Figure 6
AFP promoter/enhancer +2 c-My c shRNA delivered by F-virosomes down-regulated c-Myc . (A) Hemi fusion study in various hepatoma, untransformed and non-liver cells was done by fluorescence dequenching assay. Fusion of R18 labeled Sendai F-virosomes was determined by spectro-fluorimetry and was almost similar in the case of HepG2 and Huh7, whereas it was slightly lesser with Chang Liver cells. CHO cells, being a non-liver cell line, lack ASGPR and served as a negative control. F-virosomes with inactivated F-protein (HC: Heat Control) displayed poor fusion with the HepG2 cells. (B) Time dependant fall in the expression of c-Myc by AFPEn–Pr + 2 – myc after virosomal delivery to HepG2 cells was significantly comparable with that of LipofectamineTM 2000 (C) In HepG2 cells, AFPEn–Pr + 2 – myc construct decreases c-Myc level significantly which was comparable to that of the positive control CMVPr – myc. (D) Similar pattern was observed in the case of Huh7. (E) Down-regulation of c-Myc in untransformed Chang Liver cell line was observed only by CMVPr – myc and not by AFP promoter/enhancer driven shRNA. (F) Western Blot Analysis of c-Myc in HepG2, Huh7 and Chang Liver was in concordance with real-time PCR analysis and followed the same trend.