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Figure 1 | Breast Cancer Research

Figure 1

From: A novel role for signal transducer and activator of transcription 5b (STAT5b) in β1-integrin-mediated human breast cancer cell migration

Figure 1

STAT5b knockdown inhibits breast cancer cell migration. (a) BT-549 or MDA-MB-231 breast cancer cells were transfected with no siRNA (con), control siRNA for luciferase (siLuc), siSTAT5b SMARTpool (BT-549 cells), or siSTAT5b SMARTpool duplex #3 (MDA-MB-231 cells). Seventy-two hours after transfection, cells were plated in serum-free media in trans-well chambers. Media containing 1% fetal bovine serum (FBS; BT-549) or 10% FBS (MDA-MB-231) were placed in the lower chambers. After 3 hours (BT-549) or 6 hours (MDA-MB-231), cells were fixed, stained with crystal violet, and the number of migratory cells was counted. Results are graphed as the average number of migratory cells per field ± SEM. One-way ANOVA with Tukey's post-test was used to determine statistical significance (P < 0.05) between the following: BT-549: con and siSTAT5b (*), siLuc and siSTAT5b (black circles); n = 5. MDA-MB-231: con and siSTAT5b (*), siLuc and siSTAT5b (black circles); n = 4. (b) BT-549 breast cancer cells were transfected and plated in serum-free media in trans-well chambers for 3 hours. Media containing varying concentrations of FBS (0.1% to 10%) were placed in the lower chambers. One-way ANOVA with Tukey's post-test was used to determine statistical significance (P < 0.05) between the following: con and siSTAT5b (*), siLuc and siSTAT5b (black circles); n = 3. (c) MDA-MB-231 cells were transfected with no siRNA (con), control siRNA for luciferase (siLuc), or siRNA specific to STAT5a/b (siSTAT5b siGENOME SMARTpool), STAT5a (siSTAT5a custom oligonucleotide), or STAT5b (siSTAT5b siGENOME SMARTpool oligonucleotide 3), and trans-well migration assays were performed as described. One-way ANOVA with Tukey's post-test was used to determine statistical significance (P < 0.05) between the following: con and siSTAT5a/b (*), con and siSTAT5b (*), siLuc and siSTAT5a/b (black circles), siLuc and siSTAT5b (black circles); n ≥ 5. (a and c) Whole-cell lysates from siRNA-transfected cells were immunoblotted with antibodies specific for STAT5a, STAT5b, and β-actin as a loading control.

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