Faculty Collaboration Database

Slow channel calcium inhibition blocks proinflammatory gene signaling and reduces macrophage responsiveness. J Trauma 2002 Mar;52(3):434-42

Date

03/20/2002

Pubmed ID

11901316

DOI

10.1097/00005373-200203000-00004

Scopus ID

2-s2.0-0036119664 (requires institutional sign-in at Scopus site)   39 Citations

Abstract

BACKGROUND: This study investigates the possible intracellular mechanisms responsible for calcium antagonist protection in tissue-fixed macrophages, a central modulator of the proinflammatory phenotype.

METHODS: Rabbit alveolar macrophages were exposed to lipopolysaccharide in the presence of different specific calcium antagonists. Cellular and nuclear protein were extracted and analyzed by Western blot for the phosphorylated forms of PYK2, ERK 1/2, and p38, and nuclear translocation of NF-kappaB and AP-1. Tumor necrosis factor-alpha (TNF-alpha) expression was measured by an L929 bioassay on cellular supernatants. Statistical analysis was performed by unpaired Student's t tests.

RESULTS: Cells pretreated with 100 to 500 micromol/L of diltiazem or 50 to 100 micromol/L of verapamil, both slow channel calcium blockers, led to dose-dependent reductions in lipopolysaccharide-induced PYK2 and ERK 1/2 phosphorylation, and nuclear translocation of AP-1 when compared with controls (p < 0.05). Neither inhibitor had any significant effect on p38 or NF-kappaB translocation. EGTA an extracellular calcium chelator, had no significant effect on any intracellular process studied. A dose-dependent reduction in TNF-alpha production was demonstrated with diltiazem and verapamil (p < 0.05), with no effect induced by EGTA.

CONCLUSION: Slow channel calcium influx is essential for optimal intracellular signaling through PYK2 and ERK 1/2. This reduced intracellular signaling correlated with reduced AP-1 translocation and TNF-alpha production. Extracellular calcium chelation had no significant effect on intracellular signaling or TNF-alpha production. This study further elucidates the protective mechanism of action of calcium channel blockade by diltiazem and verapamil by reducing intracellular calcium release and down-regulating the excessive proinflammatory phenotype.

Author List

Cuschieri J, Gourlay D, Garcia I, Jelacic S, Maier RV

Author

David M. Gourlay MD Chief, Professor in the Surgery department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Analysis of Variance
Animals
Blotting, Western
Calcium Channel Blockers
Diltiazem
Dose-Response Relationship, Immunologic
Focal Adhesion Kinase 2
Lipopolysaccharides
Macrophages, Alveolar
Mitogen-Activated Protein Kinases
NF-kappa B
Protein-Tyrosine Kinases
Rabbits
Sepsis
Signal Transduction
Transcription Factor AP-1
Tumor Necrosis Factor-alpha
Verapamil