Reduction in serum requirement for culture of primary human amniotic fluid cells can be achieved by the addition of 10 growth-promoting factors to the nutrient medium. This supplemented medium preserves cell types normally found in amniotic fluid cell cultures supplemented with 20-30 per cent fetal bovine serum. The volume of amniotic fluid required to initiate culture can be as little as 1 ml. Amniotic fluid samples contaminated with red blood cells with no visible clot also grow well in the low serum medium. Cell-free amniotic fluid combined with equal parts of supplemented medium is useful in initiating cell culture.

Amniocentesis↗

PubMedDate not supplied

Placental transfer of the soy isoflavone genistein following dietary and gavage administration to Sprague Dawley rats.

Genistein, the principal soy isoflavone, has estrogenic activity and is widely consumed by humans for putative beneficial health effects. The goal of the present study was to measure placental transfer of genistein in rats as a possible route of developmental exposure. Pregnant Sprague-Dawley rats were administered genistein orally, either by diet or by gavage. Concentrations of genistein aglycone and conjugates were measured in maternal and offspring serum and brain using HPLC with isotope dilution electrospray tandem mass spectrometry. Although fetal or neonatal serum concentrations of total genistein were approximately 20-fold lower than maternal serum concentrations, the biologically active genistein aglycone concentration was only 5-fold lower. Fetal brain contained predominately genistein aglycone at levels similar to those in the maternal brain. These studies show that genistein aglycone crosses the rat placenta and can reach fetal brain from maternal serum genistein levels that are relevant to those observed in humans.

Administration, Oral↗

PubMedDate not supplied

Effects of phenoxyacetic acids on rat liver tissues.

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2,4,5-Trichlorophenoxyacetic Acid↗

PubMedDate not supplied

The heavy chain of human B-cell alloantigen HLA-DS has a variable N-terminal region and a constant immunoglobulin-like region.

The HLA-D region of the major histocompatibility complex (MHC) of man encodes polymorphic glycoproteins found predominantly on the cell surfaces of B cells and macrophages. These proteins mediate interactions, required for the induction of immune responses, among cells of the immune system and consequently are referred to as Ia (immune-response associated). Two families of Ia molecules, DR and DS (also known as DC), have been defined, the former analogous to the I-E (ref. 1) and the latter to the I-A molecules of the murine MHC. Both DR and DS molecules consist of two noncovalently associated polypeptide chains with molecular weights of 33,000 and 28,000, designated alpha and beta, respectively. The polymorphism of DR molecules is due to structural variation in the small subunit, DR beta, with the large subunit, DR alpha, being constant in structure. In contrast, both subunits DS alpha and DS beta are structurally variable when DS allotypes are compared. We have now isolated a cDNA clone from a DR7 cell line that contains the entire coding sequence for the DS alpha subunit and have compared its predicted amino acid sequence with that previously deduced from a DS alpha cDNA clone isolated from a DR4,w6 cell line. This comparison reveals that 10 of 11 amino acid differences are located within the alpha 1 (N-terminal) domain and that the alpha 2 or immunoglobulin-like domains are identical.

Amino Acid Sequence↗

PubMedDate not supplied

Structural organization of the rat thy-1 gene.

Thy-1 is a differentiation marker expressed predominantly on thymocytes, T cells and brain tissue. Its presence on murine peripheral T cells but not B cells has long been used to distinguish between these two populations of lymphocytes. Although analogues of Thy-1 have been described in several mammalian species, its tissue distribution in different species varies widely, precluding its use as T-cell-specific marker. The Thy-1 molecule is a cell-surface glycoprotein of relative molecular mass 18,000, one-third of which represents carbohydrate; the protein moieties of the rat and murine Thy-1 molecules have been sequenced and found to consist of 111 and 112 amino acids, respectively. An unusual aspect of Thy-1 is the apparent absence of a hydrophobic segment comparable to that observed in other membrane glycoproteins which would allow integration of Thy-1 within the membrane lipid bilayer. This has prompted speculation that Thy-1 is anchored to the cell surface by some other hydrophobic component such as glycolipid. Here we report the structure of thy-1 complementary DNA and genomic clones and describe the exon-intron organization of the gene. More importantly, our data indicate that Thy-1 is initially synthesized as a molecule of 142 amino acids, 31 amino acids longer at the carboxyl end than the Thy-1 molecule isolated and characterized by Campbell et al. An extremely hydrophobic region of 20 amino acids lies within this 31-amino acid stretch and may represent the transmembrane segment responsible for anchoring Thy-1 to the cell membrane.

Amino Acid Sequence↗

PubMedDate not supplied

Schedule to inject in vitro matured oocytes may increase pregnancy after intracytoplasmic sperm injection.

To ascertain the value of using immature oocytes in an intracytoplasmic sperm injection (ICSI) program, the authors designed a schedule, at 5 p.m. on day 1 (the day of oocyte retrieval) and at 8 a.m. and 2 p.m. on day 2, to recognize and inject the in vitro matured (IVM) oocytes. For the 1,166 oocytes retrieved in 107 ICSI cycles, 128 (11.0%) were at the stage of metaphase I (MI) and 113 (9.7%) at germinal vesicle. Routine ICSI for metaphase 11 oocytes was performed at 2 p.m. on day 1 (initial ICSI). In culture medium of human tubal fluid with 15% maternal serum, 85.1% (205/241) immature oocytes progressed to maturation in which 16.4% (21/128) of MI oocytes matured at 5 p.m. of day 1. The rate of normal fertilization for IVM oocytes (58.5%) was not significantly different from that of initial ICSI (64.0%). One patient received a transfer of two fertilized IVM oocytes alone that were injected at 5 p.m. of day 1, maturing from the MI stage, and achieved a normal pregnancy. The fertilized IVM oocytes were replaced along with the embryos from initial ICSI for 40 cycles that led to 14 (35%) clinical pregnancies. In 43 fertilized IVM oocytes donated for research, we observed that cleavage (95.3%) to the 2- to 4-cell stage was not distinct from that of initial ICSI (94.6%); however, the percentage of embryos of grade I and II morphology was significantly smaller (24.4% vs. 62.5%). Only five (11.6%) developed to blastocysts in vitro. Twenty-one fertilized IVM oocytes were frozen for future transfer. A schedule to inject IVM oocytes in ICSI cycles may generate more accessible embryos for fresh transfer or cryopreservation to increase the chance of pregnancy, although the embryo quality was relatively poor.

Adult↗