Simvastatin suppresses tissue factor expression and increases fibrinolytic activity in tumor necrosis factor-alpha-activated human peritoneal mesothelial cells.
BACKGROUND
Patients treated with peritoneal dialysis frequently suffer from recurrent peritonitis episodes. During peritonitis, inflammatory mediators are released and a serofibrinous exudate is formed in the peritoneal cavity, which promotes fibrosis and abdominal adhesion development. Human peritoneal mesothelial cells (HMC) play a critical role in maintaining the intraperitoneal balance between fibrinolysis and coagulation by expressing the fibrinolytic enzyme tissue-type plasminogen activator (t-PA) and its specific inhibitor, plasminogen activator inhibitor-1 (PAI-1) as well as the procoagulant protein, tissue factor.


METHODS
Cultured HMC were used to examine the effect of a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, simvastatin, on the expression of t-PA, PAI-1 and tissue factor after activation of the cells with tumor necrosis factor-alpha (TNF-alpha). Antigen concentrations in the cell supernatants were measured by enzyme-linked immunosorbent assay (ELISA). Northern blot analysis was conducted for mRNA expression. Luciferase reporter gene assays and Western blot analysis in human fibrosarcoma HT1080 cells and HMC were performed to analyze the effect of simvastatin on the transcription factors nuclear factor kappa B (NF-kappa B) and activator protein-1 (AP-1), which regulate tissue factor gene expression.


RESULTS
Incubation of HMC with TNF-alpha resulted in significantly decreased t-PA and increased PAI-1 synthesis. In the presence of simvastatin t-PA synthesis in control and TNF-alpha-treated cells dose-dependently increased, reaching 5.8-fold and 7.7-fold higher t-PA levels, respectively, at 5 micromol/L simvastatin after 48 hours. Simvastatin dose-dependently suppressed PAI-1 production in both control and TNF-alpha-treated cells. At 5 micromol/L, simvastatin lowered PAI-1 synthesis 3.4-fold and 4.0-fold, respectively, thereby also completely suppressing the TNF-alpha effect itself. Similarly, simvastatin down-regulated the expression of tissue factor and also completely opposed the TNF-alpha-induced tissue factor expression. The effects of simvastatin on t-PA, PAI-1 and tissue factor expression were prevented by mevalonate and geranylgeraniol (GG), suggesting the involvement of geranylgeranyl-modified intermediates in simvastatin's mode of action. Also, simvastatin reduced NF-kappa B- and AP-1-dependent reporter gene activity in TNF-alpha-treated HT-1080 fibrosarcoma cells and reduced the nuclear levels of p50-NF-kappa B, p65-NF-kappa B, and the AP-1 components c-fos and c-jun in HMC.


CONCLUSIONS
The HMG-CoA reductase inhibitor simvastatin is an effective stimulator of the mesothelial fibrinolytic capacity and suppresses the procoagulant activity both under normal and inflammatory conditions. Our findings provide a molecular explanation for the anti-inflammatory properties of statins in HMC and a rationale for the use of these drugs to protect peritoneal dialysis patients from peritoneal fibrosis and adhesion development during bacterial peritonitis.
Haslinger
Bettina
B
Gaubius Laboratory, TNO Prevention and Health, Leiden, The Netherlands.
Kleemann
Robert
R
Toet
Karin H
KH
Kooistra
Teake
T
D000970:Antineoplastic Agents / Q000494:pharmacology; D002478:Cells, Cultured; D004224:Diterpenes / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects; D004848:Epithelium; D005342:Fibrinolysis / Q000187:drug effects*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors / Q000494:pharmacology*; D008798:Mevalonic Acid / Q000494:pharmacology; D009852:Omentum / Q000166:cytology; D010530:Peritoneal Dialysis; D010537:Peritoneum / Q000166:cytology*; D010538:Peritonitis / Q000188:drug therapy; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics; D019821:Simvastatin / Q000494:pharmacology*; D013925:Thromboplastin / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000970:Antineoplastic Agents; D004224:Diterpenes; D019161:Hydroxymethylglutaryl-CoA Reductase Inhibitors; D017395:Plasminogen Activator Inhibitor 1; D014409:Tumor Necrosis Factor-alpha; D013925:Thromboplastin; D019821:Simvastatin; C017338:geranylgeraniol; D008798:Mevalonic Acid