Recent studies showed that tight junctions (TJs) integrity and assembly are required for blastocyst development in mouse and pig models. However, the biological functions of TJs associated with embryo implantation and maintenance of pregnancy were not investigated yet. To examine whether disrupted TJs affect further embryo development, we employed RNAi approach and inhibitor treatment. The embryos were injected with Cxadr (Coxsackievirus and adenovirus receptor) siRNA for knock down (KD) and treated with Adam10 (A Disintegrin and Metalloproteinase specific inhibitor 10; GI254023X; SI). We compared blastocyst development and paracellular sealing assay using FITC dextran uptake between control and KD or SI embryos. Finally, we transferred control and Cxadr KD or Adam 10 SI treated blastocyst to uteri of recipients. Cxadr KD and Adam 10 SI showed lower blastocyst development and more permeable to FITC-dextran. Moreover, we observed that half of KD and inhibited embryos failed to maintain pregnancies after the second trimester. Our findings suggested that TJs integrity is required for the maintenance of pregnancy and can be used as a selective marker for the successful application of assisted reproduction technologies.

Establishment of the Adherens junction (AJ) and Tight junction (TJ) are important steps in terms of morphological formation during preimplantation develoment. Particularly, TJ complex is crucial for cavitation in blastocyst. So far, many TJ protein/genes are revealed. However, the biological function and regulation of TJ were not elucidated during post implantation. We depleted several TJ and TJ associated genes using RNA interference, and examined preimplantation development with TJ. We tested functionality of paracellular sealing to determine integrity of TJ formation and examined TE differentiation indirectly using outgrowth assay in vitro. We observed defect of paracellular permeability in the TJ related genes knockdown(KD) blastocyst and abnormal outgrowth. Particularly, trophoblast cells were not stretched out in the KD groups. Finally, we did embryo transfer using the TJ genes KD and control blastocysts into surrogate mothers. We found lower of the implantation rates/ maintenance of pregnancy in the TJ KD groups (less than 40%) than in the controls (about 80%). In conclusion, TJ integrity is can be used as a selective marker for developmentally competent embryos and successful pregnancy.