Expression patterns of ZO-1/2 and their effects on porcine oocyte in vitro maturation and early embryonic development.

Zonula occludens (ZO)-1 and ZO-2 are involved in epithelial polarity maintenance, gene transcription, cell proliferation and tumor cell metastasis. Regulating ZO-1/2 expression influences the early embryonic development of mice, but whether they are involved in oocyte maturation is still poorly understood. In the present study, the expression patterns of ZO-1 and ZO-2 in porcine cumulus cells and oocytes matured in vitro and early embryos from parthenogenetic activation were detected by qRT-PCR or Western blot, and then their roles in porcine oocyte maturation and early embryo development were investigated by shRNA technology. ZO-1 and ZO-2 were found to be expressed in cumulus cells, oocytes and early embryos, while ZO-1α was expressed only in cumulus cells, morula and blastocysts. During in vitro maturation (IVM), the abundance of ZO-1 and ZO-2 in oocytes was significantly higher than that in cumulus cells at 0 h (P < 0.01), and their mRNA and protein levels displayed relatively higher expression at 0 and 18 h, respectively. Compared with the control groups, cumulus cell expansion, oocyte nucleus maturation, and subsequent cleavage were not influenced by treatment of the cumulus-oocyte complexes (COCs) with ZO-1-shRNA1, ZO-2-shRNA2 or combined ZO-1-shRNA1 and ZO-2-shRNA2 lentivirus (P > 0.05). However, the blastocyst rate was reduced by treatment of COCs with ZO-1-shRNA1 but not ZO-2-shRNA2. The total cell number of blastocysts was decreased by downregulation of ZO-1 and ZO-2 (P < 0.05). Downregulation of ZO-1 and ZO-2 also resulted in a significant decrease (P < 0.05) in the expression of Cx43, Cx45, PTX3 and PTGS2 in cumulus cells, Cx45, BMP15, ZP3 and C-KIT in MII oocytes, and Nanog in blastocysts, with the exception of HAS2 expression in cumulus cells and Oct4 expression in blastocysts (P > 0.05). Altogether, the above results indicate that ZO-1 and ZO-2 display similar expression patterns during porcine oocyte IVM and are critical to porcine oocyte maturation and early embryonic development.