published_or_final_version / abstract / Pathology / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Biological Sciences / Doctoral / Doctor of Philosophy

Graduation date: 1997

Primary cultures of stromal-vascular (S-V) cells from adipose tissue were used to investigate the regulation of preadipocyte development. Differentiation of S-V cells was found to be under hormonal control. Insulin and glucocorticoids are essential for S-V cell differentiation in culture. S-V cells from both newborn and mature pig adipose tissue and sera from both ages were used to examine the effect of age on preadipocyte development. S-V cells from newborn pigs replicated faster and appeared more responsive to serum borne factors influencing S-V cell growth and development in culture. Serum source (newborn vs mature) did not affect differentiation of S-V cells from newborn or mature pig adipose tissue. When sera from fed or fasted pigs were used to culture newborn pig S-V cells, fasted pig sera stimulated greater differentiation and decreased cell replication as indicated by DNA content of rat S-V cell culture. Lean pig serum compared to obese pig serum, increased differentiation activity in culture of S-V cells an effect which may be influenced by sex. When sera from rat and pig were subjected to gel filtration fractionation on Sephacryl S-200 column, the elution profiles of both sera were similar. Rat serum contained six additional peaks (280 nm) not present in pig serum. Rat serum fraction two (apparent molecular size 67-150 kD) promoted greater differentiation of S-V cells than other rat serum fractions or pig serum fraction two. Fraction three (apparent molecular size 17-43 kD) of both sera inhibited differentiation and lipid filling in cultures of S-V cells but only rat fraction three promoted cell proliferation. Rat and pig S-V cells have different morphology when differentiated. Differentiated rat S-V cells appeared as individual cells when cultured in serum free or serum supplemented medium while differentiated pig S-V cells appeared as individual cells in serum free medium and as a tight cluster of cells in serum supplemented medium. Both cells responded differently to sera obtained from pigs of differing ages and development of rat S-V cells was influenced by anatomic site. / Graduation date: 1992

Thesis (Ph. D.)--Rutgers University, 2009. / "Graduate Program in Chemistry." Includes bibliographical references (p. 131-144).

Includes bibliographical references (p. 85-99).

Thesis (Ph. D.)--University of Oregon, 2000. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 106-112). Also available for download via the World Wide Web; free to University of Oregon users.

Thesis (Ph. D.)--Hong Kong University of Science and Technology, 2003. / Includes bibliographical references (leaves 139-153). Also available in electronic version. Access restricted to campus users.

Deoxyribonucleic acid (DNA) replication in eukaryotic cells requires a highly complex series of protein-DNA interactions. Elucidation of the mechanisms by which DNA replication occurs is vital to the understanding of cellular growth. DNA polymerase alpha is an enzyme with a putative role in the replication of eukaryotic DNA. Modification by phosphorylation and dephosphorylation is one process by which enzymatic activity is regulated. The purpose of this research was to determine if a phosphorylation event could be of significance in the expression of DNA polymerase alpha activity. Evidence will be presented for the regulation of DNA polymerase alpha by phosphorylation. A highly purified DNA polymerase alpha fraction was prepared from Chinese hamster ovary cells. Purification procedure included ion-exchange chromatographies and affinity chromatography. Both the crude DNA polymerase alpha activity and the highly purified DNA polymerase alpha activity were stimulated six-fold by the addition of exogenous bovine cardiac muscle cyclic AMP-dependent protein kinase. Dephosphorylation of the highly purified DNA polymerase alpha fraction by alkaline phosphatase resulted in a concomitant decrease in DNA polymerase alpha activity. An endogenous protein kinase activity was detected in the highly purified DNA polymerase alpha fraction. Incubation of this fraction in a protein kinase reaction mixture including adenosine triphosphate (ATP) could stimulate DNA polymerase alpha activity to twelve-fold that observed in controls with no pre-incubation. The endogenous protein kinase activity in the highly purified DNA polymerase alpha fraction was utilized to indicate (1) the linear increase in DNA polymerase alpha activity with time of phosphorylation which was dependent on the presence of ATP, (2) the linear relationship between γ³²P-ATP incorporation and DNA polymerase alpha activity, and (3) the incorporation of labelled phosphate into DNA polymerase alpha as determined by SDS-PAGE analysis. Finally, the co-purification of the endogenous protein kinase with DNA polymerase alpha is presented. The significance of this research in relation to the heterogeneous nature reported for DNA polymerase alpha is discussed. It is speculated that the phosphorylational regulation of DNA polymerase alpha may play a role in the transformation of cells.

Thesis (Ph. D.)--University of Hawaii at Manoa, 1996. / Includes bibliographical references (leaves 103-117). / Microfiche. / xiv, 117 leaves, bound ill. 29 cm