Comparative Analysis of Gene Expression Data Reveals Novel Targets of Senescence-Associated microRNAs
Figure 4
Knock-down of CDCA2, ID4 or both does not induce premature senescence in PDL 33 IMR90 cells.
A) siRNAs designed to target the coding sequence of CDCA2 or ID4 were transfected, individually or as a mix, in PDL 33 IMR90 cells to knock-down the expression levels of target genes. After 72 h, transfected cells were incubated with BrdU for 4 h, then coverslips were fixed, incubated with a primary anti-BrdU antibody, washed and incubated with a secondary fluorescein-conjugated antibody, counterstained with Hoechst-33258 and counted by immunofluorescence. Counts of at least 1,000 cells were averaged and expressed as fold changes±SD, with respect to scrambled transfected cells (*** p<0.001). B) siRNA transfected cells were subcultivated for 10 days and stained for SA-β-gal. Counts of at least 300 cells were averaged and expressed as fold changes±SD, with respect to scrambled transfected cells.