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Phorbol esters and calcium ionophore can prime murine peritoneal macrophages for tumor cell destruction
Abstract:
Murine macrophages from sites of inflammation develop toward tumoricidal competence by exposure to a macrophage-activating factor such as interferon-gamma (IFN-gamma). To explore the biochemical transductional events initiated by IFN-gamma, peritoneal macrophages from C57BL/6J mice elicited by various sterile irritants were treated in vitro with two pharmacologic agents that mimic the action of certain second messengers. Phorbol myristate acetate (PMA) and the ionophore A23187 cooperatively reproduced the ability of IFN-gamma to prime macrophages for tumoricidal function. Neither agent alone was able to prime macrophages. The two agents acted on the macrophages, and target susceptibility to kill was not altered by PMA and A23187. Only active phorbol esters, which are known to bind and stimulate protein kinase C, were able to cooperate with A23187 to induce priming. A cell-permeable synthetic diacylglycerol (sn-1,2-dioctanoyl glycerol) could also prime for cytolysis. In the presence of PMA, A23187, and EGTA, addition of Ca++ was sufficient for priming, whereas the addition of Mg++ was much less efficient. Priming by IFN-gamma, however, was not blocked by EGTA. Efflux of 45Ca++ from preloaded cells was significantly increased by A23187 and by IFN-gamma. Quin-2/AM, an intracellular chelator of Ca++, blocked priming by IFN-gamma. In summary, the data suggest that priming of macrophages for tumoricidal function by IFN-gamma involves, at least in part, alterations in protein kinase C and in levels of intracellular Ca++.
Insights
Interferon-gamma (IFN-gamma) primes macrophages for tumoricidal function by activating protein kinase C and altering intracellular calcium levels. This priming is crucial for enhancing macrophage anti-tumor activity.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Macrophages at inflammatory sites acquire tumoricidal competence via factors like interferon-gamma (IFN-gamma).
- Understanding the biochemical pathways initiated by IFN-gamma is essential for cancer immunotherapy research.
Purpose of the Study:
- To investigate the intracellular signaling events triggered by IFN-gamma in macrophages.
- To elucidate the roles of protein kinase C and intracellular calcium in IFN-gamma-mediated macrophage priming.
Main Methods:
- Peritoneal macrophages from C57BL/6J mice were treated with phorbol myristate acetate (PMA), ionophore A23187, and other signaling modulators.
- Intracellular calcium levels were monitored using 45Ca++ efflux and Quin-2/AM.
- Macrophage tumoricidal function was assessed after treatment with various agents.
Main Results:
- PMA and A23187 synergistically primed macrophages for tumoricidal function, mimicking IFN-gamma.
- Active phorbol esters and diacylglycerol cooperated with A23187 to induce priming, indicating protein kinase C involvement.
- IFN-gamma increased 45Ca++ efflux and priming was blocked by intracellular calcium chelation (Quin-2/AM), but not by EGTA.
- Calcium (Ca++) was sufficient for priming in the presence of PMA and A23187.
Conclusions:
- IFN-gamma priming of macrophages for tumoricidal activity involves protein kinase C activation.
- Alterations in intracellular calcium levels are a key component of IFN-gamma signaling in macrophages.
- These findings provide insights into the molecular mechanisms of macrophage activation against tumors.