Sphingosine 1-phosphate induced by hypoxia increases the expression of PAI-1 in HepG2 cells via HIF-1α

Our group has recently reported that in the immortal human HepG2 liver cell line, sphingosine 1-phosphate (S1P) increases transcription of plasminogen activator inhibitor type-1 (PAI-1), the major physiological inhibitor of fibrinolysis, within 4 h. The present study aimed to elucidate the molecular mechanisms underlying this effect. PAI-1 expression was measured by reverse transcription-quantitative polymerase chain reaction and immunoblotting. It was demonstrated that S1P increased PAI-1 promoter activity but did not increase the activity of promoters lacking the hypoxia responsive element (HRE) 2. In addition, S1P transiently increased the concentration of hypoxia inducible factor (HIF)-1 alpha, a transcription factor capable of binding to HRE. When HIF-1 alpha was knocked down, the induction of transcription of PAI-1 by S1P was no longer observed. Sphingosine kinase (SPHK) activity is increased by hypoxia. It was demonstrated that increases in the concentration of the HIF-1 alpha protein induced by hypoxia were prevented by treatment with SPHK inhibitor or S1P receptor antagonists. Thus, modification of the induction of HIF-1 alpha by S1P, leading to increased transcription of PAI-1, may be an attractive therapeutic target for thrombosis and consequent inhibition of fibrinolysis associated with hypoxia.