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Isolation of variants in phagocytosis of a macrophage-like continuous cell line
Abstract:
We have isolated cloned variants in phagocytosis from a cloned continuous murine macrophage-like cell line, J 774.2. A selection procedure against Fc receptor-mediated phagocytosis was devised using IgG-coated SRBC containing a toxic drug, tubercidin, as the lethal agent. A series of variant clones deficient in Fc receptor-mediated phagocytosis were isolated. Such variants occurred at low frequency (approximately 6 X 10(-5)), were stable, and appeared to possess Fc receptors. The degree to which they were defective in phagocytosis of IgG-coated SRBC varied from clone to clone, yet all clones, were able to phagocytize latex particles. The phagocytic defect in some variants could be corrected by the addition of 8 Br-cAMP, in others, the drug was without effect. It is likely, therefore, that different variants are defective in several distinct steps critical to Fc receptor-mediated phagocytosis.
Insights
Researchers isolated macrophage variants with defects in Fc receptor-mediated phagocytosis. These stable variants, though possessing Fc receptors, showed varying phagocytic deficiencies, suggesting distinct defects in cellular processes.
Area of Science:
- Immunology
- Cell Biology
Background:
- Phagocytosis is a critical cellular process mediated by receptors like the Fc receptor.
- Understanding the molecular mechanisms of Fc receptor-mediated phagocytosis is essential for immune function.
Purpose of the Study:
- To isolate and characterize cloned variants of macrophage-like cells deficient in Fc receptor-mediated phagocytosis.
- To investigate the specific steps in Fc receptor-mediated phagocytosis that are affected in these variants.
Main Methods:
- Utilized a murine macrophage cell line (J 774.2).
- Developed a selection strategy using IgG-coated sheep red blood cells (SRBC) with a toxic drug (tubercidin) to eliminate cells with functional Fc receptor-mediated phagocytosis.
- Isolated and analyzed variant clones for phagocytic activity and Fc receptor expression.
Main Results:
- Successfully isolated stable cloned variants with reduced Fc receptor-mediated phagocytosis at a frequency of approximately 6 x 10(-5).
- These variants retained Fc receptors but exhibited varying degrees of phagocytic defect for IgG-coated SRBC.
- All isolated variants could still phagocytose latex particles, indicating a specific defect in Fc receptor-mediated uptake.
- The addition of 8-bromo-cyclic AMP (8-Br-cAMP) partially corrected the defect in some variants, while others remained unaffected.
Conclusions:
- The isolated variants represent distinct defects in critical steps of Fc receptor-mediated phagocytosis.
- The differential response to 8-Br-cAMP suggests multiple intracellular pathways are involved in this process.
- These cellular variants provide valuable tools for dissecting the molecular machinery of Fc receptor-mediated phagocytosis.