The phenomenon of aging arises from multiple, complex interactions causing dysfunction in cells and organs. In particular, fertility drastically decreases with age. Previously, we have demonstrated that the functional characteristics of the bovine oviduct and uterus change with the age-dependent upregulation of inflammation and noted that S100A9 triggers inflammatory responses in oviduct epithelial cells. In the present study, we investigated the hypothesis that S100A9 affects reproductive events to aspect such as sperm function, fertilization, and the development of the embryo in cows. To investigate the effect of S100A9 on bovine sperm, we incubated sperms in vitro with S100A9 for 5 h and observed significantly decreased sperm motility and viability. During in vitro fertilization, S100A9 treatment for 5 h did not affect the rate of fertilization, time of first division of embryos, or embryo development to blastocyst stage. Treatment of 2-cell stage embryos with S100A9 for 5 h significantly reduced the proportion of cells undergoing normal division (4-8 cell embryos) and embryo development to the blastocyst stage. In experiment involving 24 h treatment of 2-cell embryos, the development of all embryos stopped at the 2-cell stage in the S100A9-treated group. In blastocyst-stage embryos, S100A9 treatment significantly stimulated the expression of endoplasmic reticulum (ER) and the mRNA expression of ER stress markers, and activated caspase-3 with subsequent nuclear fragmentation. Pre-treatment with an ER stress inhibitor significantly suppressed caspase-3 activation by the S100A9 treatment, suggesting that S100A9 induces blastocyst dysfunction by apoptosis (via caspase-3 activation) depending on ER stress. These results indicate that direct exposure to S100A9 exerted adverse effects on sperm function and embryo development. These findings suggest that excessive dose of S100A9 may have an adverse effect to the reproductive machinery by inducing inflammation and tissue dysfunction.