RESUMO
In this study we extended earlier work to determine whether sperm respond to somatic cell apoptotic stimuli and whether apoptotic phenotypes are significant indicators of human sperm quality. We evaluated ejaculated sperm from fertile donors and subfertile patients following purification of fractions of high and low motility. In unstimulated conditions, caspase enzymatic activity was higher in motile fractions from subfertile patients than in donors, and was higher in low motility fractions from both groups. Staurosporine, but not a Fas ligand or H2O2, significantly increased caspase activity, but only in high motility fractions. Procaspase-3, -7 and -9 and low levels of active caspase-3, -7 and -9 were identified by immunoblot analysis. Apoptosis-inducing factor (AIF) was present in all samples but poly ADP-ribose polymerase-1 (PARP-1) was not detected. Phosphatidylserine translocation was significantly increased only with H2O2 treatment. In ejaculates of both subfertile and fertile men, we demonstrated the presence and activation of several proteins that are key constituents of apoptosis-related pathways in somatic cells, which may serve as markers for sperm quality.
Assuntos
Apoptose , Infertilidade Masculina/diagnóstico , Motilidade dos Espermatozoides/fisiologia , Espermatozoides/química , Espermatozoides/metabolismo , Adulto , Anexina A5/metabolismo , Fator de Indução de Apoptose , Transporte Biológico/fisiologia , Biomarcadores/análise , Biomarcadores/metabolismo , Caspases/análise , Caspases/metabolismo , Ejaculação/fisiologia , Proteína Ligante Fas , Flavoproteínas/análise , Flavoproteínas/metabolismo , Humanos , Isoenzimas/análise , Isoenzimas/metabolismo , Masculino , Glicoproteínas de Membrana/farmacologia , Proteínas de Membrana/análise , Proteínas de Membrana/metabolismo , Fosfatidilserinas/metabolismo , Poli(ADP-Ribose) Polimerases/análise , Poli(ADP-Ribose) Polimerases/metabolismo , Motilidade dos Espermatozoides/efeitos dos fármacos , Espermatozoides/citologia , Estaurosporina/farmacologiaRESUMO
Ejaculated spermatozoa, particularly in infertile men, have been shown to display morphological and biochemical features that are typical of an apoptotic phenotype in somatic cells. Deregulation of apoptosis is known to play roles in a number of disease processes, but roles for apoptosis in ejaculated spermatozoa and male infertility are poorly defined or have not been studied. Preliminary data demonstrated that populations of ejaculated spermatozoa express: (i) various degrees of plasma membrane translocation of phosphatidylserine and DNA fragmentation; and (ii) active caspase-3, the main executor of apoptosis in somatic cells, with an apparent exclusive cellular location to the mid-piece. Tests are currently being carried out on the effects of well-known apoptosis agonists and caspase inhibitors on such markers using purified populations of leukocyte-free ejaculated human spermatozoa. The main objective is to determine if somatic cell apoptosis markers are relevant indicators and/or causative factors of male infertility.
