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Tumor-promoting phorbol esters stimulate C3b and C3b' receptor-mediated phagocytosis in cultured human monocytes
Abstract:
Monocytes were isolated in high yield (approximately 80%) and purity (greater than 90%) by Percoll gradient centrifugation and incubated in Teflon culture vessels. Using this culture method, we routinely recovered 80% of the cells originally placed into culture. Studies of the C3b and C3b' receptors of these monocytes showed that the function of both receptors could be dramatically altered by treating the cells with tumor-promoting phorbol esters. Both C3b and C3b' receptors of human monocytes efficiently mediate attachment of erythrocytes coated with the corresponding ligands, but do not promote their ingestion. However, monocytes treated with phorbol myristate acetate (PMA) or phorbol didecanoate ingest C3b- and C3b'-coated erythrocytes. Phorbol esters that are inactive as tumor promoters do not stimulate C3 receptor-mediated phagocytosis. The ability of monocytes to respond to PMA by activation of C3 receptors is developmentally regulated. Freshly isolated monocytes do not take up C3b- or C3b'-coated erythrocytes in response to PMA, but after 3 d of culture they show strong PMA-stimulated uptake. The stimulatory effect of PMA on monocyte C3b and C3b' receptor function occurs within minutes, is stable for hours, is cycloheximide insensitive, and can be inhibited with colchicine. Several lines of evidence indicates that phagocytosis of C3b or C3b'-coated erythrocytes is specifically mediated by the monocytes' C3b and C3b' receptors. First, erythrocytes attached to monocytes with concanavalin A are not ingested when the monocytes are treated with PMA. Second, monocytes plated on IgG-bearing substrates lose Fc receptor activity on their nonadherent surfaces but retain the capacity to ingest C3b- or C3b'-coated erythrocytes after PMA treatment. Third, PMA-treated monocytes plated on C3b-coated surfaces lose C3b receptor activity on their nonadherent surfaces but retain the capacity to ingest C3b'-coated erythrocytes. Conversely, PMA-treated monocytes plated on C3b'-coated surfaces show reduced C3b' receptors activity on their nonadherent surfaces but retain the capacity to ingest C3b-coated erythrocytes.
Insights
Tumor-promoting phorbol esters, like PMA, activate human monocyte receptors for C3b and C3b
Area of Science:
- Immunology
- Cell Biology
- Complement System
Background:
- Monocytes play crucial roles in the immune system, including phagocytosis.
- Complement receptors, such as those for C3b and C3b', are involved in monocyte recognition of pathogens.
- The functional regulation of these receptors is key to understanding monocyte-mediated immune responses.
Purpose of the Study:
- To investigate the effect of tumor-promoting phorbol esters on the function of monocyte C3b and C3b' receptors.
- To determine if these esters can induce phagocytosis mediated by C3b and C3b' receptors.
- To explore the developmental regulation and characteristics of this induced phagocytic activity.
Main Methods:
- Isolation of human monocytes using Percoll gradient centrifugation.
- Incubation of monocytes in Teflon culture vessels to maintain cell viability and function.
- Treatment of monocytes with phorbol esters, including phorbol myristate acetate (PMA), and assessment of C3b/C3b' receptor-mediated erythrocyte attachment and ingestion.
- Investigation of the time course, stability, and sensitivity to inhibitors (cycloheximide, colchicine) of PMA-induced phagocytosis.
- Experiments using concanavalin A and substrate-coated surfaces to confirm receptor specificity.
Main Results:
- Freshly isolated monocytes efficiently attach but do not ingest C3b- or C3b'-coated erythrocytes.
- Treatment with tumor-promoting phorbol esters (PMA) induces monocytes to phagocytose C3b- and C3b'-coated erythrocytes.
- This PMA-induced phagocytosis is developmentally regulated, appearing after 3 days of culture, and is specific to the C3b/C3b' receptors.
- The effect is rapid, stable, cycloheximide-insensitive, and can be blocked by colchicine.
Conclusions:
- Tumor-promoting phorbol esters, such as PMA, can dramatically alter monocyte C3b and C3b' receptor function, enabling phagocytosis.
- This activation of C3 receptor-mediated phagocytosis is a regulated process, dependent on monocyte culture duration and specific phorbol ester properties.
- The findings highlight a mechanism by which monocytes can be induced to phagocytose complement-opsonized targets, with implications for immune regulation and disease.
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