Human Teratocarcinoma-derived Cells Research Articles

Flow cytometric evaluation of cell‐cycle progression in ethyl methanesulfonate and methyl methanesulfonate‐exposed P3 Cells: Relationship to the induction of sister‐chromatid exchanges and cellular toxicity

In order to determine the relationships among the reduction in relative cloning efficiency (RCE), sister-chromatid exchange (SCE) formation, and interference with progression through the cell-cycle, human teratocarcinoma-derived (P3) cells were exposed to either ethyl methanesulfonate or to methyl methanesulfonate. The relationship between SCE and toxicity was quantified, the progression through the cell-cycle was evaluated with flow cytometric methods, and the effects of these chemicals on cell growth and average generation time (AGT) were determined. A strong correlation existed between RCE and SCE (r = -0.978, p less than .001) which was accompanied by an inhibition of growth as evidenced by a significant (p less than .0001) negative linear effect of concentration on the relative cell count from 24 to 72 hours after exposure and by a concentration-dependent increase (p less than .0001) in the AGT. Delays in the transit through S-phase were evident 4 hours after exposure to toxic concentrations of either carcinogen and by 8 to 12 hours post-exposure at the lower concentrations. Increases in the percentage of nuclei in G2 + M, indicative of G2 arrest, occurred from 12 to 24 hours after exposure. One interpretation of these results is that those effects of EMS and MMS exposure which result in S-phase delay and G2 arrest may be those elements common to the induction of SCE and cellular toxicity.

Sister chromatid exchange frequency, cellular replication and relative cloning efficiency in human teratocarcinoma-derived cells.

To determine the relationships between the induction of specific biological responses and exposure to DNA-damaging agents, human teratocarcinoma-derived cells were exposed to either ethyl methanesulfonate or to methyl methanesulfonate, and sister chromatid exchange, cellular proliferation and relative cloning ability measured. SCE increased while cellular proliferation and relative cloning ability each decreased in a concentration-dependent manner. Methyl methanesulfonate was consistently more efficient in inducing biological responses than was ethyl methanesulfonate. When the individual responses were compared, the decrease in cellular proliferation paralleled the reduction in cloning efficiency. A strong correlation was also observed between the reduction in relative cloning ability and sister chromatid exchange frequency. Because these relationships are similar to those previously described in other mammalian cell lines, the observations in our study suggest that the P3 cell line is an appropriate choice for modeling effects of toxicant exposure in human cells.

Retinoic acid fails to induce differentiation in human teratocarcinoma cell lines that express high levels of a cellular receptor protein

Human embryonal carcinoma (EC) cells and other human teratocarcinoma-derived cells lines were not observed to differentiate in culture in response to retinoic acid. Nevertheless, they express high levels of a cellular binding protein (cRABP) which, in murine EC cells, appears to be necessary for the induction of differentiation by retinoic acid.

Partial structural analysis of concanavalin A binding glycopeptides of large size from human teratocarcinoma-derived cells: cleavage by hydrazinolysis and alkaline borohydride.

Pronase digests of cultured teratocarcinoma-derived cells (PA 1) of human origin have been previously shown to contain large-sized glycopeptides (relative mass (Mr) greater then 7400), of which 15-23% are retained by columns of concanavalin A (Con A) - Sepharose and can be eluted with 10 mM methyl alpha-D-mannopyranoside. The present data show that this fraction (A - Con A II) contains a family of glycopeptides that are degradable with anhydrous hydrazine as well as with 0.05 M NaOH - 1 M NaBH4. The cleavage products representing individual oligosaccharide chains, presumably as oligosaccharides and glycopeptides, consisted mostly of medium- (Mr 1400-6000) and small-sized (Mr less than 1400) molecules. This implies that glycopeptides bearing several oligosaccharide chains were present in A - Con A II. Most of the individual oligosaccharide chains were not bound to Con A - Sepharose, but some were retained by the lectin column in the same way as the original glycopeptides. Some of the oligosaccharides were degraded partially with endo-beta-galactosidase from Escherichia freundii suggesting the presence of GalbetaGlcNAcbeta repeats. The present findings show that A - Con A II may be different from the "embryonic" glycopeptides of mouse teratocarcinoma cells that are reportedly not cleaved by mild alkaline borohydride treatment. Instead, A - Con A II is reminiscent of the T-1 glycopeptide of glycophorin.

Identification of hyaluronic acid as a component of human teratocarcinoma-derived cells of line PA 1.

Ovarial teratocarcinoma-derived cells (PA 1) of human origin, which are known to be capable of differentiating in vitro, incorporate label from radioactive galactose and 2-amino-2-deoxy-D-glucose into hyaluronic acid. This macromolecule, which is prominent in many differentiating embryonic and malignant cells, was separated from sulfated glycosaminoglycans and from large-sized glycopeptides by chromatography on concanavalin A – Sepharose. It was identified by a number of chemical and enzymatic degradations as well as by several chromatographic methods.

Cell-associated glycosaminoglycans of human teratocarcinoma-derived cells of line PA 1.

Human teratocarcinoma-derived cells of line PA 1, which are capable of differentiating in vitro [Zeuthen, J. et al. (1980) Int J. Cancer, 25, 19-32], incorporate label from radioactive sulfate and/or glucosamine into several large-sized glycosaminoglycans including hyaluronate, chondroitin sulfate/dermatan sulfate co-polymers, heparan sulfate and keratan-sulfate-like molecules. All these polysaccharide fractions were identified by specific degradation methods. The labeled hyaluronate was degraded into a mixture of unsaturated octa-, hexa- and tetra-saccharides by a treatment with Streptomyces hyaluronidase (EC 4.2.2.1). The chondroitin sulfate/dermatan sulfate co-polymers were cleaved with chondroitin AC lyase (EC 4.2.2.5) into unsaturated disaccharides and a series of unsaturated oligosaccharides; the latter were degraded by a treatment with chondroitin ABC lyase (EC 4.2.2.4) into unsaturated disaccharides. Heparan sulfate was degraded with nitrous acid into free inorganic [35S]sulfate and a series of [35S]sulfate-labeled oligosaccharides and/or glycopeptides. The keratan-sulfate-like molecules were hydrolyzed by a treatment with endo-beta-galactosidase from Escherichia freundii into a series of distinct [35S]sulfate-labeled oligosaccharides; small oligosaccharides were liberated also from [3H]galactose-labeled molecules. The smallest one of the liberated oligosaccharides was tentatively identified as a sulfated disaccharide.

Fractionation of large glycopeptides of human teratocarcinoma-derived cells by concanavalin A-Sepharose chromatography.

Human teratocarcinoma derived cells, line PA 1, were labeled with radioactive monosaccharides and subsequently digested with pronase. Large sized glycopeptides (fraction A) were isolated by gel filtration on Bio-Gel P-10. Their chromatography on concanavalin A-Sepharose gave three subfractions, two of which were eluted with a sugar-free buffer and the third with 10 mM alpha-methyl mannoside. The first subfraction (fraction A-Con A Ia) incorporated label from [3H]galactose and [3H]glucosamine and contained the largest components of fraction A. The second and the third subfractions (fractions A-Con A Ib and A-Con A II) were glycopeptides which incorporated label from tritiated fucose, mannose, galactose, and glucosamine. Even these molecules were of large size eluting partially at the void volume from Bio-Gel P-60. The glycopeptides of fraction A-Con A Ib contained mannose, fucose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Fucose and galactose residues occupied ultimate or penultimate positions at the nonreducing termini of the oligosaccharides. N-Acetylneuraminic acid, too, was present in the glycopeptides of fraction A.