Chimera-competent eXtra-Embryonic eNdoderm (XEN) cells established from pig embryos

In this article, we report for the first time the derivation and characterization of extra-embryonic endoderm (XEN) cells from primitive endoderm (PrE) of porcine (p) embryos. The pXEN cells can be reliably and reproducibly generated from parthenote, in vitro and in vivo derived embryos. The pXEN cells retained all the hallmarks of PrE including expression of canonical PrE and XEN cell markers (GATA4, GATA6, SOX17, SALL4, FOXA2, and HNF4A). Transcriptome analysis further confirmed their XEN cell origin. The pXEN cells when introduced into blastocyst stage embryo contributed to wide-spread chimerism including visceral yolk sac, chorion, as well as embryonic gut and liver primordium in the fetus. The pXEN cells were shown to be an efficient nuclear donor for generating cloned offspring. Taken together, pXEN cells fulfil a longstanding need for a stable, chimera-competent, and nuclear transfer-compatible porcine embryonic cells with applications for agriculture and medicine.Significance StatementWe report for the first time, the derivation and characterization of extraembryonic endoderm (XEN) stem cells from porcine (p) embryos. The pXEN cells can be reliably and reproducibly derived from primitive endoderm precursors. When injected into blastocyst-stage embryos, the pXEN cells have contributed to wide-spread chimerism including visceral yolk sac, chorion of the extraembryonic membranes, as well as definitive endoderm of the fetus, primarily the embryonic gut and liver primordium. Additionally, these XEN cells have proven to be an efficient nuclear donor for generating cloned offspring. These newly discovered stem cells provide a novel model for studying lineage segregation, as well as a source for interspecies chimeras for generating endodermal organs, and for genome editing in livestock.

113 INTERACTIONS BETWEEN THE VISCERAL YOLK SAC PLACENTA AND THE UTERINE WALL IN NECROMYS LASIURUS (RODENTIA, CRICETIDAE)—AN EXPERIMENTAL MODEL TO UNDERSTANDING THE MATERNAL–FETAL EXCHANGE

The establishment of pregnancy results from the interaction between the trophoblast and maternal tissues. Evolutionarily, the yolk sac (YS) is the only fetal membrane present in all vertebrate taxa. In mammals, it results in the development of the YS placenta, which is responsible for maternal–fetal exchange in the beginning of gestation. Its functions are related to hematopoiesis, selective transfer of various substances including proteins, iron transfer and histiotrophic nutrition. Necromys lasiurus is a rodent species from South America, which is related to species which are used as laboratory animals. Herein, we show data on the early development of the YS to promote a better understanding of its relationship with the uterine wall. Ten implantation sites of Necromys were obtained from a breeding group at the UFERSA, Mossoró, Brazil. Samples were examined by means of histology, immunohistochemistry for vimentin (1:200) and Proliferating Cell Nuclear Antigen (PCNA; 1:800) and scanning and transmission electron microscopy. In early pregnancy the visceral YS was in places closely attached to the uterine wall. At these attachments, the uterus possessed infolded structures with a high-columnar epithelium associated with blood vessels and surrounded by vimentin positive glands. In these areas, the YS became compact in shape and we observed an intimate association of both visceral yolk sac and endometrial luminal epithelium. A strong response to periodic acid Schiff, including plenty of granular vesicles, indicated secretory activity of the endodermal cells, such as release of histiotrophe. Large intercellular spaces, vacuoles and glycogen were present in the cytoplasm of these cells. On the other hand, in areas where the visceral YS was near to the labyrinthine region of chorioallantoic placenta, it was highly villous. The villi were formed by endoderm cells with hexagonal shape. The visceral YS was supplied by fetal vessels, which form the blood islands, structures related to the fetal hematopoiesis. A prominent microvilli area was observed on the surface of the endoderm cells. Additionally, vacuoles, electron-dense inclusions and dense droplets indicative of high secretory and transfer activities were seen inside of these cells. According to PCNA labelling, the mesothelial layer in the region near to uterine consisted of proliferative cells. Although early gestation represents a crucial developmental stage, information about the interaction between the trophoblast and maternal tissues is sparse. Embryonic mortality, a major cause of reproductive failure in livestock species, can be related to interactive problems between maternal and fetal tissues that affect exchange of nutrients, hematopoiesis and other vital functions. Supported by grants from FAPESP (Proc. 09/53392-8).