Studies on the granulocyte cell surface oxidase.
thesisposted on 2023-05-26, 22:21 authored by Dewar, Catharine Lynne
This thesis examined the possible role of the microfilaments and microtubules in controlling the activity of the granulocyte plasma membrane oxidase. There have not been any detailed investigations of this topic published to date. The conflicting literature on the nucleotide specificity and location of the oxidase within the granulocyte was examined. In vitro techniques for isolating granulocytes from human peripheral blood and for quantitating the initial rate of plasma membrane oxidase activity were developed and used for these investigations. The involvement of microfilaments and microtubules in oxidase activity was studied by using pharmacological agents known to disrupt these structures in vitro (cytochalasin B, and colchicine, vinblastine and vincristine, respectively). Control experiments were also performed to ensure that microfilament and microtubule disruption by these agents provided the best explanation for the results presented herein. Correlative experiments were conducted to determine if a change in hexose monophosphate shunt activity was associated with the observed changes in plasma membrane oxidase activity brought about by the drug treatments. Experiments on non-phagocytic granulocytes showed that microfilament disruption led to enhanced oxidase activity, while microtubule disruption produced a dual effect : a paradoxical enhancement (with low doses or brief exposures to the agents) and an impairment in oxidase activity. These results suggested that microfilaments act as a constraint against, while microtubules are required for plasma membrane oxidase activity. The requirement for microtubules was highlighted in experiments where the usual enhancement in oxidase activity with cytochalasin B was ablated when the granulocytes were concomitantly incubated with a microtubule-disrupting agent. (With phagocytic granulocytes qualitatively similar results were observed. However, those experiments were not suitable for studying cell surface oxidase activity because the presence of ingestable. Particles made it impossible to determine whether plasma membrane and/or granule oxidase activity was being measured.) Based on the spectrophotometric and electron microscopic results presented and the relevant findings from other investigators, a scheme concerning the participation of the cytoskeletal elements in granulocyte plasma membrane oxidase activity was devised. In this scheme the microfilaments physically constrain the oxidase within the plasma membrane, and thereby hold the activity of the oxidase in check. Thus, microfilament disruption is envisaged to result in enhanced oxidase activity by freeing the enzyme. However, the oxidase is viewed as optimally active only as long as the microtubules are present to maintain the plasma membrane topography suitable for the enzyme's activity.
Rights statementCopyright 1979 the Author - The University is continuing to endeavour to trace the copyright owner(s) and in the meantime this item has been reproduced here in good faith. We would be pleased to hear from the copyright owner(s). Thesis (Ph.D.)--University of Tasmania, 1981. Bibliography: l. 184-202