Figure 1

shRNA-mediated downregulation of elastase decreases proliferation of MDA-MB-231 cells. MDA-MB-231 cells were selected that stably expressed control, scrambled shRNA (231-Control1 and 231-Control2) or elastase-specific shRNA (231-Elastase1 and 231-Elastase2). A) Down-regulation of elastase was confirmed by confocal microscopy of each clone and the parental cells using Alexa Fluor 555 (red) and Alexa Fluor 488 (green) goat anti-rabbit secondary antibodies against an elastase primary antibody along with DAPI nuclear stain. B) qRT-PCR was performed on RNA from the control and elastase shRNA treated cells. A Student's t-test was used to determine the significance of the down-regulation (**P < 0.01). C and D) Cells of each clone were counted every day for five days. Three independent growth curves were generated in this manner and the average of each cell count was graphed. Error bars indicate standard error and differences between parental and elastase shRNA cells were detected using a Student's t-test (*P < 0.05, ***P < 0.001). E and F) Cells of each clone were subjected to clonogenic assays, the resulting clones were enumerated and subjected to Student's t-test (**P < 0.01, ***P < 0.001).