Fig. 2

BCL-xL and miR-377 co-regulation mediates ABT-199 resistance. RT-PCR analysis of BCL-xL, miR-377, and BCL-2 expression levels after treatment with 5-Aza for 72 h in (a) OC-R and (b) S6-R cells. BCL-xL, miR-377, and BCL-2 expression levels in OC-R and S6-R were normalized to those in parental OC and S6 cells. DMSO treatment and BCL-2 expression served as negative controls. (c) Parental OC, S6 and ABT-199R OC-R and S6-R cells were treated with 5-Aza and cell viability was assessed after 72 h by annexin-V/FITC and propidium iodide staining and analyzed by flow cytometry. (d) Cell viability, analyzed by flow cytometry and (e) BCL-xL expression, assessed by immunoblot after transfecting S6-R cells with 200 nM of Non-Target (N.T), miR-377 mimic, or siBCL-xL. S6-R cells were transfected with 1 μg of pMIG-Bcl-xL expression plasmid and 24 h later transfected again with 200 nM of miR-377 mimic. After an additional 24 h, analysis of BCL-xL expression by RT-PCR (f) and cell viability (g) were determined. BCL-xL expression and cell viability were normalized to untransfected S6-R cells. Cell viability (h) and BCL-xL expression (i) were assessed in S6-R cells treated with 5-Aza for 24 h and then at 48 h following transfection with miR-377 inhibitor (72 h total time). Significance was determined by a t-test