5 mL acetonitrile and 5 mL water, respectively, loading with 5 mL aqueous samples, washing with 1 mL acetonitrile, and eluting with 3 mL methanol. MIPs can selectively recognize, effectively trap and preconcentrate BPA over a concentration range of 2-20 microM. Recoveries ranged from 94.03 to 105.3%, with a relative standard deviation lower than 7.9%. Under the optimal condition, molecularly imprinted SPE recoveries of spiked human serum, pig urine, tap water and shrimp were 65.80, 82.32, 76.00 and 75.97%, respectively, when aqueous samples were applied directly. Compared with C18 SPE, a better baseline, better high-performance liquid chromatography separation efficiency and higher recoveries were achieved after molecularly imprinted SPE.