Molecular and Cellular Correlates of Sperm Viability Associated with Male Fertility

Fertility is classified as one of the most limiting aspects plaguing successful mammalian reproduction. In addition, sequential collaborations between a quality spermatozoon and oocyte are essential for successful fertilization, and key in subsequent embryonic development. Male fertility has been consistently declining in mammals. Currently, cost efficient and reliable methods predicting fertility are lacking. As a result, the objectives of this research were to establish molecular markers to predict male fertility by determining sperm cellular phenotype variability among bulls of varying fertility by focusing on the roles of protein markers, TUBB 2C, HSP10, HXK1 and SOD1; establish expression characteristics of ITGB5 protein in sperm, oocytes and early embryos; and finally; determine expression level variability and functions of miRNAs, miR-214 and miR-25 in sperm from bulls of varying fertility. We found that while no significant differences occurred in the expression levels of our proteins in high and low fertility bulls, correlations were discovered between SOD1, HSPE1 and fertility, and subpopulations consisting of intact, partially damaged and completely damaged acrosomes in flow cytometry and microscopy experiments were identified. Results also showed that expression levels of itgb5 were significantly higher in the 2-cell bovine 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 the MII oocytes. Phylogenetic analyses confirmed conservation of itgb5 across multiple species, further indicating its potential importance and functional role(s) in fertilization and potential as a marker of fertility. Micro-RNA studies revealed differential expression levels of miR-25 and miR- 214 for all samples analyzed, yet there was no significant difference in expression of the microRNAs when comparing high fertility and low fertility bulls. Also, processes associated with spermatogenesis, such as cellular growth, transduction, translation and transformation which are necessary for fertility, were shown to be targets of miR-214 and miR-25. These results imply that sperm proteins and sperm-bore miRNAs could potentially be targeted as molecular markers of fertility. The findings are important because they illuminate the molecular and cellular underpinnings of gamete quality that influence successful fertilization and early development