Fig. 3

EVs are released by early embryos via apocrine and merocrine secretory mechanisms. Upper panel: Schematic representation of the embryo after apposition/attachment to the luminal epithelium. Trophoblast cells are facing the luminal epithelium to maintain contact between the developing embryo and the receptive endometrium. The basal cell surface of trophoblast cells stays in close contact with elongated endodermal cells. Porcine trophoblast secretes EVs via apocrine (when the apical part of the secretory cell containing vesicles pinches off and enters the uterine lumen) and merocrine (when vesicles are released from the cell via exocytosis) mechanisms. Trophoblast cells also receive molecules from the uterine environment. Clathrin-mediated endocytosis and pinocytosis are portals for uptake of molecules and particles, including EVs. Left panel: Cross section of the embryonic cells on day 12 of pregnancy. A Different in size and shape EVs (yellow arrows) covered by a double membrane (higher magnification) were seen attached to the plasma membrane at the apical surface of trophoblast cells. B A merocrine type of secretion was evident at the apical surface of the trophoblast cells, as exemplified by the fusion of numerous MVBs with plasma membrane (black arrows) and vesicles ready to be released into the extracellular space (higher magnification). C The apocrine type of secretion (pink arrows) was also observed, as the cytoplasmic fragments containing the numerous secretory vesicles. Right panel: Cross section of the trophectoderm on day 16 of pregnancy. D Micrograph showing close connections between trophoblast cells through tight junctions. Close to the apical surface of cells, secretory vesicles and endosomes were observed. Apocrine-type secretion (pink arrows) was also observed. E EVs (yellow arrows) attached to the apical surface. F MVBs (black arrow) filled with vesicles were located under the cell membrane at the apical surface. G Probable clathrin-mediated endocytosis was observed. Clathrin-coated pits were formed at the apical surface of the trophoblast cell membrane. Double-membrane EVs were visible inside the clathrin-coated pits (higher magnification). AS – apical surface, APS – apocrine secretion, CCP – clathrin-coated pit, DP – day of pregnancy, EC – endodermal cell, E – endosomes, M – mitochondria, MRS – merrocrine secretion, MV – microvilli, MVB – multivesicula bodies, N – nucleus, RER – rough endoplasmic reticulum, SER – smooth endoplasmic reticulum, TC – trophoblast cell, TJ – tight junctions, UL – uterine lumen, Vs – vesicles