Supplementary Figure 4: LIF stimulates STAT3:SOCS3 induction in breast cancer cells with low metastatic potential.
(a,c,e) Western blot for LIFR, pSTAT3 (Y705), total Stat3, and β-actin or vinculin (loading control) after 15 or 30 min treatment with PBS (vehicle control), recombinant OSM (50 ng ml−1) or recombinant LIF (50 ng ml−1) in (a) PyMT and (c) D2.0R cells with low metastatic potential and (e) D2A1 cells with high metastatic potential. Blots represent 3 independent biological replicates/group. (b,d,f) SOCS3 mRNA levels after 1 or 6 h treatment with PBS, recombinant OSM (50 ng ml−1) or recombinant LIF (50 ng ml−1). h: Mann-Whitney test. j,l: Student’s unpaired t-test.n = 3 biological replicates, each being an average from 3 independent experiments. (g,i) Western blot for LIFR, pSTAT3 (Y705), total Stat3, and vinculin (loading control) after 15 or 30 min treatment with PBS (vehicle control), recombinant OSM (50 ng ml−1) or recombinant LIF (50 ng ml−1) in (g) 4T1 parental cells and (i) MDA-MB-231 parental cells. Blots represent 3 independent biological replicates/group. (h,j) SOCS3 mRNA levels after 1 or 6 h treatment with PBS, recombinant OSM (50 ng ml−1) or recombinant LIF (50 ng ml−1) in (h) 4T1 parental cells and (j) MDA-MB-231 parental cells. One-Way ANOVA. n = 3 biological replicates, each being an average from 3 independent experiments. Unprocessed blots in Supplementary Fig. 9. Graphs represent the mean/group and error bars represent standard error of the mean (SEM). ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001, and ∗∗∗∗P < 0.0001.
