Growth factors, hypoxia-inducible factor 1-alpha (HIF-1α) and klotho protein all have very important functions in the male reproduction; however their role in the regulation of seasonal reproductive processes in the male European bison remains unclear. Similarly, although the uterus masculinus is very frequently found in the bison, its importance and functions remain unknown. It is likely that, this organ may have secretory functions and thus be a target for various regulatory factors. Therefore, the aim of this study was to investigate expression and activity of several factors: vascular endothelial growth factor (VEGF-A), fibroblast growth factor (FGF-2), transforming growth factor β1 (TGF-β1), nerve growth factor (NGF), insulin-like growth factor receptor (IGF-IR β), hypoxia-inducible factor 1 alpha (HIF-1α), and klotho protein in the uterus masculinus, immediately after the season of the reproductive activity (November and December). Our study reveals that the growth factor expression levels are significantly higher in November, when compared to December, while expression of HIF-1α and klotho was higher in December. These results provide novel data on differences in the expression levels of several factors in the uterus maculinus of European bison bulls after the breeding season. The described factors may, therefore, be potent regulators of the seasonal reproduction.