Abstract
Bacterial lipopolysaccharide (LPS) is a potent inflammatory agent capable of producing liver injury, the pathogenesis of which depends on numerous mediators, including thrombin. Previous studies showed that thrombin promotes LPS-induced liver injury independent of its ability to form fibrin clots. In isolated, buffer-perfused livers from LPS-treated rats, thrombin added to the perfusion buffer caused dose-dependent liver injury with an EC50 value of 0.4 nM, consistent with activation by thrombin of a protease-activated receptor (PAR). Actions of thrombin at PARs can be mimicked by thrombin receptor-activating peptides (TRAPs). TRAPs for PAR-1 reproduced the injury caused by thrombin in isolated livers, suggesting that one mechanism by which thrombin promotes LPS-induced liver injury is by activating PAR-1. Immunocytochemistry demonstrated the presence of PAR-1 on sinusoidal endothelial cells and Kupffer cells but not on parenchymal cells or neutrophils. Previous studies showed that thrombin interacts with neutrophils in the genesis of liver injury after LPS treatment. To explore this interaction further, the influence of thrombin on mediators that modulate neutrophil function were evaluated. Inhibition of thrombin in LPS-treated rats prevented liver injury but did not prevent up-regulation of cytokine-induced neutrophil chemoattractant-1, macrophage inflammatory protein-2, or intercellular adhesion molecule-1. Thrombin inhibition did, however, prevent neutrophil (PMN) degranulation in vivo as measured by plasma elastase levels. In addition, elastase concentration was increased in the perfusion medium of livers isolated from LPS-treated rats and perfused with TRAPs. These results suggest that activation of PAR-1 after LPS exposure promotes PMN activation and hepatic parenchymal cell injury.
Footnotes
-
This work was supported by National Institutes of Health Grant DK50728. F.M. and B.C. were supported, in part, by National Institutes of Health Training Grant T32 ES07255. B.C. was also supported by NRSA ES05866 from National Institutes of Health.
-
DOI: 10.1124/jpet.102.046391
- Abbreviations:
- LPS
- lipopolysaccharide
- PMN
- neutrophil
- PAR
- protease-activated receptor
- TRAP
- thrombin receptor-activating peptide
- SEC
- sinusoidal endothelial cell
- ICAM-1
- intercellular adhesion molecule-1
- FBS
- fetal bovine serum
- PBS
- phosphate-buffered saline
- SFFLRN
- Ser-Phe-Phe-Leu-Arg-Asn
- TFLLR
- Thr-Phe-Leu-Leu-Arg
- NRLFFS
- Asn-Arg-Leu-Phe-Phe-Ser
- RLLFT
- Arg-Leu-Leu-Phe-Thr
- ALT
- alanine aminotransferase
- DAPI
- 4′6-diamidino-2-phenylindole dihydrochloride
- CINC-1
- cytokine-induced neutrophil chemoattractant-1
- MIP-2
- macrophage inflammatory protein-2
- RT-PCR
- reverse transcription-polymerase chain reaction
- ELISA
- enzyme-linked immunosorbent assay
- ANOVA
- analysis of variance
- EU
- endotoxin unit
- Received November 6, 2002.
- Accepted January 21, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|