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Subcellular localization of the eosinophil chemotactic factor (ECF) and its inactivator in human polymorphonuclear
Abstract:
An eosinophil chemotactic factor (ECF) of low MW can be released from human polymorphonuclear leucocytes (PMN) on stimulation with the Ca-ionophore, arachidonic acid and during phagocytosis. After a rapid rise of ECF activity in the supernatant a steep fall of in its activity occurred at the later times of secretion suggesting a mechanism of ECF inactivation. ECF obtained at the later times of secretion represents a stable biological activity and does not decrease on further incubation. In addition, intact PMN and ECF combined do not lead to its inactivation, while incubation of homogenized PMN with ECF decreased its activity. These data suggest the presence of an inactivator for ECF within human PMN. The purpose of the study was to localize ECF and its inactivator within human PMN. After cell disruption, differential and equilibrium gradient centrifugation, subcellular components of human PMN can be obtained which reveal eosinophilotactic (ECF) or ECF-inactivating activity. ECF activity can be recovered (in a structurally bound state) from the microsomal fraction of unstimulated and stimulated PMNs, while another portion is obtainable as a soluble, low mol. wt ECF. The PMN-derived ECF inactivator can be recovered from the peroxidase positive (azurophilic) granules and has a mol. wt of 60,000 and less. We suggest that low mol. wt ECF is derived from the plasma membrane of PMN which can be inactivated by components of the azurophilic granules. The mechanism of inactivation is still unresolved.
Insights
Human polymorphonuclear leucocytes (PMN) release eosinophil chemotactic factor (ECF). ECF is inactivated by a component within PMN, localized to azurophilic granules, suggesting a novel regulatory mechanism.
Area of Science:
- Immunology
- Cell Biology
Background:
- Human polymorphonuclear leucocytes (PMN) release eosinophil chemotactic factor (ECF).
- ECF activity decreases over time, suggesting inactivation.
- Intact PMN do not inactivate ECF, but homogenized PMN do, indicating an intracellular inactivator.
Purpose of the Study:
- To localize ECF and its inactivator within human PMN.
- To understand the mechanism of ECF regulation.
Main Methods:
- Cell disruption and differential centrifugation.
- Equilibrium gradient centrifugation to isolate subcellular components.
- Assay of ECF and ECF-inactivating activity in isolated fractions.
Main Results:
- ECF activity was found in the microsomal fraction and as soluble low molecular weight ECF.
- An ECF inactivator was recovered from peroxidase-positive (azurophilic) granules.
- The inactivator has a molecular weight of 60,000 and less.
Conclusions:
- Low molecular weight ECF is likely derived from the PMN plasma membrane.
- Azurophilic granules contain an ECF inactivator.
- The precise mechanism of ECF inactivation remains to be elucidated.