Figure 4. Involvement of the MARK pathway and 14-3-3β protein in statin-induced regulation of endothelial TM.

A) Decreased nuclear content and concomitant increase in cytoplasmic content of 14-3-3β protein after exposure to atorvastatin for 4–8 hours.

HUVEC treated with vehicle (V) or atorvastatin (A) for up to 24 hours. Changes in nuclear (top) and cytoplasmic (bottom) 14-3-3β protein levels were analyzed by western blot.

B) Densitometric analysis of blots shown in panel A.

C) Inhibition of MEK activation enhances the effect of atorvastatin on TM and tPA, but not onCTGFandPAI-1.

HUVECs were treated for 24 hours with vehicle (V), atorvastatin (A), pretreated with mevalonate for 30 minutes before atorvastatin treatment (M+A), PD98059 alone (Pd), U0126 alone (U), or pre-treated with PD98059 (P+A) or U0126 (U+A) for 30 minutes before atorvastatin treatment. TM, tPA, CTGF and PAI-1 mRNAs were quantified by real-time RT-PCR.

D) Knockdown of 14-3-3β augments the effect of atorvastatin on endothelial expression of TM and tPA.

siRNA was used to knock down the 14-3-3β gene. Forty-eight hours later, HUVECs or HCAECs were treated with vehicle or atorvastatin for 24 hours. TM and tPA mRNAs were quantified by real-time PCR.

Atorvastatin: 1×10⁻⁵M; mevalonate: 5×10⁻⁴M; PD98059 and U0126: 1×10⁻⁵M.

* p < 0.05 versus vehicle.