Figure 3.

Effect of H- and L-chain siRNA on de novo ferritin subunit synthesis in LECs. (A, B) LECs were seeded in six-well plates, then transfected with nontargeting control or FH- or FLsiRNA. Two and 4 days later, the cells were incubated with ³⁵S-methionine for 20 hours. A sample for TCA precipitation was removed from the cell lysates, followed by immunoprecipitation of ferritin from the lysates. The solubilized immunoprecipitate was subjected to 12% SDS-PAGE, and radioactivity in the ferritin H- and L-chain bands was normalized to the total amount of radioactivity incorporated into protein (TCA). The data are expressed as a percentage of that of control cells transfected with nontargeting siRNA. The histograms represent the mean ± SEM of at least eight samples. *Significantly different from control (P < 0.05); ANOVA and Tukey's test. (C) A representative gel showing newly synthesized H- and L-chain ferritin subunits labeled with ³⁵S-methionine after siRNA treatment.