Calcium and calmodulin regulate lipopolysaccharide-induced alveolar macrophage production of tumor necrosis factor and procoagulant activity.

Abstract

Alterations in macrophage (M phi) function are responsible, in part, for adult respiratory distress syndrome and multiple organ failure developing in patients with sepsis. Elucidation and control of these M phi mechanisms during sepsis are crucial to our understanding of this disease and, ultimately, to improving survival of these patients. To investigate the involvement of calcium flux in endotoxin-induced alveolar M phi production of tumor necrosis factor (TNF) and procoagulant (PC) activity. Rabbit alveolar M phi obtained by bronchoalveolar lavage were exposed to endotoxin in the form of lipopolysaccharide (LPS) extracted from Escherichia coli 0111:B4 in the presence of different specific calcium agonists and antagonists. The TNF expression was measured in the supernatant by L929 bioassays. The PC activity was determined in cell lysates by a one-step coagulation assay. Macrophages activated by LPS produce enormous levels of TNF and PC. Either W7 (20 mumol/L), a calmodulin antagonist, or TMB-8 (50 mumol/L), which prevents calcium release from the endoplasmic reticulum, inhibited production of both TNF and PC activity. Verapamil (50 mumol/L) alone or combined with TMB-8 significantly inhibited both TNF and PC production by LPS-stimulated M phi. Elevating intracellular calcium ([Ca2+]i), using the calcium ionophore, A23187, or thapsigargin alone, did not induce M phi production of TNF but significantly augmented LPS-stimulated TNF production. Our results indicate that increased intracellular calcium causing signal transduction activation through the calmodulin pathway is a necessary, but insufficient, component of the LPS signaling in M phi.