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Upregulation of iNOS and phosphorylated eNOS in the implantation-induced blastocysts of mice
Misato Seki1, Eisaku Takeuchi1, Emiko Fukui1,2
1Laboratory of Animal Breeding and Reproduction, Division of Animal Science, School of Agriculture Utsunomiya University Utsunomiya, Tochigi Japan.
Purpose:
This study aimed to examine expressions of iNOS and phosphorylated eNOS (p-eNOS) in implantation-induced blastocysts. We also examined the upstream of p-eNOS.
Methods:
To address the protein expressions in implantation-induced blastocysts, we performed immunohistochemical analysis using a delayed implantation mouse model. Immunostaining for iNOS, p-eNOS, and p-Akt was done. To address the relationship between p-eNOS and p-Akt, activated blastocysts were treated with an Akt inhibitor, MK-2206.
Results:
iNOS expression was at low levels in dormant blastocysts, whereas the expression was significantly increased in the activated blastocysts. Double staining of p-eNOS and p-Akt in individual blastocysts showed colocalization of p-eNOS and p-Akt of the trophectoderm. p-eNOS and p-Akt expressions were at low levels in dormant blastocysts, whereas both of them were significantly increased in the activated blastocysts. Both dormant and activated blastocysts showed significant positive correlations between p-eNOS and p-Akt. MK-2206 treatment for activated blastocysts showed that blastocysts with lower p-Akt had significantly lower p-eNOS levels.
Conclusions:
iNOS and p-eNOS, Ca2+ independent NOS, are upregulated by E2 in the blastocysts during implantation activation. Furthermore, p-eNOS is upregulated in implantation-induced blastocysts downstream of p-Akt.
