Differential effect of diltiazem and TA-3090 on calcium homeostasis of neutrophils.
The effects of diltiazem and TA-3090, an 8-chloro analog of diltiazem, on cellular responses and calcium homeostasis of human neutrophils were investigated. TA-3090, at 10 to 20 microM, enhanced lysozyme release and superoxide generation induced in neutrophils by n-formyl-methionyl-leucyl-phenylalanine (FMLP). Higher concentrations of TA-3090 inhibited responses at IC50s between 70 and 85 microM. Diltiazem by comparison inhibited responses at an IC50 of about 200 microM. The two drugs had little or no effect on early signaling events: inositol 1,4,5-trisphosphate formation triggered by FMLP was not affected. Moreover, 500 microM TA-3090 or diltiazem did not significantly affect FMLP-triggered Ca2+ transients. (Cytoplasmic free Ca2+ levels ([Ca2+]i) were monitored in fura-2-loaded neutrophils.) Diltiazem alone caused a limited influx of extracellular Ca2+ which increased basal [Ca2+]i by twofold. Internal Ca2+ stores were not released. TA-3090, in contrast, induced a biphasic rise in [Ca2+]i--an initial mobilization of intracellular Ca2+ stores was followed after 10-15 min by a persistent influx of extracellular Ca2+ which increased [Ca2+]i to 1.3 +/- 0.7 (SD) microM. Complementary studies with semipermeabilized neutrophils showed that TA-3090 but not diltiazem directly released Ca2+ from intracellular stores. In TA-3090-treated cells, lactate dehydrogenase release was correlated with delayed influx of extracellular Ca2+. The chelation of extracellular Ca2+ by EGTA prevented LDH release. Present results show that TA-3090 and diltiazem initially blocked cell signaling at steps subsequent to phospholipase C activity. With TA-3090-treated cells, elevated [Ca2+]i ensuing from prolonged incubations likely activated inappropriate reactions leading to cell lysis and death.
Wong
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K
Department of Medicine, University of Alberta, Edmonton, Canada.
Kwan-Yeung
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Ng
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D002118:Calcium / Q000097:blood*; D002121:Calcium Channel Blockers / Q000494:pharmacology* / Q000633:toxicity; D003571:Cytochalasin B / Q000494:pharmacology; D004110:Diltiazem / Q000494:pharmacology* / Q000633:toxicity / Q000031:analogs & derivatives; D006706:Homeostasis / Q000187:drug effects*; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000096:biosynthesis; D007424:Intracellular Fluid / Q000378:metabolism; D009113:Muramidase / Q000097:blood; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D015290:Second Messenger Systems / Q000187:drug effects; D013481:Superoxides / Q000378:metabolism; D013841:Thiazepines / Q000494:pharmacology* / Q000633:toxicity
D002121:Calcium Channel Blockers; D013841:Thiazepines; D013481:Superoxides; D003571:Cytochalasin B; C056595:clentiazem; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015544:Inositol 1,4,5-Trisphosphate; D009113:Muramidase; D004110:Diltiazem; D002118:Calcium