Fig. 6

Focused inhibition of let-7 miRNAs regulates gamma-globin and HbF levels in human erythroblasts ex vivo. Let-7a-TuD and control transductions were investigated for a alpha-, mu-, theta- and zeta-globins, and b beta-, delta-, gamma- and epsilon-globins. RT-qPCR analyses were performed at culture day 14. c Representative HPLC tracings. HbF and HbA peaks are labeled on each graph (y-axis, mVolts; x-axis, elution time in minutes). d HPLC analysis of hemoglobin from let-7a-TuD compared to control transductions. HPLC samples were collected at culture day 21. Let-7a-TuD effects in e CA1 and f GCNT2 mRNA levels. RT-qPCR analyses were performed at culture day 14. Open bars represent control and black bars represent let-7a-TuD. Mean value ± SD of five independent donors for each condition. p values were calculated using two-tailed Student’s t-test. Western blot analyses of g CA1 and h GCNT2 in the cytoplasmic extracts of three independent donors at culture day 14 upon let-7a-TuD compared to control transductions. Blots were probed with anti-CA1 or anti-GCNT2 antibodies as indicated. Beta-Actin was used as loading control. C, control (negative control vector) transduction; 7a-TuD or 7a, let-7a tough decoy design. *p < 0.05