RESUMO
OBJECTIVE: To evaluate the expression pattern of Dazl (deleted in azoospermia-like) protein in the mouse preimplantation embryo. DESIGN: Experimental study. SETTING: Medical research laboratory in a university hospital. ANIMAL(S): Twenty female 28- to 35-day-old FVB mice. INTERVENTION(S): Embryo collection at 1.5, 2.5, and 3.5 days postcoitus (plug date, 0.5 d postcoitus) to examine the Dazl protein expression from the two-cell embryo to the blastocyst. MAIN OUTCOME MEASURE(S): Dazl protein expression was analyzed by immunofluorescent staining. RESULT(S): There is abundant expression of Dazl protein in the cytoplasm of the blastomere. Strong fluorescent signals of Dazl protein expression were found in preimplantation embryo cytoplasm, including two-cell, eight-cell, morula, and blastocyst. CONCLUSION(S): By using an antibody raised against mouse Daz-like protein (Dazl), we showed that Dazl protein is present in all cleaving stages of the preimplantation embryo. This is the first report on the protein expression of a Dazl gene during embryogenesis in mice. However, further study is needed to evaluate the molecular functional role of Dazl.
Assuntos
Blastocisto/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Células Cultivadas , Fase de Clivagem do Zigoto/metabolismo , Citoplasma/metabolismo , Desenvolvimento Embrionário/fisiologia , Feminino , Camundongos , Camundongos Endogâmicos , GravidezRESUMO
OBJECTIVE: To explore the role of CDC25B protein in postmeiotic germ cells. DESIGN: In vitro experiment. SETTING: University-based reproductive genetics laboratory. PATIENT(S): Fertile and infertile volunteers. INTERVENTION(S): Reverse transcription-polymerase chain reaction (RT-PCR), real-time RT-PCR, and immunostaining for CDC25B. MAIN OUTCOME MEASURE: Expression profiling of CDC25B in human spermatozoa. RESULT(S): Four splicing variants (CDC25B1, B2, B3, and B4) are expressed in human spermatozoa. Immunofluorescence staining showed strong homogeneous staining in the midpiece of spermatozoa, and weak staining in the principal piece and cytosol of the head. The messenger RNA (mRNA) transcript level of CDC25B was increased in sperm samples of men with asthenospermia. CONCLUSION(S): The expression of CDC25B in different cellular compartments of human spermatozoa suggests that there are diverse noncell-cycle-related functions of CDC25B in terminally differentiated human germ cells.
Assuntos
Proteínas de Ciclo Celular/genética , Espermatozoides/metabolismo , Fosfatases cdc25/genética , Adulto , Astenozoospermia/genética , Astenozoospermia/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/fisiologia , Imunofluorescência , Expressão Gênica , Humanos , Isoenzimas/genética , Isoenzimas/metabolismo , Masculino , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Distribuição Tecidual , Fosfatases cdc25/metabolismo , Fosfatases cdc25/fisiologiaRESUMO
BACKGROUND: DAZ gene family is crucial for human spermatogenesis that requires the precise co-ordination of cell cycle events. CDC25A is recognized as the downstream substrate of DAZ gene family and is thought to function on the M-phase regulation of cell cycles. We investigated the expression profiles of CDC25A in the testes of infertile men and evaluated the relationship between CDC25A levels and testicular phenotype, clinical hormonal parameters and sperm retrieval results. METHODS: The protein and mRNA transcript levels of CDC25A in the testes of 40 azoospermic men were determined by immunohistochemistry and quantitative real-time-PCR. CDC25A in human spermatozoa was investigated by western blotting and immunofluorescence staining. RESULTS: The CDC25A protein was expressed mainly in spermatocyte, spermatid and spermatozoa. CDC25A transcript levels were significantly decreased (P = 0.0009) in patients with spermatogenic failure, especially in men with meiotic arrest and Sertoli cell-only syndrome. Significantly higher CDC25A transcript levels were detected in patients with successful sperm retrieval than in patients with failed sperm retrieval (P = 0.005). CONCLUSIONS: Decreased CDC25A is associated with spermatogenic failure and failed sperm retrieval in infertile men. Further studies are necessary to explore the functional roles of CDC25A in human spermatozoa.
