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The blastocyst achieves on-time implantation by entosis

Posted by , on 1 September 2015

The process of embryo implantation consists of multiple steps: blastocyst apposition, adhesion to uterine luminal epithelial (LE) cells, and removal of the epithelial cells encasing the blastocysts. How the blastocyst trophectoderm breaches the luminal epithelial barrier has been studied for decades, the mechanism of the abstraction of LE cells was not clearly understood. Since the discovery of cell apoptosis mechanism, a putative thought has been that LE cells around the implanting embryo undergo this programmed cell death once they are attached with the blastocyst (Joswig et al, 2003; Parr et al, 1987). This mechanism indicates that degeneration of LE cells is a maternal intrinsic response to implanting embryos (Finn & Hinchliffe, 1964; Krehbiel, 1937). Although others suggested that trophoblast cells have a role in triggering epithelial cell apoptosis (Joswig et al, 2003), it is believed that trophoblast cells need to wait passively for the autolysis of apoptotic epithelial cells in order to get in contact of stromal cells.

However, our study shows that embryos play an active role in removing LE cells (Li et al, 2015). We have shown that LE cells in direct contact with the blastocyst are endocytosed by trophoblast cells by adopting the nonapoptotic cell-eat-cell process (entosis) in the absence of Caspase3 activation. About half a century ago, Finn and McLaren observed that degenerated uterine epithelial cells are taken up by the trophoblast cells (Finn & McLaren, 1967). The endocytosed epithelial cells were named as “W-body”. However, these notions were primarily based on observations of cell integrity and structure, but it was not clear whether it is entosis or phagocytosis, since both of them involve engulfment of one cell by another cell. In phagocytosis, only dead or dying cells are engulfed, whereas live cells are internalized in entosis. Our results showed that trophoblast cells actively engulf proximate live epithelial cells. This observation challenges the dogma that uterine epithelial cells undergo apoptosis attributed by maternal responses with minimal role played by embryonic cells in eliminating the LE cells.

Although our results demonstrated that trophoblast cells are able to entosize epithelial cells both in vivo and in vitro, we cannot rule out the possibility that the involvement of other mechanisms to remove LE cells during implantation. In fact, we observed that in the absence of embryos, epithelial cells undergo apoptosis in a longer time span in pseudopregnant mice. It would be interesting to examine whether blockage of entosis or epithelial cell apoptosis could completely block implantation and cause pregnancy failure. It is possible that embryos will still be able to complete implantation without entosis, although the implantation process may be delayed. However, most defects during the early implantation events result in either pregnancy failure or late stage pregnancy defects (Cha et al, 2012; Wang & Dey, 2006).

Although trophoblast cells are known to be invasive, our report is the first one to show that they are able to endocytose live cells. Trophoblast cells also play a key role during placentation. It has been shown that maternal endothelial cells are substituted by trophoblast cells (Cross, 2005). The mechanism is still not clear. Is it possible that entosis also plays a role during placentation?

We have shown previously that LE cells begin to lose their apicobasal polarity pending blastocyst implantation (Cha et al, 2014; Daikoku et al, 2011). The transition of LE cells from a high to a low polar state is critical to implantation, since the LE retains high polarity in mice with uterine inactivation of Msx, Klf5 or overexpression of Wnt5a, and blastocysts remain encased within the intact epithelium past day 6 of pregnancy which results in implantation failure (Cha et al, 2014; Daikoku et al, 2011; Sun et al, 2012). Therefore the initial phase of the implantation process requires a transition from high to lower polar state in the epithelium. In addition, usually epithelial cells in vivo are anchored onto the basement membrane, adhered to neighboring epithelial cells by junctional proteins, and aligned as a single layer. The cell-in-cell structure of entosis suggests that the engulfed cell loses connections to its neighboring LE cells and basal lamina, causing epithelial cells to be more flexible and thus easier to be engulfed. Therefore, it would be interesting to examine whether entosis requires decreases in epithelial polarity.

Our results showed that some trophoblast cells on the surface of the blastocyst became enlarged and formed dome-shaped bulges during entosis. The mechanism of reorganization of trophoblast cytoskeleton is not clear. Our study also showed that entosis is only observed in mural trophectoderm, which differentiates into trophoblast giant cells. The giant cell layer is the border between fetal and maternal tissues, and functions as a protective shell for fetal growth. We speculate that the early entosis of maternal cells by the cells in mural trophectoderm makes the future giants cells chimera of maternal and fetal origin, and thus facilitating the protective role of trophoblast giant cells.

 

References:

Cha J, Bartos A, Park C, Sun X, Li Y, Cha SW, Ajima R, Ho HY, Yamaguchi TP, & Dey SK (2014). Appropriate crypt formation in the uterus for embryo homing and implantation requires Wnt5a-ROR signaling. Cell reports, 8 (2), 382-92 PMID: 25043182

Cha J, Sun X, & Dey SK (2012). Mechanisms of implantation: strategies for successful pregnancy. Nature medicine, 18 (12), 1754-1767 PMID: 23223073

Cross JC (2005). How to make a placenta: mechanisms of trophoblast cell differentiation in mice–a review. Placenta, 26 Suppl A PMID: 15837063

Daikoku T, Cha J, Sun X, Tranguch S, Xie H, Fujita T, Hirota Y, Lydon J, DeMayo F, Maxson R, & Dey SK (2011). Conditional deletion of Msx homeobox genes in the uterus inhibits blastocyst implantation by altering uterine receptivity. Developmental cell, 21 (6), 1014-25 PMID: 22100262

Finn CA, & Hinchliffe JR (1964). Reaction of the Mouse Uterus during Implantation and Deciduoma Formation as Demonstrated by Changes in the Distribution of Alkaline Phosphatase Journal of reproduction and fertility, 8, 331-338 PMID: 14248593

Finn CA, & McLaren A (1967). A study of the early stages of implantation in mice Journal of Reproduction and Fertility, 13, 259-267 : 10.1530/jrf.0.0130259

Joswig A, Gabriel HD, Kibschull M, & Winterhager E (2003). Apoptosis in uterine epithelium and decidua in response to implantation: evidence for two different pathways. Reproductive biology and endocrinology, 1 PMID: 12801416

Krehbiel RH (1937). Cytological Studies of the Decidual Reaction in the Rat during Early Pregnancy and in the Production of Deciduomata Physiological Zoology, 10, 212-234

Li Y, Sun X, & Dey SK (2015). Entosis allows timely elimination of the luminal epithelial barrier for embryo implantation. Cell reports, 11 (3), 358-365 PMID: 25865893

Parr EL, Tung HN, & Parr MB (1987). Apoptosis as the mode of uterine epithelial cell death during embryo implantation in mice and rats. Biology of reproduction, 36 (1), 211-225 PMID: 3567276

Sun X, Zhang L, Xie H, Wan H, Magella B, Whitsett JA, & Dey SK (2012). Kruppel-like factor 5 (KLF5) is critical for conferring uterine receptivity to implantation. Proceedings of the National Academy of Sciences of the United States of America, 109 (4), 1145-1150 PMID: 22233806

Wang H, & Dey SK (2006). Roadmap to embryo implantation: clues from mouse models. Nature reviews. Genetics, 7 (3), 185-199 PMID: 16485018

 

 

 

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