Figure 2.

The knockdown of SPR inhibited the proliferation of MDA-MB-231 and MDA-MB-468 breast cancer cells. (A) The SPR expressions in the non-tumorigenic mammary gland epithelial cell lines MCF10A and MDA-MB-231 and MDA-MB-468 triple-negative breast cancer cells were analyzed using western blot. (B, C) The MDA-MB-231 and MDA-MB-468 breast cancer cells were transfected with NC, siSPR#1, siSPR#2, and siSPR#3 for 4 h, then replaced with fresh DMEM containing 10% FBS for another 48 h culture. mRNA and protein samples were harvested, the mRNA expression of SPR was analyzed using qRT-PCR (B), and the SPR protein expression was analyzed using western blot (C). (D) After the transfection of siSPR#3, 1000 cells were seeded into 6-well plates and cultured for 14 days, then the cells were stained with a crystal violet solution. (E) After the transfection of siSPR#3, 5×10⁴ cells were seeded into 12-well-plates and cultured for 3 days, and the cells were counted every 24 h. (F) After the transfection of siSPR#3 and culturing the cells to 90% confluence, the cells were harvested, and the mTOR, p-mTOR, p70S6k, and p-p70S6k expressions were analyzed using western blot. The data are represented as the mean ± SD in triple. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.