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Akihiro Yoneda, Masashi Kashima, Shigeki Yoshida, Kei Terada, Shoma Nakagawa, Akiko Sakamoto, Koji Hayakawa, Keita Suzuki, Junji Ueda and Tomomasa Watanabe

The molecular mechanism by which sperm triggers Ca2+ oscillation, oocyte activation, and early embryonic development has not been clarified. Recently, oocyte activation has been shown to be induced by sperm-specific phospholipase Cζ (PLCζ). The ability of PLCζ to induce oocyte activation is highly conserved across vertebrates. In the present study, porcine PLCζ cDNA was identified and the nucleotide sequence was determined. The expression pattern of porcine PLCζ mRNA during the period of postnatal testicular development was shown to be similar to that of mouse PLCζ. PLCζ mRNA expression in the pig and mouse was detected only in the testes when the elongated spermatids had differentiated, and was detected from day 96 after birth in the pig. Histological examination of porcine testis during the period of postnatal development revealed the presence of spermatozoa from day 110 after birth. These findings suggest that the synthesis of PLCζ mRNA starts when spermiogenesis is initiated. Microinjection of porcine PLCζ complementary RNA into porcine oocytes demonstrated that porcine PLCζ has the ability to trigger repetitive Ca2+ transients in porcine oocytes similar to that observed during fertilization. It was also found that porcine PLCζ cRNA has the potential to induce oocyte activation and initiate embryonic development up to the blastocyst stage.

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Yumiko Miyazaki, Akihito Horie, Hirohiko Tani, Masashi Ueda, Asuka Okunomiya, Koh Suginami, Eiji Kondoh, Tsukasa Baba, Ikuo Konishi, Tamayuki Shinomura and Yukiyasu Sato

The endometrium extracellular matrix (ECM) is essential for embryo implantation. Versican, a large chondroitin sulfate proteoglycan that binds hyaluronan and forms large ECM aggregates, can influence fundamental physiological phenomena, such as cell proliferation, adhesion and migration. The present study investigated the possible role of versican in human embryo implantation. Versican V1 expression and secretion in human endometrial epithelial cells (EECs) was most prominent in the mid-secretory phase. Versican expression in EECs significantly increased after treatment with estrogen and progesterone, but not by estrogen alone. We also established versican V1-overexpressing Ishikawa (endometrial cancer cell line) cells (ISKW-V1), versican V3-overexpressing (ISKW-V3) and control GFP-overexpressing (ISKW-GFP) Ishikawa cells. By the in vitro implantation model, the attachment ratio of BeWo (choriocarcinoma cell line) spheroids to the monolayer of ISKW-V1, but not of ISKW-V3, was found significantly enhanced compared with attachment to the ISKW-GFP monolayer. The conditioned medium derived from ISKW-V1 (V1-CM) also promoted the attachment of BeWo spheroids to the ISKW monolayer. However, this attachment-promoting effect was abolished when V1-CM was pretreated with chondroitinase ABC, which degrades chondroitin sulfate. Therefore, out of the ECM components, versican V1 may facilitate human embryo implantation.