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PubMed1998

Differential expression of p57kip2, a maternally imprinted cdk inhibitor, in normal human placenta and gestational trophoblastic disease.

Evidence has recently been provided to support a role for genomic imprinting in the regulation of embryonic implantation and development and placental growth, as well as in the pathogenesis of proliferative trophoblastic diseases. The cyclin-dependent kinase inhibitor p57KIP2 has recently been recognized as a maternally imprinted gene. We investigated p57KIP2 expression in first-trimester normal placentas from interrupted pregnancy, spontaneous abortions, and different types of proliferative trophoblastic diseases using single- and double-marker immunohistochemical techniques. In normal placenta, nuclear p57KIP2 expression was observed at high frequency (up to 100%) in extravillous trophoblast, cytotrophoblast, and implantation-site interstitial trophoblast, but was absent in syncytiotrophoblast. p57KIP2 was also expressed in the stromal cells of maternal decidua, which was one of the few adult tissues retaining p57KIP2 expression (most other adult tissues investigated were negative). p57KIP2 expression was either absent or low in all cases of diploid/tetraploid complete moles (20 cases) and in three cases of gestational choriocarcinoma. On the other hand, all spontaneous abortions (12 cases) and triploid partial moles (19 cases) showed p57KIP2 levels comparable to those observed in normal placenta. These findings are in line with the hypothesis that deregulation of genomic imprinting, particularly the loss of cell-cycle inhibitors such as p57KIP2, is involved in the abnormal development of androgenetic trophoblastic proliferations. In addition, this simple immunohistochemical analysis could provide a useful diagnostic marker in difficult cases.

Choriocarcinoma↗
PubMed2001

Genetic and developmental analysis of X-inactivation in interspecific hybrid mice suggests a role for the Y chromosome in placental dysplasia.

It has been shown previously that abnormal placental growth, i.e., hyper- and hypoplasia, occurs in crosses and backcrosses between different mouse (Mus) species. A locus that contributes to this abnormal development has been mapped to the X chromosome. Unexpectedly, an influence of fetal sex on placental development has been observed, in that placentas attached to male fetuses tended to exhibit a more pronounced phenotype than placentas attached to females. Here, we have analyzed this sex dependence in more detail. Our results show that differences between male and female placental weights are characteristic of interspecific matings and are not observed in intraspecific Mus musculus matings. The effect is retained in congenic lines that contain differing lengths of M. spretus-derived X chromosome. Expression of the X-linked gene Pgk1 from the maternal allele only and lack of overall activity of two paternally inherited X-linked transgenes indicate that reactivation or lack of inactivation of the paternal X chromosome in trophoblasts of interspecific hybrids is not a frequent occurrence. Thus, the difference between male and female placentas seems not to be caused by faulty preferential X-inactivation. Therefore, these data suggest that the sex difference of placental weights in interspecific hybrids is caused by interactions with the Y chromosome.

Animals↗
PubMed1990

Human neuraminidase is a 60 kDa-processing product of prosaposin.

Human neuraminidase was purified from placenta as part of a large molecular weight complex with lysosomal beta-galactosidase and carboxypeptidase. Passage of this purified complex through a sialic acid-affinity column (fetuin-agarose) retained a minor 60 kDa protein which was eluted with 100 mM N-acetylneuraminic acid. This 60 kDa protein is recognized in Western blots of the purified complex by an anti-prosaposin antibody which at the same time was able to inhibit neuraminidase activity in the preparation. Furthermore, probing of cultured skin fibroblasts of patients affected with neuraminidase deficiency using the antiprosaposin antibody revealed an abnormal 57 kDa protein. These results indicate that the 60 kDa protein is derived from prosaposin and has the characteristics of a neuraminidase.

Antibodies↗
PubMed2002

Baboon placentas express soluble and membrane-bound Paan-AG proteins encoded by alternatively spliced transcripts of the class Ib major histocompatibility complex gene, Paan-AG.

The human class Ib major histocompatibility complex (MHC) molecule, HLA-G, is unique in its limited polymorphism, high expression in the placenta and generation of multiple transcripts by alternative splicing. The proteins encoded by these transcripts are believed to modulate maternal-fetal immunological relationships during pregnancy. The baboon placenta contains messages encoded by a novel MHC gene, Paan-AG, which is evolutionarily related to the HLA-A locus, but shares unique characteristics with HLA-G. In this study, we show that the Paan-AG message is alternatively spliced to generate at least seven transcripts. One of these transcripts retains intron 4 and encodes a soluble glycoprotein with three external domains and a unique 21-amino-acid sequence at the carboxyl terminus, similar to soluble HLA-G1. This glycoprotein was detected in first trimester placental villous cytotrophoblast and syncytiotrophoblast, and in extravillous cytotrophoblast cells in the basal plate in term placenta. Four of the transcripts ( Paan-AG1, Paan-AG2, Paan-AG3, Paan-AG4) encode membrane-bound class Ib MHC glycoprotein isoforms. Paan-AG1 protein expression was similar to that of sPaan-AG, while Paan-AG2 protein was not detected in these tissues. The other two transcripts ( Paan-AGx and Paan-AGxi) contain a truncated exon 3 and multiple stop codons. Paan-AG1 and Paan-AGx transcripts were detected in a number of non-placental tissues, but these transcripts contained multiple stop codons. Because of the structural similarities and common features of organ-specific expression and splicing of the message, studies on Paan-AG may be of value in dissecting the functions of the class Ib proteins in human pregnancy.

Alternative Splicing↗
PubMed2005

Stem cell characteristics of amniotic epithelial cells.

Amniotic epithelial cells develop from the epiblast by 8 days after fertilization and before gastrulation, opening the possibility that they might maintain the plasticity of pregastrulation embryo cells. Here we show that amniotic epithelial cells isolated from human term placenta express surface markers normally present on embryonic stem and germ cells. In addition, amniotic epithelial cells express the pluripotent stem cell-specific transcription factors octamer-binding protein 4 (Oct-4) and nanog. Under certain culture conditions, amniotic epithelial cells form spheroid structures that retain stem cell characteristics. Amniotic epithelial cells do not require other cell-derived feeder layers to maintain Oct-4 expression, do not express telomerase, and are nontumorigenic upon transplantation. Based on immunohistochemical and genetic analysis, amniotic epithelial cells have the potential to differentiate to all three germ layers--endoderm (liver, pancreas), mesoderm (cardiomyocyte), and ectoderm (neural cells) in vitro. Amnion derived from term placenta after live birth may be a useful and noncontroversial source of stem cells for cell transplantation and regenerative medicine.

Amnion↗
PubMed1979

Differentiated regions of human placental cell surface associated with attachment of chorionic villi, phagocytosis of maternal erythrocytes and syncytiotrophoblast repair.

Scanning electron micrographs of human placental cell surface show: (1) Differentiated zones of trophoblast which may be covered by fewer 'microvilli' than the adjacent syncytial cell surface and which extend as a narrow, usually distal protrusion of the chorionic villus. This narrow outgrowth terminates as a fractured end. Presumably since preparations were obtained from therapeutic terminations of pregnancy or Caesarian deliveries these broken ends represent the yield point in the anchoring 'villi' ruptured as a result of surgery. Similar anchoring 'villi' with fractured ends were observed in unfixed material with the use of Nomarski interference contrast microscopy. (2) It appears that, during apparent phagocytic uptake of maternal erythrocytes by syncytiotrophoblast, cell surface lining the forming vacuole still retains an irregular microvillous surface. This observation indicates the potential location of phagocytosis receptors for red blood cells in the placental cell surface. (3) Areas of human placenta which appears to have been damaged and may be undergoing repair exhibit masses of cells with conspicuous microvillar cell surfaces. The origin of these cells is discussed in relation to the usual processes of syncytiotrophoblast formation.

Cell Membrane↗
PubMedDate not supplied

Characterization of antibodies to a placenta-specific antigen cross-reacting with a choriocarcinoma cell line.

Antibodies were raised in rabbits to purified human term placental villous plasma membrane. These were cytotoxic to human peripheral blood lymphocytes and manifested cross-reactivity to kidney and liver. After absorption with these tissues, reactivity was retained with placental villous plasma membrane. The placental-specific antiserum was fully reactive with BeWo choriocarcinoma cells, but was devoid of reaction with a human myeloma tumor cell line (GM2132). The antibodies were cytotoxic to BeWo in the presence of complement, as determined by trypan blue uptake, 51Cr release, and inhibition of uptake of 3H-thymidine.

Animals↗
PubMed1990

Placenta increta in the second trimester of pregnancy: report of a case.

A rare case of placenta increta in the second trimester of pregnancy is reported. The patient was at 15 weeks gestation when the pregnancy, which had been complicated by a maternal rubella infection, was terminated at a regional hospital. However, vaginal bleeding persisted after the operation in spite of medication to control bleeding. Curettage of the uterine cavity one month later failed to reveal any retained placental tissue or other pathology. Therefore, an exploratory laparotomy was performed, yet nothing particular was found in the peritoneal cavity. So, the patient was transferred to our department. Sonography revealed a lower uterine mass of 4.0 x 3.3 cm in size. A persistently low serum hCG titer was also found. Placenta accreta was highly suspected. Three doses of methotrexate were given to control bleeding, yet without results. Hysterectomy was finally performed. A histological study revealed placenta increta.

Adult↗
PubMed1990

Binding of thiopental in neonatal serum.

Protein binding of thiopental was studied in 21 samples of neonatal serum (from placental blood) and compared with protein binding in ten healthy volunteers. These infants ranged between 32 and 43 weeks of gestational age (mean, 37.7 weeks) and the adult age range was from 27 to 54 years (mean, 35.4 years). Because the unbound fraction of the drug is responsible for its pharmacologic effect, a marked difference in the protein binding between neonates and adults may be relevant to the clinician. Blood obtained from freshly delivered placentas or from adult volunteers was allowed to clot and the serum separated from the sample. A portion of the serum was sent for protein and bilirubin analysis and the remainder retained for study. This latter serum was combined with four concentrations of thiopental. These specimens were then ultrafiltered and the amount of thiopental in the ultrafiltrate (unbound) compared with the prefiltered amount (total), as measured by reverse-phase high-performance liquid chromatography. The binding studies were repeated at pH 7.2, 7.4, and 7.6 in both the adult and neonatal serum. Total protein and albumin are significantly less in neonatal serum, whereas bilirubin (total and direct) is significantly higher in neonatal serum than in adult serum (P less than 0.01). Neonatal serum was associated with significantly more unbound thiopental than adult serum at all levels of pH studied (P less than 0.005). Increasing the pH resulted in less free drug in both groups, but this reached statistical significance only in the adult group (P less than 0.025). Drug concentration had no effect on binding in the range examined.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1988

Two systems in vitro that show insulin-stimulated serine kinase activity towards the insulin receptor.

Two systems in vitro are described that show insulin-stimulated phosphorylation of the insulin receptor on serine residues. In the first system, insulin receptor was purified partially from Fao rat hepatoma cells by direct solubilization of the cells in Triton X-100 and chromatography on wheat-germ-agglutinin-agarose. Phosphorylation of these preparations with [gamma-32P]ATP in the presence or absence of insulin resulted in 32P incorporation exclusively into phosphotyrosine residues. Serine kinase activity towards the insulin receptor was reconstituted by adding extracts of Fao cells. Prior exposure of the cells to insulin stimulated serine kinase activity towards the insulin receptor in extracts 7.2-fold. A receptor serine kinase activity enhanced by treatment of cells with cyclic AMP analogues was also retained in the reconstituted system. In the second system, insulin receptor and insulin-sensitive serine kinase activity towards the insulin receptor were co-purified from human placenta. The protocol involved preparation of membranes, before solubilization and chromatography on wheat-germ-agglutinin-agarose, by using gentle procedures designed not to disrupt a potentially labile association between the insulin receptor and the serine kinase. Serine kinase activity in these preparations towards the insulin receptor was stimulated up to 10-fold by insulin, and the stoicheiometry of serine phosphorylation was estimated to be approx 0.8 mol/mol of insulin receptor for phosphorylations performed in the presence of insulin. Thus a preparation of insulin receptor is described for the first time that is phosphorylated to high stoicheiometry on serine in an insulin-dependent manner. Conditions that facilitate recovery and assay of serine kinase activity are defined and discussed. These systems provide a basis for characterizing the nature of the insulin-sensitive serine kinase that phosphorylates the insulin receptor, and defining its role in insulin action and control of receptor function.

Animals↗
PubMed1990

Changes in the proportion of type I and type III collagen in the developing and retained bovine placentome.

The proportions of Type I and Type III collagen were evaluated from gestational, postpartum-retained, and released bovine placental membranes. Placentomes were excised at 90, 150, 210, and 270 days of gestation (n = 32) and from postpartum-retained (2 and 12 h, n = 8) and released (2 h, n = 4) membranes. Placentome components were processed for collagen, hydroxyproline, protein, and dry weight determination. Collagen extracts were separated by SDS-PAGE. Densitometry was used to establish the proportions of collagen alpha chains (Type I = 2 alpha 1 + 1 alpha 2; Type III = 3 alpha 1). No difference in the proportion of maternal caruncular Type I and Type III collagen was found. The proportion of Type I fetal cotyledonary collagen was lowest (p less than 0.05) at Day 90 of gestation but did not differ between Days 150, 210, 270, or between retained and released fetal membranes. The proportion of Type III fetal cotyledonary collagen was greatest (p less than 0.05) at Day 90. Retained fetal cotyledons had a greater (p less than 0.05) proportion of Type III collagen than did released fetal cotyledons. Therefore, although hydroxyproline content was not different between retained and released fetal membranes, the retained bovine fetal cotyledon was characterized by disproportionate amounts of Type III collagen as compared to the fetal cotyledon that was not retained.

Animals↗
PubMed1987

Phosphorylation of alpha-tubulin carboxyl-terminal tyrosine prevents its incorporation into microtubules.

Insulin receptor kinase phosphorylated tubulin in an insulin-dependent fashion. Two different populations of phosphotubulin were found. In tubulin dimers containing tyrosine at the carboxyl-terminal of their alpha subunit, phosphate was incorporated in that residue, and the phosphorylated protein did not assemble into polymers. In tubulin dimers lacking this tyrosine residue, phosphate was incorporated into different tyrosine residues located in other parts of the molecule, and the phosphoprotein retained its capacity to polymerize.

Animals↗
PubMed1991

Characterization of endocrine cell lines immortalized by a temperature-sensitive mutant SV40.

An ideal in vitro model for the study of endocrine functions would be one in which cells could propagate in culture and express their specialized functions. Most endocrine studies to date have relied on primary cell culture or on the use of tumor cell lines. This report describes the characterization of three endocrine cell lines immortalized by transfecting endocrine cells with a temperature-sensitive mutant SV40 virus. Rabbit endometrium (HRE-H9), human placenta (SPA209-10) and rat pituitary (RP) cells were immortalized by SV40 virus, a temperature-sensitive (ts) mutant in the A gene, which encodes the large tumor antigen that is required for the maintenance of transformation. The transformed phenotype of the SV40 tsA mutant-immortalized cell line can be reversed simply by a shift in temperature. At the permissive temperature (34 degrees C), all three types of cells exhibited a transformed phenotype, which is characterized by high cell density growth and by the overgrowth of nontransformed cell layers. However, at the non-permissive temperature (40 degrees C) these cells reverted to a non-transformed phenotype as demonstrated by a marked decrease in the overgrowth of nontransformed layers and by the expression of differentiated functions. At the non-permissive temperature (40 degrees C), the endometrial cell line was capable of synthesizing beta-endorphin, and it exhibited hormonally regulated expression of the transfected hybrid uteroferrin gene construct. The human placenta cell line was capable of secreting GnRH upon stimulation by cAMP, forskolin, theophyllin, PGE, catecholamine and Ca++ channel stimulators. Moreover, the rat pituitary cell line was capable of synthesizing and secreting growth hormone (GH) which was stimulated by GHRH and cAMP. The advantage of the temperature-sensitive cell lines is that a single cell line is the source of both the normal and transformed states; thus, studies are internally controlled. These results demonstrate that tsA mutants of SV40 virus are the best available agents for immortalizing mammalian endocrine cells that retain differentiated functions.

Animals↗
PubMed1986

The nucleotide sequences of baboon chorionic gonadotropin beta-subunit genes have diverged from the human.

The placental glycopeptide hormone chorionic gonadotropin (CG) is involved in establishing and maintaining pregnancy. The hormone consists of two different non-covalently associated subunits termed alpha and beta. In man there are seven closely linked genes coding for beta CG-like peptides, but only three of these appear capable of expression in the placenta. The organization of beta CG-like genes in man and baboon appears to be similar. We demonstrate here that the baboon genome contains multiple copies (at least five) of beta CG-related genes, and that these genes are closely linked in the genome. Nucleotide sequence data from several beta CG cDNA clones indicates that at least two of these beta CG-related genes are expressed in the baboon placenta. Analysis of beta CG sequences from baboons and human subjects demonstrates that these genes have diverged markedly between species. In contrast, these sequences are remarkably homogeneous within their respective genomes. Gene conversion events may be responsible for retaining the high degree of identity among the various beta CG gene family members. Knowledge of beta CG sequences from baboon may lead to development of a long-term antipregnancy vaccine. The ability of CG antibodies to interfere with the maintenance of pregnancy can now be investigated within a homologous system.

Animals↗