Fig. 4.

Silencing of OPN by small interfering RNA (siRNA) in PH-Fibs results in attenuated cell proliferation. A and B: different siRNAs (siRNA 1–3) were tested for their ability to block OPN protein expression in PH-Fibs and siRNA-3 was chosen as the best one. As shown by both Western blotting (A) and real-time PCR analysis (B), treatment of PH-Fibs with siRNA-3 (50 nM, 48 h for RT-PCR, 72 h for Western blotting) mediated significant downregulation of OPN expression. C: siRNA3-mediated OPN silencing in PH-Fibs resulted in decreased cell proliferation as shown by MTT assay (72-h time point). D: addition of recombinant OPN (rOPN; 10 or 20 ng/ml) to siRNA-treated PH-Fibs restored and even elevated their proliferative potential, as determined by cell count assessment. Data shown are representative of at least 3 independent experiments. NT, not treated; SCR, treated with scrambled RNA; siRNA 1–3, treated with siRNA 1, 2, or 3. *P < 0.05; **P < 0.01.