Vascular Heme Oxygenase-1 Induction Suppresses Tissue Factor and Plasminogen Activator Inhibitor-1 Expressions

Objective - Heme oxygenase-1(HO-1), the rate-limiting enzyme of heme degradation, has recently been considered to have protective roles against various pathological conditions. 10 years have passed since we lost the first and the only patient of HO-1 deficiency. Since the patient of HO-1 deficiency showed endothelial cell injury and extremely enhanced coagulation and fibrinolytic parameters, we examined the effect of HO-1 modulation on tissue factor (TF) and plasminogen activator inhibitor-1 (PAI-1) expression on endothelial cells.

Methods and Results -

1. Human umbilical vein endothelial cell (HUVEC) was stimulated with hemin (100mM), HO-1 inducer, and mRNA and protein levels for HO-1, TF and PAI-1 were examined. Total RNA was extracted from HUVEC, and was analyzed by real time RT-PCR. Protein expression levels of HO-1, TF and PAI-1 were measured by ELISA. Hemin stimulation increased HO-1 mRNA levels by 20 times. On the other hand, TF mRNA and antigen levels were minimum even after 8 hours of stimulation. Importantly, hemin stimulation reduced PAI-1 mRNA more than half after 4 hours.

2. After HO-1 induction by hemin (100 mM) for 6 hours, HUVEC cultures were exposed to 10 ng/ml tumor necrosis factor (TNF).

Prior exposure to hemin significantly increased HO-1 mRNA by 60 times in 30 minutes after stimulation with TNF. However, TNF alone could not induce HO-1 mRNA and protein levels in HUVEC. Although stimulation with TNF enhanced expressions of both TF and PAI-1 mRNA, they were significantly inhibited more than half by prior treatment with hemin. TF antigen levels were similarly decreased (5.0 to 0.7 pg/ml). PAI-1 antigen levels were also inhibited by prior treatment with hemin (1.8 to 0.1 ng/ml)(3) To see if hemin effect on HUVEC is due to HO-1 production, HO-1 inhibitor tin-protoporphyrin IX (SnPP-IX) was added to the cultures. The inhibitor effect of hemin on TF and PAI-1 productions was cancelled when HUVEC was cocultured with SnPP-IX.

Conclusions - These results indicate that hemin exert inhibitory effect on TF and PAI-1 expressions through HO-1 production. Induction of HO-1 may be beneficial in the prevention of thrombosis associated with inflammation.