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Mitogenic activity of cytoplasmic membranes isolated from L-forms of Staphylococcus aureus
Abstract:
Cytoplasmic membranes of L-forms of Staphylococcus aureus exerted a strong mitogenic effect on splenocytes of athymic nude mice as well as normal mice, while a cytoplasmic fraction of the same bacteria did not show definite mitogenicity. The mitogenic principle(s) of the membrane fraction was resistant to treatment with trypsin and was heat stable (at 100 C for 10 min). The active principle(s) in the insoluble residue of the membrane fraction digested with trypsin was not extracted with cold acetone, but could be solubilized by extraction with a cold chloroform-methanol mixture (2:1, v/v). The mitogenic principle(s) in the extract was fractionated by silicic acid column chromatography. Among five fractions separated by chromatography, fractions eluted with chloroform-methanol mixtures (1:1 and 1:20, v/v) were found to be strongly mitogenic. The cytoplasmic membranes of the L-forms also exerted a definite mitogenic effect on guinea pig splenocytes, but not on the thymocytes.
Insights
Staphylococcus aureus L-form membranes strongly stimulate mouse splenocytes, unlike cytoplasmic fractions. This bacterial membrane mitogen is heat-stable and extracted by chloroform-methanol, indicating a lipid-based immune stimulant.
Area of Science:
- Immunology
- Microbiology
- Biochemistry
Background:
- Bacterial components can modulate immune responses.
- L-forms of bacteria, lacking cell walls, may possess unique immunomodulatory properties.
Purpose of the Study:
- To investigate the mitogenic potential of Staphylococcus aureus L-form cytoplasmic membranes.
- To characterize the nature of the mitogenic principle(s) present in these membranes.
Main Methods:
- Preparation of cytoplasmic membrane and cytoplasmic fractions from Staphylococcus aureus L-forms.
- Assessment of mitogenic activity on mouse splenocytes (athymic and normal) and guinea pig splenocytes/thymocytes.
- Biochemical treatments including heat, trypsin digestion, and solvent extraction (acetone, chloroform-methanol).
- Fractionation using silicic acid column chromatography.
Main Results:
- Cytoplasmic membranes, but not cytoplasmic fractions, of Staphylococcus aureus L-forms exhibited significant mitogenic activity on mouse splenocytes.
- The mitogenic activity was resistant to heat (100°C) and trypsin digestion.
- The active principle was extracted by a chloroform-methanol mixture and further purified by column chromatography.
- Fractions eluted with specific chloroform-methanol ratios demonstrated potent mitogenicity, suggesting a lipid-based molecule.
Conclusions:
- Staphylococcus aureus L-form cytoplasmic membranes contain potent, heat-stable, and trypsin-resistant mitogenic factors.
- These factors are likely lipid in nature, based on extraction and chromatographic behavior.
- Bacterial membranes, particularly from L-forms, represent a significant source of immunomodulatory molecules.