macrophage(MΦ) activation is a complex process and not yet fully understood in that mΦativating signals are received from extracellular medium and transduced across the cell membrane. The mechanism by which MΦ's acqulre the tumoricidal
activity
is
of particular interest. This study mainly deals with the role of cAMP and Ca2+ in the tumoricidal activity of MΦ's. Lipopolysaccharides(LPS) and other MΦ activating signals induce the functional responses in MΦ's LPS was treated at a
concentration of
10μgm/ml which is redcognized as effective triggering concentration. To compare the difference between LJPS and linterferon-γ(IFN-γ) signal transduction, peritoneal mΦ's (PM's) and alveolar MΦ's(AM's) lackin LPS-binding sites, sites, were
used.
To prove MΦactivation, NBT reduction test as microbicidal activity and L929 tumor cell cytotoxicity and tumor necrosis fctor(TNF0 production as tumoricidal activities were investigated. PM's were activated with LPS and AM's were activated
IFN-γ(10U/ml). To examine the role of intracellular cAMP and Ca2+ in the MΦactivation, intracellular cAMP level was checked every 10 minutes by competitive binding assay using [3H]-cAMP and intracelluar Ca2+ concentration change using
fluorescent
dye
tetraacetoxymethoylester of Quin-2(Quin-2/AM). The effect of extracellular Ca2+ was checked by 45CaCl2 uptake every 5 minutes.
As a result of those experiments ,cAMP analogues such as DbcAMP an Prostaglandin E2(PGE2) inhibited L929 tumor cell cytotoxicity, NBT reduction and TNF production. CAMP inhibitors such as alloxan and indomethacin increased tumor cell
cytotoxicity.
When
treated with intracellular Ca2+ chelator, Quin-2/AM, TNF production was decreased significantly comparing to control group. But extracellular Ca2+ chelator, EDTA did not have any significant change in tumor cell cytotoxicity and TNF production.
When MΦ's were activated with LPS, intracellular cAMP level was greatly decreased within 10 minutes and recovered slowly to the normal level. The cAMP is thought to be kept at certain level in normal state, but decreased transiently as the
signal
arrived at cell membrane.
When treatd with LPS, intracelular Ca2+ level was increased within 1 minute(probably at 30 or 40 seconds) but the was no Ca2+ uptake from extracellular medium. Interestingly, when MΦ's were activated with IFN-γ, there was increase in both
intracellular and extracellular Ca2+ level, significantly. Also, Ca2+ ionophore, A23187 had priming effect in the presence of LPS, and was thought to be related to the alteration in phospholipid arrangement of MΦmembrane.
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