Epididymal epithelial cells cultured in vitro prolong the motility of bovine sperm

It is well known that the epididymis is an excellent environment to maintai n sperm viability. Therefore, we used different sections of bovine epididym is (caput, corpus, and cauda) to develop epithelial cell culture monolayers to identify factors that will increase sperm survival in the freezing-thaw ing process. Each epididymal section was dissected and treated with collage nase to obtain epithelial cell clusters. The cells were cultured in RPMI-16 40 medium with 10% serum at 38.5 degreesC. A confluent monolayer was obtain ed after 5-7 days in culture and preliminary characterization using cytoker atin antibody indicated that the cell culture contained 85%-95% of epitheli al cells. These cellular cultures were tested for their ability to maintain motility of epididymal and frozen-thawed spermatozoa. Washed spermatozoa w ere added to obtain a final dilution of 1 x 10(6) spermatozoa/mL. The motil ity of frozen-thawed spermatozoa was also recorded after incubation in cond itioned media. Our results show that cocultures of spermatozoa and epididym al cell monolayers for 24 and 48 hours were beneficial for maintaining epid idymal and frozen-thawed sperm motility (36.0% and 20.4%) compared with spe rmatozoa cultured with fibroblast cells or in the absence of a cell monolay er (0%; P < .01). The conditioned medium provides favorable conditions for sperm motility. Results with conditioned medium on maintenance of frozen-th awed sperm motility suggest that epididymal cells in vitro secrete benefici al factors that prolong the sperm survival.