Journal: Oncotarget
Article Title: Novel role of miR-29a in pancreatic cancer autophagy and its therapeutic potential
doi: 10.18632/oncotarget.11928
Figure Lengend Snippet: A. qPCR analysis of miR-29 family members in normal human ductal epithelial cell lines (HPNE and HPDE) and pancreatic cancer cell lines (Panc-1, MIA PaCa-2, COLO 357, BxPC-3, AsPC-1) (n=4). Data represented as average fold change ( ΔΔ CT) ± standard error of the mean (S.E.M.) B. Pancreatic cancer cell lines (Panc-1 and MIA PaCa-2) were seeded into 96-well plates, transfected with control (CTRL) or miR-29a mimics, treated with indicated concentrations of gemcitabine (GEM) for 24 hours post-transfection, and viability was measured at 72 hours post-treatment using the Cell Counting Kit-8 (CCK-8). Average absorbance (A450) is represented (n=8) ± S.E.M. C. Panc-1 cells were transfected with CTRL or miR-29a mimics, treated with 10μM GEM for 48 hours and lactate dehydrogenase (LDH) release was determined by substrate based activity assay (fluorescence 560/590nm). Average relative percent cytotoxicity are represented (n=3-4) ± S.E.M. D. Panc-1 cells were transfected with CTRL or miR-29a mimics. 24 hours post-transfection cells were treated with 10μM GEM for 24 hours, lysed, and caspase activity was determined by absorbance using Apo-ONE Homogeneous Caspase-3/7 Assay according to manufacturer's protocol. Average relative fluorescence (RFU, 490/530nm) are represented (n=4) ± S.E.M. E. Panc-1 transfected with CTRL or miR-29a mimics, treated with 10μM GEM for 12 hours and 15ug of total cell protein lysate was subjected to western blot analysis for procaspase-3, cleaved caspase-3, and GAPDH was used as loading control. Relative quantification of band intensities normalized to GAPDH are shown below respective blots. All experiments were repeated 3-4 times and representative data is presented. *p<0.05, **p<0.01, non-significant (n.s.).
Article Snippet: Normal human pancreatic epithelial cell lines HPNE (ATCC, CRL-4023) and HPDE (AddexBio, T0018001) were grown in Dulbecco's Modified Eagle Medium (DMEM) (Life Technologies, 11965-092) supplemented with 10% fetal bovine serum (FBS).
Techniques: Transfection, Control, Cell Counting, CCK-8 Assay, Activity Assay, Fluorescence, Western Blot, Quantitative Proteomics
