Isolation and characterization of a bovine trophectoderm cell line derived from a parthenogenetic blastocyst (original) (raw)
Related papers
Placenta, 1995
We have established in culture a feeder-dependent cell line, termed TEI, from a 9 day, pre-implantation, porcine embryo. TEI cells were observed by light and electron microscopy, and characterized by immunocytochemistry: the morphology, cytology and ultrastructure of this cell line are described. The cells display epithelial characteristics, as revealed using immunof2uorescence microscopy with antibody against cytokeratins of simple epithelia, but not with antibody against vimentin. The cells demonstrate many morphological and cytochemical features in common with trophectoderm of the intact porcine blastocyst. For example, TEl cells are polarized and possess tight junctions at their borders, similar to those found in trophectoderm of the pre-implantation embryo. Moreover, TEI cells label positively for the porcine trophectoderm-specz$c monoclonal antibody, SN1/38. Thus, by several important criteria TEI is deduced to be a porcine trophectoderm cell line. 0143--1004/95/070643+16 $12.00/O 0 1995 W. B.
In Vitro Cellular & Developmental Biology - Animal, 2005
A cell line, BPE-1, was derived from a parthenogenetic 8-d in vitro-produced bovine blastocyst that produced a cell outgrowth on STO feeder cells. The BPE-1 cells resembled visceral endoderm previously cultured from blastocysts produced by in vitro fertilization (IVF). Analysis of the BPE-1 cells demonstrated that they produced serum proteins and were negative for interferon-tau production (a marker of trophectoderm). Transmission electron microscopy revealed that the cells were a polarized epithelium connected by complex junctions resembling tight junctions in conjunction with desmosomes. Rough endoplasmic reticulum was prominent within the cells as were lipid vacuoles. Immunocytochemistry indicated the BPE-1 cells had robust microtubule networks. These cells have been grown for over 2 yr for multiple passages at 1:10 or 1:20 split ratios on STO feeder cells. The BPE-1 cell line presumably arose from embryonic cells that became diploid soon after parthenogenetic activation and development of the early embryo. However, metaphase spreads prepared at passage 41 indicated that the cell population had a hypodiploid (2n ϭ 60) unimodal chromosome content with a mode of 53 and a median and mean of 52. The cell line will be of interest for functional comparisons with bovine endoderm cell lines derived from IVF and nuclear transfer embryos.
Trophectoderm differentiation in the bovine embryo: characterization of a polarized epithelium
Reproduction, 1998
Blastocyst formation is dependent on the differentiation of a transporting epithelium, the trophectoderm, which is coordinated by the embryonic expression and cell adhesive properties of E-cadherin. The trophectoderm shares differentiative characteristics with all epithelial tissues, including E-cadherin-mediated cell adhesion, tight junction formation, and polarized distribution of intramembrane proteins, including the Na\p=n-\K ATPase. The present study was conducted to characterize the mRNA expression and distribution of polypeptides encoding E-cadherin, \g=b\-catenin, and the tight junction associated protein, zonula occludens protein 1, in pre-attachment bovine embryos, in vitro. Immunocytochemistry and gene specific reverse transcription\p=n-\polymerase chain reaction methods were used. Transcripts for E-cadherin and \g=b\-catenin were detected in embryos of all stages throughout pre-attachment development. Immunocytochemistry revealed E-cadherin and \g=b\-catenin polypeptides evenly distributed around the cell margins of one-cell zygotes and cleavage stage embryos. In the morula, detection of these proteins diminished in the free apical surface of outer blastomeres. E-cadherin and \g=b\-catenin became restricted to the basolateral membranes of trophectoderm cells of the blastocyst, while maintaining apolar distributions in the inner cell mass. Zonula occludens protein 1 immunoreactivity was undetectable until the morula stage and first appeared as punctate points between the outer cells. In the blastocyst, zonula occludens protein 1 was localized as a continuous ring at the apical points of trophectoderm cell contact and was undetectable in the inner cell mass. These results illustrate that the gene products encoding E-cadherin, \g=b\-catenin and zonula occludens protein 1 are expressed and maintain cellular distribution patterns consistent with their predicted roles in mediating trophectoderm differentiation in in vitro produced bovine embryos.
Molecular Reproduction and Development, 2008
The expression of interferon-tau (IFN-t) is essential for bovine embryo survival in the uterus. An evaluation of IFN-t production from somatic cell nuclear transfer (NT)-embryo-derived primary trophectoderm cultures in comparison to trophectoderm cultured from parthenogenote (P) and in vitro matured, fertilized, and cultured (IVP) bovine embryos was performed. In Experiment 1, the success/ failure ratio for primary trophectoderm colony formation was similar for IVP and NT blastocysts [IVP ¼ 155/ 29 (84%); NT 104/25 (81%)], but was decreased (P ¼ .05) for P blastocysts [54/43 (56%)]. Most trophectoderm colonies reached diameters of at least 1 cm within 3-4 weeks, and at this time, 72 hr conditioned cell culture medium was measured for IFN-t concentration by antiviral activity assay. The amount of IFN-t produced by IVP-outgrowths [4311 IU/mL (n ¼ 155)] was greater (P < .05) than that from NT-[626 IU/mL (n ¼ 104)] and P -[1595 IU/ mL (n ¼ 54)] derived trophectoderm. Differential expression of IFN-t was confirmed by immunoblotting. In Experiment 2, colony formation was again similar for IVP and NT blastocysts [IVP ¼ 70/5 (93%); NT 67/ 1 (99%)] and less (P < .05) for P blastocysts [65/ 27 (70%)]. Analysis of trophectoderm colony size after 23 days in culture showed a similar relationship with P-derived colonies being significantly smaller in comparison to IVP and NT colonies. A differential expression of IFN-t was also observed again, but this time as measured over time in culture. Maximal IFN-t production was found at day-14 of primary culture and diminished to a minimum by the 23rd day. Mol.
PLOS ONE, 2015
Despite being successfully used to produce live offspring in many species, somatic cell nuclear transfer (NT) has had a limited applicability due to very low (>1%) live birth rate because of a high incidence of pregnancy failure, which is mainly due to placental dysfunction. Since this may be due to abnormalities in the trophectoderm (TE) cell lineage, TE cells can be a model to understand the placental growth disorders seen after NT. We isolated and characterized buffalo TE cells from blastocysts produced by in vitro fertilization (TE-IVF) and Hand-made cloning (TE-HMC), and compared their growth characteristics and gene expression, and developed a feeder-free culture system for their long-term culture. The TE-IVF cells were then used as donor cells to produce HMC embryos following which their developmental competence, quality, epigenetic status and gene expression were compared with those of HMC embryos produced using fetal or adult fibroblasts as donor cells. We found that although TE-HMC and TE-IVF cells have a similar capability to grow in culture, significant differences exist in gene expression levels between them and between IVF and HMC embryos from which they are derived, which may have a role in the placental abnormalities associated with NT pregnancies. Although TE cells can be used as donor cells for producing HMC blastocysts, their developmental competence and quality is lower than that of blastocysts produced from fetal or adult fibroblasts. The epigenetic status and expression level of many important genes is different in HMC blastocysts produced using TE cells or fetal or adult fibroblasts or those produced by IVF.
Human Reproduction, 1996
The results of experiments in which horseradish peroxidase (HRP) was used to mark single trophectoderm or inner cell mass (ICM) cells in situ in mouse blastocysts have led to the proposal that growth of the trophectoderm depends on stem cells located in the inner cell mass. Thus, the finding that the visual centre of clones formed following labelling of the central polar trophectoderm cell in early or expanding blastocysts was consistently shifted towards or into the mural trophectoderm was attributed to their displacement by ICM-derived cells. However, the frequency with which central polar cells were displaced is likely to have been overestimated by using the visual centre of descendant clones as the index of their location. Also, the possibility that displacement of central polar cells was an artefact of the marked temporary interruption of their cycling that resulted from labelling was not discounted. Furthermore, no attempt was made to ascertain whether cells located elsewhere in the polar trophectoderm also moved murally, as expected if there is a general displacement of such cells. In the present study, labelling of either the central or a peripheral polar trophectoderm cell with HRP was achieved without obviously perturbing their subsequent proliferation. Moreover, displacement was assessed by recording the location of the proximal boundary rather than the visual centre of the resulting clones. Even by this conservative criterion, the majority of labelled central cells moved towards or into the mural trophectoderm. In marked contrast, however, labelled peripheral polar cells moved murally in only a minority of cases. The remainder either retained their original position or moved towards rather than away from the central polar region. Such an anisotropic pattern of growth of the polar trophectoderm is not readily explicable in terms of recruitment of cells from the ICM. Rather, it accords with the view that the polar trophectoderm is a proliferative centre, and suggests that movement murally of its surplus cells may be restricted circumferentially, possibly through anchorage of the junctional trophectoderm cells that extend processes over the free surface of the ICM.
Trophectoderm Lineage Determination in Cattle
Developmental Cell, 2011
The trophectoderm (TE) and inner cell mass (ICM) are committed and marked by reciprocal expression of Cdx2 and Oct4 in mouse late blastocysts. We find that the TE is not committed at equivalent stages in cattle, and that bovine Cdx2 is required later, for TE maintenance, but does not repress Oct4 expression. A mouse Oct4 (mOct4) reporter, repressed in mouse TE, remained active in the cattle TE; bovine Oct4 constructs were not repressed in the mouse TE. mOct4 has acquired Tcfap2 binding sites mediating Cdx2-independent repression-cattle, humans, and rabbits do not contain these sites and maintain high Oct4 levels in the TE. Our data suggest that the regulatory circuitry determining ICM/TE identity has been rewired in mice, to allow rapid TE differentiation and early blastocyst implantation. These findings thus emphasize ways in which mice may not be representative of the earliest stages of mammalian development and stem cell biology.
Cell Biology International Reports, 1990
Tissue sections of periimplantation pig conceptuses (days 9-15 of pregnancy) were incubated with antiserum to the basic protein (BP), a major secretory protein of filamentous pig blastocysts. Bound antibody was detected by the peroxidase-antiperoxidase method. BP was restricted to trophectoderm in conceptuses which had made the transition from a spherical to an ovoid shape having a diameter of greater than 5 mm (day 11). Tubular (day 11, lo-20 mm) and filamentous (day 11-15) conceptus trophectoderm contained BP. These results suggest that BP synthesis commences at the time of rapid trophoblast growth. 1 ennessee Agricultural Experiment Station.
Differentiation of ICM cells into trophectoderm
The American journal of pathology, 1988
It has been established previously that when inserted in the blastocyst E Ca 247 preferentially differentiates into trophectoderm in vitro. If the concept that tumors are caricatures of the process of tissue renewal is correct, then some cells from the inner cell mass (ICM), the normal counterpart of embryonal carcinoma, should be able to differentiate into trophectoderm. This has been a controversial issue. Four experiments are now reported that support the idea that ICM can differentiate into trophectoderm: 1) ICM from early blastocysts after classical immunosurgery made blastocysts in vitro; 2) ICM obtained from early blastocysts by immunosurgery using antigens other than histocompatibility ones made blastocysts in vitro; 3) ICM from early blastocysts, in which the trophectodermal cells had been labeled, contained no labeled cells following immunosurgery; and 4) In reconstruction experiments, polar and mural trophectodermal cells attached to ICM from late blastocysts failed to mu...
Trophectoderm in Control of Murine Embryonal Carcinoma
Cancer Research, 1984
It has been shown previously that the intact blastocyst of the mouse can regulate tumor formation and colony formation of murine embryonal carcinoma. This effect is consistent with the close histogenetic correspondence between embryonal carci noma and the inner cell mass of the blastocyst. The ability of inner cell mass, blastocele fluid, and inner and outer surfaces of trophectoderm to abrogate colony formation of a variety of malignant tumors has now been tested. Direct contact of the embryonal carcinoma cells with the blastocele surface of tro phectoderm proved to be necessary for abrogation of colony formation of embryonal carcinoma. This effect was not seen with any of the other tumors tested. Some tumors, which lack a normal cellular counterpart in the blastocyst, grew poorly in the blastocele unless a fistula was made in the wall of the blastocyst. Colony formation of the embryonal carcinoma was regulated in blastocysts with fistulas, but the other tumors were not regulated under these conditions. It is concluded that colony formation of embryonal carcinoma cells is regulated by direct contact with the trophectoderm of its corresponding embryonic field in an unknown but specific manner.