Supplementary Figure 7.

Cyclophilin inhibition by siRNA. HepG2215 cells were transfected with siRNAs specific for CypA, CypC, or CypD, or scramble siRNA (negative control) at baseline. (A) The cells subsequently were cultured for up to 96 hours and the expression of the respective cyclophilin mRNA was quantitated by real-time PCR in cell extracts collected every 24 hours, relative to the expression in the controls: cells transfected with scramble siRNA. Mean values and the SD of 3 independent experiments are shown. (B) In the same experiment, the knock-down effect of siRNA at a protein level was assessed by quantitation of CYPA in the HepG2215 cells by ELISA. (C) The specificity of siRNA in reducing the CYPA expression in HepG2215 cells was tested by Western blotting to detect CYPA, CYPB, and CYPD. (D) Western blot of HuH-7 cells, transfected with CypA short hairpin RNA (shRNA), confirms the marked reduction of CYPA in these cells. *P values less than .05, and **P values less than .01 for Cyp siRNA-treated vs scramble siRNA-treated cells.