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Rat mast cells permeabilised with streptolysin O secrete histamine in response to Ca2+ at concentrations buffered in
Abstract:
Rat peritoneal mast cells have been permeabilised by treatment with streptolysin O which generates membrane lesions of macromolecular dimensions. In the presence of Ca2+ buffered at concentrations in the micromolar range, the permeabilised mast cells release histamine, beta-N-acetylglucosaminidase and lactate dehydrogenase. Release of the two secretory components (but not lactate dehydrogenase) has an obligatory requirement for a nucleoside triphosphate and micromolar concentrations of Ca2+. Inosine triphosphate (ITP) supports the release reaction better than ATP does. It is concluded that the secretory materials are released from the cells by an exocytotic mechanism, while lactate dehydrogenase leaks from the cells through the toxin-generated lesions. By initially withholding and then supplying Ca2+ to the permeabilised cells, it is shown that the exocytotic secretory reaction can persist even when the cytosol is depleted of the bulk of soluble proteins. The streptolysin O treated mast cell preparation represents a simplified system with which to study the mechanism of exocytosis.
Insights
This study shows that rat mast cells release histamine and beta-N-acetylglucosaminidase via exocytosis, requiring calcium and nucleoside triphosphates. Lactate dehydrogenase, however, leaks through toxin-induced membrane pores.
Area of Science:
- Cell Biology
- Immunology
- Biochemistry
Background:
- Rat peritoneal mast cells are crucial immune cells involved in allergic responses.
- Mast cell degranulation releases inflammatory mediators.
- Understanding the exocytosis mechanism is vital for therapeutic development.
Purpose of the Study:
- To investigate the mechanism of exocytosis in permeabilized rat mast cells.
- To determine the role of calcium and nucleoside triphosphates in mediator release.
- To differentiate between exocytotic release and non-specific leakage.
Main Methods:
- Mast cells were permeabilized using streptolysin O, creating membrane lesions.
- Mediator release was measured in the presence of varying calcium and nucleoside triphosphate concentrations.
- Lactate dehydrogenase release was monitored as a marker for non-specific leakage.
Main Results:
- Permeabilized mast cells released histamine and beta-N-acetylglucosaminidase in a calcium-dependent manner.
- Nucleoside triphosphates, particularly inosine triphosphate (ITP), were essential for this secretory release.
- Lactate dehydrogenase was released independently of these factors, indicating leakage through pores.
- Exocytosis persisted even after depletion of soluble proteins, demonstrating its robustness.
Conclusions:
- Secretory component release occurs via an exocytotic mechanism requiring calcium and nucleoside triphosphates.
- Lactate dehydrogenase release is due to non-specific leakage through streptolysin O-induced pores.
- The streptolysin O-treated mast cell system provides a simplified model for studying exocytosis.