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Title: A study of polyamine oxidase activity in mammalian cells : a role in the regulation of cell growth
Author: MacGowan, Sian H.
Awarding Body: University of Aberdeen
Current Institution: University of Aberdeen
Date of Award: 1986
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We were unable to successfully measure polyamine oxidase activity in vitro in BHK-21/C13 cells, despite modifications to the published method, including the separation of products from unreacted substrate on ion-exchange chromatography columns. We established a sensitive, reproducible HPLC method for the measurement of free polyamines and their acetyl derivatives in cell extracts and body fluids. Optimum conditions for assay of the enzyme from freshly isolated hepatocytes were established, together with some kinetic parameters for the PAO in hogenates of FIH. Polyamine anti-metabolites such as MGBG and aminoguanidine were found to be ineffective on PAO activity in vitro. PAO activity in primary cultures of hepatocytes varied with time in culture, being high initially, then falling up to 24h, followed by a rise and plateau by 48h. Sodium butyrate was a potent, but reversible inhibitor of cell division and uptake of [3H]-thymidine in BHK-cells and PyY-cells. Protein synthesis was not effected. It caused increased excretion of polyamines from both growing and confluent cells, probably as a result of altered membrane permeability. MGBG also inhibited cell division and altered efflux of polyamines from the cells. It was, however, difficult to implicate polyamine oxidase activity in the alteration of intracellular polyamine levels under these conditions. Double-labelling of the cells with [3H]- and [14C]-putrescine showed that confluent and MGBG-treated cells, particularly the latter, exhibited some degradative activity, possibly that of polyamine oxidase. We suggest that PAO activity is highest in growth-inhibited cells and that the enzyme probably acts upon cytosolic polyamines and not those in other compartments of the cell, such as the nucleus.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Biochemistry Biochemistry Molecular biology Cytology Genetics