RESUMEN
MicroRNAs modulate male fertility by regulating gene expression. In this study, dynamics of sperm miR-15a, miR-29b and miR-34a from high fertility (HF) and low fertility (LF) bulls using RT-qPCR were evaluated. Bioinformatic tools were employed to ascertain genes of interest of the sperm miRNAs. The expression levels of p53, BCL2, BAX and DNMT1 in bull spermatozoa were determined by immunoblotting. MicroRNA levels of miR-15a and miR-29 were higher in LF sires when compared with those present in HF bulls. Expression levels of miR-34a did not differ between the two groups. We found an inverse correlation between miR-15a and bull fertility. MiR29-b was also negatively associated with fertility scores. BCL2 and DNMT1 were higher in HF bulls while BAX was higher in the LF group. Our data showed a positive correlation between BCL2 and bull fertility. In addition, DNMT1 was positively associated with bull fertility. Furthermore, levels of BAX were negatively linked with bull fertility scores. Identification of miRNAs found in the spermatozoa of sires with different in vivo fertility helps understand the alterations in the fertilising capacity from cattle and other mammals. These potential biomarkers can be used in reproductive biotechnology as fertility markers to assess semen quality and predict male fertility.
Asunto(s)
Bovinos/fisiología , Fertilidad/genética , MicroARNs/metabolismo , Análisis de Semen/veterinaria , Espermatozoides/metabolismo , Animales , Biomarcadores/metabolismo , Cruzamiento , Biología Computacional , ADN (Citosina-5-)-Metiltransferasa 1/genética , Regulación de la Expresión Génica/fisiología , Masculino , Proteínas Proto-Oncogénicas c-bcl-2/genética , Análisis de Semen/métodos , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Integrins have been shown to act as signalling receptors, and they primarily recognise extracellular matrix ligands on the oocyte surface. However, their possible roles in oocyte activation and embryo development are not clearly understood. The objectives of this study were to evaluate expression of Integrin Subunit Beta 5 (ITGß5) in bovine sperm, oocytes, and early embryos and to ascertain the evolutionary conservation of ITGß5. To accomplish these objectives, we used western blotting to study expression levels of ITGß5 protein in sperm and RT-qPCR to determine expression levels of ITGß5 transcripts in oocytes and embryos. We have also used bioinformatic analysis to determine the evolutionary conservation of the ITGß5 protein among various species. Western blotting showed that ITGß5 protein was detectable in bull sperm. Moreover, results of RT-qPCR showed that levels of ITGß5 were significantly higher in the two-cell embryos, followed by the 8-16-cell embryos. However, no significant difference in expression levels were noted for the morula and blastocyst stages as compared to MII oocytes. Bioinformatic analysis revealed that ITGß5 is conserved among various species. We conclude that expression of ITGß5 in bovine gametes and embryos implies an important role in fertilisation and embryogenesis.
Asunto(s)
Cruzamiento , Bovinos/embriología , Desarrollo Embrionario , Fertilización In Vitro/veterinaria , Cadenas beta de Integrinas/metabolismo , Animales , Bovinos/genética , Biología Computacional , Embrión de Mamíferos/embriología , Embrión de Mamíferos/metabolismo , Evolución Molecular , Femenino , Cadenas beta de Integrinas/genética , Masculino , Oocitos/metabolismo , Alineación de Secuencia , Espermatozoides/metabolismoRESUMEN
Decreasing mammalian fertility and sperm quality have created an urgent need to find effective methods to distinguish non-viable from viable fertilising spermatozoa. The aims of the present study were to evaluate expression levels of ?-tubulin 2C (TUBB2C), heat shock protein 10 (HSP10), hexokinase 1 (HXK1) and superoxide dismutase 1 (SOD1) in spermatozoa from Holstein bulls with varying fertility using western blotting and to analyse the biological networks of these key sperm proteins using a bioinformatics software (Metacore; Thomson-Reuters, Philadelphia, PA, USA). The rationales behind this study were that the sperm proteins play crucial roles in fertilisation and early embryonic development in mammals and ascertaining the biological networks of the proteins helps us better understand sperm physiology and early mammalian development. The results showed that expression of SOD1 was higher in spermatozoa from high fertility bulls (PPin vivo bull fertility. The findings are important because they illuminate molecular and cellular determinants of sperm viability and the identified protein markers can be used to determine bull fertility.
