Differentially expressed genes in human trophoblast cells with different degrees of invasiveness

Abstract

s: R.T.C. and T.G.W.M.S. Canada 1996 A.15 Differentially Expressed Genes in Human Jrophoblast Cells with Different Degrees of Invasiveness. G. Huch, H.-P. Hohn, H.-W. Denker; Institute for Anatomie, Universrty of Essen, Medical School, D-45122 Essen, Germany. The majority of molecular control mechanisms of trophoblast invasiveness are unknown so far. We have analyzed gene expression of normal trophoblast cells and of JAr malignant trophoblast cells with differing invasive potential by DDRTPCR (differential-display-RT-PCR). In JAr cells, high invasiveness was induced with phorbol-12-my ristoyl-13-acetate while for low invasiveness cells were treated with dibutyryl-CAMP (Placenta 14, 1993, A44). Normal trophoblast cells were isolated from first trimester (highly invasive) and term placentae (“non”-invasive), According to DDRT-PCR, major differences’in mRNA-patterns were seen between normal and malignant trophoblast cells, between early and term trophoblast cells, but not between JAr cells with different treatments Differentially generated PCR-products of normal trophoblast were reamplified, cloned into the vector pCRII, and sequenced. Data base searches identified only a few of the cloned fragments. One of them isolated from first trimester trophoblast showed 100% homology to j31-integrin which has been suggested on the protein level to be involved in directing trophoblast invasion (Damsky et al., 1994, Development 120, 3657). This result demonstrates that DDRT-PCR is a reliable tool to detect changes in gene expression as related to regulation of trophoblast differentiation/invasiveness. Since RNA probes for cloned PCR-fragments usually representing 3’-regions (mostly untranslated) did not give reliable results in in-situ-hybridizations we are constructing a cDNA library from first trimester human placentae. This and another one from term placenta is being screened for the respective coding regions as a basis for the construction of appropriate probes. Supported by Dr:Mildred-Scheel-Stiftung fiir Krebsforschung Maternal Growth Hormone Treatment Increases Placental Diffusion Capacity but not Fetal or Placental Growth in Sheep. J.E.Harding, P.C.Evans and P.D.Gluckman, Research Centre for Developmental Medicine and Biology, Department of Paediatrics, University of Auckland, Auckland, New Zealand. Growth hormone (GH) treatment increases circulating concentrations of glucose, fatty acids and IGF-1. We hypothesised that maternal GH treatment may enhance fetal and placental growth. Twelve chronically catheterised pregnant sheep were treated with GH O.lmg/Kg SC bd for 10d from 125f0.3d (meanfSE), while 11 controls received saline. GH increased placental capacity for simple diffusion, measured by steady state 14C-urea clearance. Maternal and fetal blood urea fell, but there were no other changes in fetal or placental metabolism. Fetal and placental weights were similar in GH and control groups at 134f0.3d (fetus 4.3f0.2 vs 4.3+0.3Kg, placenta 441f36 vs 411*42g). These data suggest placental diffusion capacity does not limit fetal growth under these circumstances. Human MASH2 (HASH2) Maps lo Chromosome 11~15 and is Expressed in Extravillus Trophoblast. :,Oudeians, M.Alders, J.Postmus, Lvan Wijk, H.Hodgess, A.K.Hadjantonskis, J.Bliek, M.de Meulemeester, A.Westerfeld, P.Little, and MMannens. Departments of Clinical Chemistry, Free University Hospital, Amsterdam, Human Genetics, Univ. of Amsterdam, and Biochemistry, Imperial College London. Mice deficient for the Mash-2 gene coding for the lineage-specific transcription factor of the basic helix-loop-helix (bHLH) family die at 10 days post coitum due to placental failure as a consequence of deficient spongiotrophoblast formation. By screening of chromosome 1 l-specific cosmids with a degenerate FCR probe, the human Mash2 (HASHI) was isolated. By pulsed field gel electrophoresis, HASH2 was found to map to chromosome 11~15.5 proximal to and in close vicinity of IGF.2 in the syntenic human chromosome region of mouse chromosome 7. This region harbours a cluster of imprinted genes (Mash2, Ins-2, IGF-2, and H19). In the bHLH-region, the human gene was 92.7% and 98% homologous to mouse MASH2 at the DNA and peptide levels, respectively. HASH2 expression was studied by non-radioactive RNA in situ hybridization of early human trophoblast cells of normal and androgenetic (molar) origin. in early human placentae, HASH2 expression was found in extravillus trophoblast cells only. Chorionic villas trophoblast, placental stromal cells as well as maternal placental bed giant cells were negative. In trophoblast cells of androgenetic origin, a differential distribution pattern was observed. Extravillus trophoblast cells with malignant potential (invasive moles) were positive, while extravillus trophoblast cells from complete moles without malignant potential were negative. These expression data suggest, although indirectly, that HASH2 is genetically imprinted with preferential activation of the