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Characterization of membrane permeability alterations induced in Vero cells by Clostridium perfringens enterotoxin
Abstract:
Alterations in plasma membrane permeability induced by Clostridium perfringens enterotoxin were studied using Vero (African green monkey kidney) cells which were radioactively labeled with four markers of different molecular size. The markers were alpha-amino[14C]isobutyric acid (Mr 103), 3H-labeled nucleotide (Mr approx. 300), 51Cr label (Mr approx. 3000) and [3H]RNA (Mr>25000). Over a 2h period, enterotoxin caused significant release of aminoisobutyric acid, nucleotides and 51Cr label but not RNA. The effects of enterotoxin on label release were dose- and time-dependent. The rate of release of markers was dependent upon their size. Permeability alterations could be detected within 15 min with a high dose of enterotoxin. Gel chromatography of released material was used to determine that markers of Mr 3000 but not 25000 leaked from permeabilized cells. It was concluded that enterotoxin is producing functional 'holes' of limited size in the membrane. Permeability changes due to enterotoxin treatment differed between confluent and nonconfluent (growing) cells. We propose that the primary action of the enterotoxin is to interact with the plasma membrane and produce functional 'holes' of defined size. The resultant alterations in membrane permeability cause the loss of essential cellular substances which inhibits processes such as macromolecular synthesis and eventually leads to cell deterioration and death.
Insights
Clostridium perfringens enterotoxin creates functional holes in cell membranes, allowing small molecules to leak out. This toxin-induced membrane permeability alters cell function and leads to cell death.
Area of Science:
- Cell Biology
- Molecular Biology
- Toxicology
Background:
- Clostridium perfringens enterotoxin (CPE) is a potent toxin.
- CPE disrupts cellular integrity.
- Understanding CPE's mechanism is crucial for cell biology and medicine.
Purpose of the Study:
- To investigate the effects of CPE on plasma membrane permeability.
- To determine the size selectivity of CPE-induced membrane pores.
- To elucidate the mechanism of CPE-mediated cell damage.
Main Methods:
- Vero cells were radioactively labeled with markers of varying molecular sizes.
- Cells were treated with CPE, and the release of labeled markers was measured over time.
- Gel chromatography was used to analyze the size of molecules that leaked from cells.
Main Results:
- CPE caused dose- and time-dependent release of small molecular weight markers (amino acids, nucleotides, ~3000 Da Cr label).
- Larger molecules like RNA (>25000 Da) were not released, indicating size-selective pore formation.
- Permeability changes were observed within 15 minutes of high-dose CPE exposure.
- Differences in permeability changes were noted between confluent and nonconfluent cells.
Conclusions:
- CPE induces the formation of functional pores of limited size in the plasma membrane.
- These pores allow the passage of molecules up to approximately 3000 Da.
- CPE-mediated loss of cellular substances inhibits vital processes, leading to cell death.