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. 2019 Oct 10;11(10):1527. doi: 10.3390/cancers11101527

Figure 3.

Figure 3

Figure 3

PAK1 induces mammosphere formation and regulates hallmark breast CSC processes. The PAK1 inhibitor IPA-3 and the PAK1 inactivator ivermectin were included in mammosphere induction cultures for one week. (A,B) The effect of IPA-3 and ivermectin on mammosphere formation. Primary mammospheres were treated with IPA-3 (10 and 20 µM) or ivermectin (2.5 and 5 µM). Images were captured by microscopy. (C,D) Effect of PAK1 on mammosphere formation assessed using PAK1 siRNA. PAK1-silenced cells were seeded in ultralow plates and cultured for seven days. Images were captured by microscopy at 10× magnification. The data are presented as the mean ± SD; n = 3; * p < 0.05 vs. control. (E) Effect of ivermectin, which inactivates PAK1, on the breast CSC hallmark CD44+/CD24−-positive cell population. The ivermectin (5 μM)-treated CD44+/CD24−-cell population was analyzed by flow cytometry. Ten thousand cells were collected. Gating was based on binding of the control antibody (red cross). (F) Effect of ivermectin on ALDH-positive cells. The breast cancer cells were analyzed using an ALDEFLUOR assay. The right panel shows ALDH-positive cancer cells with N, N-diethylaminobenzaldehyde (DEAB), and the left panel represents ALDH-positive cancer cells without DEAB. The ALDH-positive population was gated in a box (red dotted line box).