AIMTo investigate the effect of different peroxisome proliferator-activated receptors (PPARs) activators on plasminogen activator inhibitor-1 in HepG-2 cell line and explore the effect of PPARs on PAL-1 gene expression.

METHODSStearic acid, oleic acid, linoleic acid, fenofibrate, pioglitazone were used in the treatment of HepG-2 cell culture. The level of PAI-1 and PPARs mRNA was measured by reverse transcription-polymerase chain reaction (RT-PCR) and the level of PAI-1 activity and PPARs protein was determined by colorimetric assay and western blotting respectively.

RESULTSThe mRNA and activity of PAI-1 significantly increased in the groups of oleic acid and linoleic acid compared with the control, but decreased in the group of fenofibrate. There were no significant changes in both groups of stearic acid and pioglitazone. The alterations in the level of PPARs mRNA and protein were not detected in all the treated groups compared with the control.

CONCLUSIONPeroxisome proliferator-activated receptors activators play important roles in the PAI-1 gene expression and regulation. It is likely mediated by the activation of PPARalpha, but there might be other mechanisms.

Fenofibrate ; pharmacology ; Hep G2 Cells ; Humans ; Linoleic Acid ; pharmacology ; Oleic Acid ; pharmacology ; Peroxisome Proliferator-Activated Receptors ; agonists ; metabolism ; Plasminogen Activator Inhibitor 1 ; genetics ; metabolism ; RNA, Messenger ; genetics

The balance between tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) regulates fibrinolysis. PAI-1 expression increases in atherosclerotic arteries and vascular smooth muscle cells (VSMCs) are one of major constituents of atheroma. We investigated the impact of lysophosphatidylcholine (lysoPC), an active component of oxidized low-density lipoprotein, on the plasminogen activator system of the rat VSMCs. The lysoPC stimulated the protein and gene expressions of PAI-1 but did not affect the protein expression of t-PA. Fibrin overlay zymography revealed that lysoPC increased the activity of PAI-1 in the conditioned media, while concurrently decreasing that of free t-PA. Vitamin E inhibited the lysoPC-induced PAI-1 expression. Further, lysoPC increased the intracellular reactive oxygen species (ROS) formation. Caffeic acid phenethyl ester, an inhibitor of NF-kappaB, blocked this lysoPC effect. Indeed, lysoPC induced the NF-kappaB-mediated transcriptional activity as measured by luciferase reporter assay. In addition, genistein, an inhibitor of protein-tyrosine kinase (PTK), diminished the lysoPC effect, while 7,12-dimethylbenz[a]anthracene, a stimulator of PTK, stimulated PAI-1 production. In conclusion, lysoPC does not affect t-PA expression but induces PAI-1 expression in the VSMC by mediating NF-kappaB and the genistein-sensitive PTK signaling pathways via oxidative stress. Importantly, lysoPC stimulates the enzyme activity of PAI-1 and suppresses that of t-PA.
Animals ; Benz(a)Anthracenes/pharmacology ; Caffeic Acids/pharmacology ; Cells, Cultured ; Genistein/pharmacology ; Lipoproteins, LDL/metabolism ; Lysophosphatidylcholines/*pharmacology ; Muscle, Smooth, Vascular/cytology/*drug effects/metabolism ; NF-kappa B/antagonists & inhibitors/metabolism ; Oxidative Stress/drug effects ; Phenylethyl Alcohol/analogs & derivatives/pharmacology ; Plasminogen Activator Inhibitor 1/agonists/genetics/*metabolism ; Protein Kinase Inhibitors/pharmacology ; Protein-Tyrosine Kinases/antagonists & inhibitors/metabolism ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species/metabolism ; Signal Transduction/drug effects ; Tissue Plasminogen Activator/*metabolism ; Transcription, Genetic/drug effects ; Up-Regulation/drug effects ; Vitamin E/pharmacology

The peroxisome proliferator activated receptor (PPAR)gamma agonist is used as antidiabetic agent with antihyperglycemic and antihyperinsulinemic actions. Beyond these actions, antifibrotic effects have been reported. We examined antifibrotic effects of PPARgamma agonist and interaction with angiotensin receptor antagonist in the unilateral ureteral obstruction (UUO) model. After UUO, mice were divided to four groups: no treatment (CONT), pioglitazone treatment, L158809 treatment, and L158809+ pioglitazone treatment. On day 14, CONT mice showed severe fibrosis and all treated mice showed decreased fibrosis. The immunohistochmistry of PAI-1, F4/80 and p-Smad2 demonstrated that their expressions were increased in CONT group and decreased in the all treated groups compared to CONT. PAI-1 and p-Smad2 determined from Western blotting, among treated groups, was decreased compared to CONT group. The expression of TGF-beta1 from real time RT PCR showed markedly increased in the CONT group and decreased in all treated groups compared to CONT. These data suggest the pioglitazone inhibited tubulointerstitial fibrosis, however, the synergism between pioglitazone and L158809 is not clear. Considering decreased expression of PAI-1 and TGF-beta/Smad2 in the treated groups, PAI-1 and TGF-beta are likely linked to the decreased renal tubulointerstitial fibrosis. According to these results, the PPARgamma agonist might be used in the treatment of renal fibrotic disease.
*Angiotensin Receptor Antagonists ; Animals ; Antigens, Differentiation/metabolism ; Disease Models, Animal ; Fibrosis ; Hypoglycemic Agents/pharmacology ; Kidney/metabolism/*pathology ; Male ; Mice ; Mice, Inbred C57BL ; PPAR gamma/*agonists ; Plasminogen Activator Inhibitor 1/metabolism ; Smad2 Protein/metabolism ; Thiazolidinediones/pharmacology ; Transforming Growth Factor beta1/genetics/metabolism ; Ureteral Obstruction/metabolism/pathology