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Date of Award
1993
Document Type
Dissertation - Pacific Access Restricted
Degree Name
Doctor of Philosophy (Ph.D.)
Department
Pharmacy
First Advisor
James Blankenship
First Committee Member
David S. Fries
Second Committee Member
Timothy Smith
Third Committee Member
Paul A. Richmond
Fourth Committee Member
Denis Meerdink
Abstract
Polyamines are widely distributed in living organisms. Although the complexities of the growth and differentiation of normal and malignant cells are not fully understood, it is known that polyamines are involved in these processes.
Induction of neurite outgrowths is a well documented effect of nerve growth factor (NGF) on PC12 cells. NGF at 50 ng/ml induced neurite outgrowths in about 90% of the cells after 8 days of treatment. In addition to morphological differentiation, we observed a significant (p (is less than) 0.01) decrease in the rate of growth of the cells.
Inhibition of the rate-limiting enzyme of polyamine synthesis, ornithine decarboxylase, by DFMO significantly (p (is less than) 0.01) inhibited the growth of PC12 cells and slightly inhibited morphological differentiation induced by NGF.
Previous studies have shown that increased levels of NBacetylspermidine lead to stimulation of cell growth. In the present study, an inhibitor of N8-acetylspermidine deacetylase, 7-[N-(3-aminopropyl)amino]heptan-2-one (APAH), was used to study the effects of N8-acetylspermidine on growth and differentiation of PC12 cells.
APAH was found to stimulate the growth rate and induce morphological differentiation of these cells. Maximum cell densities were increased (p (is less than) 0.01) by 17.01%, 12.32% and 24.05% compared to control at concentrations of 10-5 M, 10-6M, and 10-7M APAH, respectively. Maximum cell differentiation was observed about 8 days after initiation of treatment. At concentrations of 10-5 M, 10-6M, and 10-7M APAH, 30.37%, 26.5%, and 34% of the cells were differentiated, respectively.
Coaddition of APAH and NGF to the medium inhibited the cell growth and this inhibition was about the same as the inhibition induced by NGF alone. N8-acetylspermidine increased cell growth at concentrations of 10-5 M, 10-6M, and 10-7M. However, a significant increase was observed only from day 4 to day 7 of treatment. At the same concentration, maximum morphological differentiation was observed on day 7 with 15.5%, 14.25% and 12.05% of the cells differentiated by 10-5 M, 10-6M, and 10-7M APAH, respectively.
When the cells received simultaneous treatment with APAH and N8-acetylspermidine, a small synergistic effect on the stimulation of cell growth and morphological differentiation was observed.
Genistein, an inhibitor of protein tyrosine kinase, is known to inhibit cell growth and induce differentiation of some cell lines. At a concentration of 40 μg/ml, inhibition of PC12 cell growth (p (is less than) 0.01) was observed but the morphological differentiation of the cells was not induced. The combination of genistein and NGF led to inhibition of cell growth not significantly different from the inhibition induced by NGF alone. When added in the presence of APAH, genistein showed inhibition of cell growth similar to the growth observed by genistein alone and morphological differentiation was about the same as the differentiation induced by APAH.
Our study is the first report of an enzyme inhibitor of polyamine metabolism inducing differentiation in PC12 cells. It focuses on the possible role for nuclear N8-acetylation of spermidine in induction of neurite outgrowths in these cells and in processes regulating differentiation in general.
Pages
98
Recommended Citation
Menezes, Ana Maria Soares. (1993). Effects of N(8)-acetylspermidine deacetylase inhibition on growth and differentiation of PC12 cells. University of the Pacific, Dissertation - Pacific Access Restricted. https://scholarlycommons.pacific.edu/uop_etds/2957
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