Efficacy of model recombinant miR-7-5p to regulate target gene expression in human NSCLC cells. (A) BioRNAGly- and BioRNALeu-carried miR-7-5p are processed to target miR-7-5p in A549 and H1975 cells. Cells were transfected with 15 nM BioRNA/miR-7-5p, control BioRNA, or Lipofectamine 3000 (vehicle) for 48 h, and miR-7-5p levels were determined by selective stem–loop RT qPCR assay. Commercial miRCURY LNA miR-7-5p mimic (Mimic miR-7-5p) and control RNA (Mimic Control) from Qiagen were used for comparison. (B) Impact of BioRNA/miR-7-5p on a well-known miR-7-5p target, epidermal growth factor receptor (EGFR) in A549 and H1975 cells after 72-h transfection, as determined by western blot analyses. Surprisingly, BioRNAGly/miR-7-5p more effectively reduced EGFR protein levels than BioRNALeu/miR-7-5p besides control RNA. Therefore, the effects of BioRNAGly/miR-7-5p versus the mimic on (C) multidrug resistance-associated protein 1 (MPR1) and (D) voltage-dependent anion channel protein 1 (VDAC1) protein levels were further defined. miR-7-5p and targeted protein levels were normalized to corresponding U6 and β-actin or total protein, respectively, and vehicle control groups were set as 1.0. All values are mean ± SD (N = 3 biological replicates per group). (****) P < 0.0001; (***) P < 0.001; aP < 0.05, as compared to respective control RNA; bP < 0.05, compared to vehicle control; cP < 0.05, compared to Mimic miR-7-5p; dP < 0.05, compared to BioRNALeu/miR-7-5p (one-way ANOVA with Bonferroni post hoc tests).
