Trophectoderm mechanics direct epiblast shape upon embryo implantation

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Sex ratio trajectory in mouse

Skewing of the sex ratio towards males occurs in humans. The possible explanation for excess male births could be a preference for Y-bearing sperm at fertilization and/or selective elimination of female embryos during pregnancy. In this study, we have tested the sex ratio in the preimplantation embryo (2–3 cells stage/closest possible primary sex ratio), the post-implantation embryo (day E7.5), and at birth (secondary sex ratio) on a homogenous (genetic, environmental, and dietary) population of mice to ascertain the biological reason i.e., male preference at fertilization or female elimination during pregnancy or both. Primary sex ratio on early preimplantation embryos (2–3 cells stage) was studied on 598 embryos and secondary sex ratio (at birth) on 721 pups using PCR-based sexing (both X & Y chromosome-specific) besides sex ratio of 80 post-implantation embryos (day E7.5). We have also investigated whether the fat content (high & low) of the diet affects the sex ratio. We observed a skewed sex ratio (more female) in preimplantation embryos (0.436; 95 % CI 0.39, 0.48), and post-implantation embryos (0.462; 95 % CI 0.35, 0.57) but reverse skewing (more male) at birth (0.539; 95 % CI 0.5, 0.58). We also observed that high-fat diet promoted male sex ratio at birth (0.657; 95 % CI 0.57, 0.74) whereas a low-fat diet had the opposite effect (0.46; 95 % CI 0.36, 0.56) but no effect at fertilization (2–3 cells stage embryos). This indicates selective elimination of female embryo and fetus throughout pregnancy in mice, more so with a high-fat diet.     

The major role of glutathione in the metabolism and excretion of N,N-dimethyl-4-aminoazobenzene in the rat

In the normal rat given a single dose of one mg N,N-dimethyl-4-aminoazobenzene (DAB) via the hepatic portal vein the following biliary metabolites reached their maximal rates of excretion in the sequence: 4'-sulphonyloxy-DAB, N-(glutathione-S-methylene)-4-aminoazobenzene (GSCH2AB), 4'-sulphonyloxy-N-methyl-4-aminoazobenzene (4'-sulphonyloxy-MAB) 4'-sulphonyloxy-GSCH2AB and MAB-4'-beta-glucuronide. The unusual and relatively unstable N-methylene glutathione conjugates were major metabolites accounting for up to 70% of the whole. It was shown that all the 4-aminoazobenzene (AB) and perhaps all of the 4'-sulphonyloxy-AB, which may be observed in bile, are artefacts due to decomposition of GSCH2AB and 4'-sulphonyloxy-GSCH2AB respectively and that biliary excretion of N-methyl oxidised products of MAB and 4'-hydroxy-MAB is dependent on their conversion to the GSH conjugates, GSCH2AB and 4'-hydroxy-GSCH2AB respectively. Sulphotransferase inhibition by pentachlorophenol caused a reduction in the excretion of all sulphate conjugates, but biliary excretion as a whole was not reduced significantly due to a compensatory increase in the excretion of MAB-4'-beta-glucuronide and the appearance of 4'-OH-GSCH2AB. Glutathione (GSH) depletion by diethylmaleate caused a reduction in biliary metabolites of DAB by lowering the levels of GSH conjugates. This was because the amount of N-methyl oxidation of MAB and 4'-hydroxy-MAB were proportional to the amount of GSH present. The fall in N-methyl oxidation was not compensated for by an increase in 4'-hydroxylation and was accompanied by a delay in the appearance of 4'-hydroxylated metabolites. The administration of potential precursors of 4'-sulphonyloxy-GSCH2AB establishes the sequence of reactions resulting in its formation to be 4'-hydroxylation, N-methyl oxidation, GSH conjugation and O-sulphation.     

In vitro attachment and symmetry breaking of a human embryo model assembled from primed embryonic stem cells

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Cytoskeletal control of the apical surface transformation of rat uterine epithelium

Summary— During early pregnancy, in the lead up to blastocyst implantation, the apical cell surface of luminal epithelial cells of the rat uterus undergo a dramatic shape transformation. This study aims to investigate the role of the cytoskeleton in this apical transformation by considering the effects of the drugs cytochalasin D and colchicine on the uterine luminal cell surface. The results are determined using transmission and scanning electron microscopy. In vivo exposure to cytochalasin D during oestrus, as well as on day 1 of pregnancy, did not affect the long, regular surface microvilli. This drug, however, did disrupt the terminal web within the apical cytoplasm of these cells. Disruption of microfilament (MF) polymerization by cytochalasin D on day 4 of pregnancy induced a cell surface transformation, resulting in the appearance of numerous irregular projections normally present during blastocyst implantation on day 6 of pregnancy. Colchicine did not alter the uterine microvilli of oestrus or day 1 pregnant tissue. Unlike the effect of cytochalasin D, colchicine-induced microtubule (MT) disruption on day 4 of pregnancy did not increase irregular projections and hence this treatment did not result in the cell surface appearance associated with blastocyst implantation. These results indicate that the disruption of MF, rather than MT, contributes to the transformation of the uterine luminal cell surface during the lead up to blastocyst attachment.     

A strategy for the suppression of tumorigenesis induced by biomaterials: Restoration of transformed phenotype of polyetherurethane-induced tumor cells by Cx43 transfection

Biomaterials such as polyetherurethans (PEUs) are the scaffolding, which is indispensable for the development of the bio-artificial organs. However, PEUs can induce tumors in subcutaneous implantation sites in rat. We have shown that the different inhibitory potential of gap junctional intercellular communication (GJIC) on the surface of the biomaterials, including PEUs, is a key step in determining the tumorigenic potential. Here we show that suppression of a gap junctional protein connexin 43 (Cx43) plays an important role in in vivo tumorigenesis induced by PEUs for the first time and that Cx43 transfection may be an effective strategy for preventing tumorigenesis induced by biomaterials. Rat tumor cell line U41 is derived from tumors in the subcutaneous implantation of PEU films. The GJIC and the expression of Cx43 were suppressed in U41. The restoration of normal phenotype, such as reduction of growth rate, recovery of contact inhibition and loss of colony formation ability in soft agar, was achieved by Cx43 transfection. These results strongly suggest that suppression of Cx43 expression plays an important role in the development of rat malignant fibrous histiocytoma (MFHC) caused by PEUs and that Cx43 transfection is effective for prevention of tumorigenesis induced by PEUs. This revised version was published online in August 2006 with corrections to the Cover Date.