FERTILIZING ABILITY OF BOVINE SPERMATOZOA COCULTURED WITH OVIDUCT EPITHELIAL-CELLS

The effects of bovine oviduct epithelial cell monolayers (OECM), derived from the different segments of the oviduct and under various conditions, on oocyte penetration by bovine sperm were determined by in vitro cell culture techniques. The oocyte penetration rate by sperm treated with OECM+OECM-conditioned medium was 93% (155 of 166). Sperm penetration rates in OECM+fresh medium and in OECM-conditioned medium alone were 66% (115 of 173) and 52% (87 of 167), respectively, significantly lower (p < 0.01) than in OECM+OECM-conditioned medium. However, the percentage of penetrated oocytes in OECM-conditioned medium alone was significantly higher than in control (11% = 16 of 148;p < 0.01). In both the OECM+OECM-conditioned medium and the OECM+fresh medium groups, oocyte penetration rates by sperm cocultured with OECM derived from the isthmic segment were significantly lower than those by sperm cocultured with OECM derived from the ampullary segment (44% = 43 of 98 vs. 72% = 68 of 94, and 42% = 49 of 117 vs. 66% = 72 of 110, respectively; p < 0.01). Sperm penetration rates were low after insemination in the OECM-conditioned medium alone derived from either the isthmic or the ampullary segments (20% = 20 of 100 and 19% = 17 of 81, respectively). However, the sperm penetration rate was improved significantly when OECM-conditioned medium was obtained from whole-oviduct OECM (57% = 60 of 105;p < 0 01). The effect of OECM derived from different segments on ability of sperm binding and maintaining motility was also evaluated in vitro. After 2 h of coculture, more than half the sperm attached to OECM regardless of their origin. Sperm were gradually released from OECM in the whole oviduct and ampullary segments after 3 h of coculture; however, sperm remained attached to isthmic OECM after 12 h of coculture. After 48 h of coculture, the motility of unattached sperm in either isthmic OECM or whole-oviduct OECM was significantly higher than in ampullary OECM (52% + 3, 55% +/- 4, and 25% +/- 4, respectively; P < 0.01). When sperm were cocultured with ampullary OECM, the percentage of motile sperm was significantly lower from 24 h after coculture than percentages obtained after coculture with whole-oviduct OECM and isthmic OECM (26% +/- 4, 63% +/- 3, and 61% +/- 4, respectively; p < 0.01). These results suggest that sperm capacitation was synergistically induced by means of both attachment to OECM and exposure to OECM-conditioned medium, and that sperm attachment to the ampullary epithelium enhanced sperm capacitation. These results also suggest that the isthmus epithelial cells are important in maintaining spertn motility and that attachment of sperm in the isthmus may act to reduce the number of sperm arriving at the ampulla in vivo.