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C F Graham

Publications and source records attributed to C F Graham.

At least 37 records · Page 2Linked to original sources

Clonal analysis of the change in growth phenotype during embryonal carcinoma cell differentiation.

Retinoic acid has been shown to induce the differentiation of mouse embryonal carcinoma cells. Previous workers have reported that bulk cultures of the differentiated derivatives have a slower growth rate and a reduced capacity to form tumours. We have analysed this change in growth rate for a sub-tetraploid EC cell line, PC13 clone 5 MA2, at a clonal level and have shown that the production of cells with a slower growth rate is not a result of cell selection. We have also demonstrated that the action of retinoic acid on growth rate is delayed for approximately 48 h and that the new growth phenotype, once attained, is stable. Finally we have confirmed at a clonal level that the differentiated derivatives of EC cells exposed to retinoic acid have a reduced capacity to form tumours. Clones of EC cells exposed to retinoic acid for longer than 96 h are unable to form tumours in a 30-day period, whilst 87% of their untreated counterparts are able to do so.

Animals

Epidermal growth factor receptors increase during the differentiation of embryonal carcinoma cells.

Mouse teratocarcinoma stem cells (embryonal carcinoma, or EC cells) bind very small amounts of mouse epidermal growth factor (EGF) and the latter hormone seems to have no stimulatory effect on the growth of two cloned lines of EC cells. However, when EC cells are induced to differentiate into large flat endodern-like cells (END cells), EGF receptors increase in number reaching a plateau in 6 to 8 days. At 8 to 10 days after induction, END cells multiply very slowly, but when EGF is added (3 x 10(-10) M) to the medium, cell division is stimulated and a further change in morphology occurs. This letter describes the binding characteristics and numbers of the EGF receptors on EC and END cells and shows that exogenous retinoic acid increases the numbers of EGF receptors on END cells. We were unable to find endogenous competing factors produced by EC cells. Such factors could account for the lack of detectable binding of EGF on these cells. As EC cells differentiate to END cells, so the ability of the cells to form tumours is reduced. Since this change is accompanied by an increase in the number of EGF receptors there may be a relationship between these two events.

Animals

Formation and consequences of cell patterns in preimplantation mouse development.

The behaviour of groups of cells was studied in culture during preimplantation mouse development. The following observations were made with intact embryos minus the zona pellucida and with embryos whose cells had been dissociated and recombined. The form of the 4-cell embryo was related to the behaviour of the first cell to divide to this stage. The form of the 8-cell embryo depended on contact between the groups of four cells each derived from a single cell at the 2-cell stage. The form of the 16-cell embryo depended on cell movement during and after division from the 8- to the 16-cell stage. These results suggest that the morphogenetic movements of these early embryonic cells are principally governed by continuous cell interactions after fertilization. The cell surfaces of the embryos were examined with scanning electron microscopy in an attempt to discriminate between the mechanisms which could account for these movements.

Animals

Growth and differentiation of an embryonal carcinoma cell line (C145b).

Several cell and tumour lines were isolated from a single-embryo-derived teratocarcinoma and their karyotypes and differentiation in adult hosts recorded. The majority of cells contained normal karyotypes by banding. The cells were injected into blastocysts and although they sometimes colonized the yolk sac, they never colonized the embryo. Thus the possession of a normal karyotype is not a sufficient condition for embryo colonization. The loss of growth capacity was investigated by studying differentiation and tumourigenicity in a variety of circumstances. The change in appearance from an EC cell morphology to a big flat cell in culture leads to retardation of growth in adult hosts. When EC cells are injected into a blastocyst, the ability to grow progressively both in culture and in adult hosts is lost.

Animals

Appearance of interferon inducibility and sensitivity during differentiation of murine teratocarcinoma cells in vitro.

Pluripotential embryonal carcinoma (EC) cells do not produce interferon after treatment with a wide variety of inducers, nor are they sensitive to its action. Several differentiated lines derived from the EC cells, however, both produce and are sensitive to mouse interferon. Differentiation of EC cells in vitro is accompanied by development of interferon inducibility and sensitivity.

Cell Differentiation

Cell division and cell allocation in early mouse development.

Cell division was observed in intact and dissociated mouse embryos between the 2-cell stage and the blastocyst in embryos developing in culture. Division to the 4-cell stage was usually asynchronous. The first cell to divide to the 4-cell stage produced descendants which tended to divide ahead of those cells produced by its slow partner at all subsequent stages of development up to the blastocyte stage. The descendants of the first cell to divide to the 4-cell stage did not subsequently have short cell cycles. The first cell or last cell to divide from the 4-cell stage was labelled with tritiated thymidine. The embryo was reassembled, and it was found that the first pair of cells to reach the 8-cell stage contributed disproportionately more descendants to the ICM when compared with the last cell to divide to the 8-cell stage.

Animals

Features of cell lineage in preimplantation mouse development.

The cell lineage of the mouse was studied from the 2-cell stage to the blastocyst. Lineage to the 8-cell stage was followed under the microscope. Each cell from the 2-cell stage divided to form two daughter cells which remained attached. Subsequently, these two daughters each produced two descendants; one of these descendants regularly lay deep in the structure of the embryo while the other was peripheral. Lineage to the blastocyst was followed by injecting oil drops into cells at the 8-cell stage, and then following the segregation of these drops into the inner cell mass and trophectoderm. Between the 8-cell stage and the blastocyst, the deep cells contributed more frequently to the inner cell mass than did the peripheral cells.

Animals

Analysis of the forms of acetylcholinesterase from adult mouse brain.

The solubilization of 80% of the acetylcholinesterase activity of mouse brain was performed by repeated 2h incubations of homogenates at 37 degrees C in an aqueous medium. Analysis of the soluble extract by gel filtration on Sephadex G-200 showed that up to 80% of the enzyme activity was eluted in a peak which was estimated to consist of molecules of about 74000mol.wt. This peak was called the monomer form of the enzyme. After 3 days at 4 degrees C, the soluble extract was re-analysed and was eluted from the column in four peaks of about 74000, 155000, 360000 and 720000 mol.wt. Since the total activity of the enzyme in these peaks was the same as that in the predominantly monomer elution profile of fresh enzyme, we concluded that the monomer had aggregated, possibly into dimers, tetramers and octomers. Extracts of the enzyme were analysed by polyacrylamide-gel electrophoresis and the resulting multiple bands of enzyme activity on gels were shown to separate according to their molecular sizes, that is by molecular sieving. All these forms had similar susceptibilities to the inhibitors eserine, tetra-isopropyl pyrophosphoramide and compound BW 284c51 [1,5-bis-(4-allyldimethylammoniumphenyl)pentan-3-one dibromide]. Thus the forms of the enzyme in mouse brain which can be detected by gel filtration and polyacrylamide-gel electrophoresis may all be related to a single low-molecular-weight form which aggregates during storage. This supports similar suggestions made for the enzyme in other locations.

Acetylcholinesterase

Development of parthenogenetic and fertilized mouse embryos in the uterus and in extra-uterine sites.

Mouse eggs were activated with hyaluronidase in vitro and subsequently transferred to the oviduct. In the female reproductive tract they formed morulae and blastocysts which died soon after implantation. Haploid blastocysts were transferred beneath the kidney capsule and here some formed disorganized egg-cylinder structures in a week. Morulae and blastocysts from haploid and diploid parthenogenones were also transferred beneath the testis capsule. Two to four months later the growths which had formed were sectioned. They contained neural tissue, pigment, keratinized epithelium, glandular epithelium, ciliated epithelium, cartilage, bone, muscle, adipose tissue, and haemopoietic tissue. The range of cell types was similar to that produced by fertilized control blastocysts except that the parthenogenones did not form identifiable yolk-sac carcinoma or embryonal carcinoma cells. The growths from haploid and diploid parthenogenones in the testis were stained with Feulgen and their DNA content measured. Growths from diploid embryos contained the normal diploid amount of DNA while growths from haploid embryos contained less than this amount. Cell cultures were prepared from the growths. The cells which were investigated contained no Y chromosome, suggesting that they were derived from the embryonic cells rather than the cells of the male host. These cells contained a near diploid chromosome number, although some of them were originally derived from haploid embryos.

Animals