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The National Agriculture and Food Research Organization (NARO) has developed a stable and long-lasting system for producing conditioned medium that improves the quality of bovine in vitro fertilized embryos. The three major challenges for this system to work, including (1) establishment of an immortalized bovine oviduct epithelial cell line, (2) confirmation of the development-promoting effects of the conditioned medium, and (3) isolation of the effective factors contained in the conditioned medium, have been resolved. This technology is expected to improve the quality of in vitro fertilized embryos, thereby increasing conception rates after embryo transfer.

In dairy and beef cattle production, embryo transfer using in vitro fertilization (IVF) technology is widely practiced, and the number of embryo transfers in Japan is several with tens of thousands of transfers performed annually in Japan and increasing each year. However, the conception rate of IVF eggs has remained at around 40% for more than 30 years, making improvement in this area a long-standing technical challenge.

One effective approach to improving the quality of IVF embryos is to replicate the in vivo fertilization environment. A well-known method involves using conditioned medium derived from the culture of oviduct epithelial cells collected from the oviduct where fertilization takes place. However, conventional cell cultures have a short lifespan, making it difficult to produce conditioned medium consistently.

To address this issue, NARO has established a spontaneously immortalized bovine oviduct epithelial cell line with a semi-permanent proliferation ability, developed a system to continuously produce the conditioned medium using this cell line, and confirmed the development-promoting effects of the medium on the quality of IVF eggs.

For determining the quality of IVF embryos, we used a set of evaluation criteria that we believe are the most reliable and practical among existing reports for predicting pregnancy success. The four evaluation criteria are based on the normal kinetics and morphology of the embryos during the first two days after fertilization. The conditioned medium was found to increase the percentage of embryos that met the recommended criteria.

Further analysis showed that the sediment recovered after ultracentrifugation of the conditioned medium exhibited similar developmental benefits. This sediment can be used as a convenient supplement to other media and is expected to be widely used in the field of IVF embryo production.

With the establishment of a stable production system for conditioned medium, this technology is expected to significantly improve the quality of bovine IVF embryos, leading to improved conception rates in cattle breeding. Its practical application could contribute to increased efficiency and productivity in the livestock industry.

Norikazu Miyashita, Satoshi Akagi, Tamas Somfai, Yuji Hirao (2024) Serum-free spontaneously immortalized bovine oviduct epithelial cell conditioned medium promotes the early development of bovine in vitro fertilized embryos. Journal of Reproduction and Development 70:42-48. doi.org/10.1262/jrd.2023-031

This work was supported by grants from the Research project for technologies to strengthen the international competitiveness of Japan's agriculture and food industry and from NARO.