by Tai-Tung Yip. / Bibliography: leaves 191-199 / Thesis (Ph.D.)--Chinese University of Hong Kong, 1985

A cloned metastatic variant of a murine tumor cell line, Lewis lung carcinoma (LLC(C3), was synchronized by double exposure to thymidine (5 x 10-4 M) at G1/S boundary. The effects of Prostaglandin (PG)E1, PGE2, PG synthetase inhibitors, and an analog of cAMP (dbut-cANP) on the life cycle of synchronized LLC(C3) cells were examined. When added to cells in G1 phase, PGE1 and dbut-cAMP enhanced 3H-thymidine uptake during S phase. Both of these agents suppressed S phase when they were added to cells entering S phase. The addition of PGE2 to LLC(C3) cells in G1 phase had no effect on S phase of the synchronized LLC(C3) cells. Prostaglandin E2 suppressed S phase when it was added at the beginning of S phase. A low concentration of PGE2 was more suppressive to S phase than was a high concentration. Indomethacin, an irreversible PG synthetase inhibitor, and piroxicam, a reversible PG synthetase inhibitor, suppressed S phase of synchronized LLC(C3) cells. These results suggest that prostaglandins can regulate the cell cycle of LLC(C3) cells through modulation of cAMP levels.

Thesis (M.S.)--University of Hawaii at Manoa, 2006. / Includes bibliographical references (leaves 83-86). / x, 86 leaves, bound ill. 29 cm

Bibliography: leaves 141-169. / xix, 170, [63] leaves, [49] leaves of plates : ill. (chiefly col.) ; 30 cm. / Title page, contents and abstract only. The complete thesis in print form is available from the University Library. / Calcitonin (CT) is a 32 amino acid peptide hormone, known to inhibit osteoclastic bone resorption by direct interaction with cell surface calcitonin receptors (CTR). This study aims to define the intracellular mechanisms by which CT treatment results in decreased cellular proliferation. Results show, for the first time, that the growth regulating actions of CT are receptor isoform specific. In addition, CT inhibition of cellular proliferation occurs by arresting cells in the G2 phase of the cell cycle via a p21 mediated mechanism, which is at least partially activated through the Erk1/2 Map Kinase pathway. / Thesis (Ph.D.)--University of Adelaide, Dept. of Orthopaedics and Trauma, 2000

No description available.

A new approach has been introduced to characterize the epidermal growth factor receptors and their relation to the mechanism of cell growth control using hybrid cells made between human EGF responsive cells and mouse A9 cells incapable of EGF binding. BALBc mice were immunized with human carcinoma A431 cells carrying an extraordinary high number of EGF receptors; antisera were used to identify the human nature of EGF receptors in these hybrid cells. One of the hybrid lines, C2B5, that retains only one human chromosome, an X/7 translocation, and a nearly complete mouse parental genome was used to analyze the relationship of the binding ability and certain post-receptor functions to the cellular mitogenic response. It was shown that the ability to bind, internalize and degrade the ligand and/or its receptor is not sufficient for cells to respond to the mitogen. Spleen cells from mice immunized with A431 cells were fused with mouse myeloma P3NP cells. One of the isolated hybridoma lines, B4G7, secreted a monoclonal antibody of the IgG class which inhibits the binding of ¹²⁵I-EGF to A431 and human fibroblasts, but not of mouse 3T3 cells. This inhibition was partial (65-70%) and Scatchard analysis of the binding data suggested that antibody preferentially interacts with a low affinity class of EGf receptors. The antibody specifically precipitated EGF receptor from radiolabeled cells. This monoclonal antibody was crosslinked to subunit A of toxic ricin through a disulfide bond. The resulting conjugate inhibited protein synthesis of A431 cells at 4 x 10⁻¹¹M and exhibited substantial cell killing. Using this conjugate we isolated a variant of A431 cells, designated C1-B7, with approximately 30 times less EGF binding capacity. Contrary to the parental A431, this variant is resistant to EGF-induced suppression of cell growth and appears to have lost most of the low affinity receptors. The high affinity type EGF receptors retained by the variant are 170,000 Mr and susceptible to EGF-induced phosphorylation, presumably on tyrosine residues. In membrane prepared from this variant, besides the EGF receptor, a low molecular weight component of as yet unknown nature is highly phosphorylated in an EGF-independent manner.

Ras proteins are a special class of proteins that mediate cell growth signals. Their importance lies in the fact that they are products of a proto-oncogene. This means that under certain conditions the gene that determines its structure is altered and a mutant protein results that is involved in the transformation of normal cells to cancer cells. The actual function by which the protein acts in the signal pathway is not known. However it is known that they act as a switch, undergoing a cycle involving the exchange of guaninosine nucleotides in the binding site. This thesis uses computer simulations to study the energetics of this binding, with the long term aim of developing a drug to inhibit the transforming activity of the oncogenic protein. To begin with, a model of the protein based on a crystal structure is built. Using Molecular dynamics the motion of this model is studied. A possible mechanism by which one half of the nucleotide cycle could be induced is investigated, with the result that phosphorylation of the protein may be involved. The main part of the thesis is then devoted to using the free energy perturbation (FEP) method to calculate the difference in Gibbs binding free energy between the nucleotides in the protein. Using histamine as a model, a method of dealing with charged, flexible molecules is developed; namely the inclusion of a reaction field and comprehensive conformational analysis. The results from the associated calculations are seen to be very close to experimental data. The same procedures are then applied to the much more complex ras: nucleotide system with less successful results, the reason for which is mostly due to the restriction of limited computer resources to tackle such a problem. The conclusion is that given the resources and by using the techniques developed in this thesis, this type of calculation is a feasible way to study such systems.

Call number: LD2668 .T4 1984 M44 / Master of Science

Cellular transformation is accomplished in vitro through the concerted action of growth factors and oncogenes. This association has demonstrated that malignant growth results from aberrations in pathways that normally operate to control proliferation. Activation of genes that code for growth factors, their receptors, and/or molecules essential in the transduction of signals from the cell surface to the nucleus are all potential mechanisms by which tumor cells could establish a selective growth advantage over normal cells. This dissertation addresses the question of what oncogenic mechanisms are important in the development and progression of human melanoma. These studies show that melanoma growth is regulated by endogenous substances produced by the melanoma cells themselves (autocrine stimulation), as well as by exogenous substances supplied by neighboring cells and platelets (paracrine stimulation). These factors work to drive the expression of the transformed phenotype for melanoma as evidenced by induction of serum-free soft agar growth. Human platelets were found to the the richest source of paracrine growth promoters. The factor from human platelets was characterized and partially purified. Melanoma cells respond to this 60,000 molecular weight, disulfide-bond-containing protein in colony formation assays. In addition, the protein has endothelial cell growth factor activity. Purified fractions which promoted optimal colony formation for human melanoma cells also maximally stimulated monolayer growth of bovine aortic endothelial cells, while melanocytes were nonresponsive. This implies that melanoma cells are expressing receptors for a protein which plays no known or apparent role in the normal growth of melanocytes. Melanoma cells are sensitive to growth regulatory molecules of autocrine and paracrine nature. This dissertation provides clues to the genetic lesions which have occurred in these melanoma cells to influence their proliferation. The aberrations appear to reside in those genes important in growth factor pathways at the level of endogenous production and misguided response to exogenous factors through receptor expression. We can not hope to fully inhibit the proliferation of tumor cells until we identify and understand those forces which drive their growth. These studies have increased our knowledge of those signals which stimulate melanoma cellular proliferation, and thus provide insight into important therapeutic targets.

The in vitro study of muscle cell growth is hampered by the presence of non-muscle cells, particularly fibroblasts. The heterobifunctional cross-linking agent, N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) has been used to create a novel "toxic growth factor" to address the problem. Epidermal growth factor (EGF), which stimulates fibroblast but not satellite cell proliferation, was conjugated via SPDP to a potent ribosome inhibitor, pokeweed antiviral protein (PAP). By preferentially binding to fibroblasts, it was hoped that EGF-PAP could cytotoxically eliminate fibroblasts from primary cultures of rat skeletal muscle satellite cells. While EGF-PAP did prove to be a fibroblast cytotoxin, it could not completely eliminate them from cell cultures. Low dose-time exposures improved the ratio of multinucleated cells to mononucleated cells (percent fusion) by up to 66% over controls, but increased concentrations, or durations of EGF-PAP treatment, proved detrimental to satellite cell growth and/or differentiation.