Fig. 1

Grem1 is highly expressed in 66cl4 and GREM1 correlates with RFS in breast cancer patients. (a) Scatter plot of in vivo versus in vitro differential expression of mRNAs encoding secreted proteins in 66cl4 and 67NR. A positive number indicates higher expression in 66cl4, whereas a negative number indicates higher expression in 67NR. The 28 factors were significantly overexpressed in both 66cl4 cells and 66cl4 primary tumors (cut-off: fold-change ≥1.5, p-value ≤0.05) are indicated by name. (b) Relationship between GREM1 gene expression and RFS in breast cancer patients using KM plotter. High and low expression were defined as above and below median. (c) Q-PCR analysis of Grem1 mRNA expression in 67NR and 66cl4 cell lines in vitro (n = 3). Fold change is relative to Actb and Tbp. Results are shown as mean ± SEM. Student’s t-test, **0.001 < P < 0.01. (d) Representative gremlin1 immunoblot (n = 4) of whole lysates of the 67NR and 66cl4 cell lines without or with PTI for 6 h. (e) Levels of gremlin1 in conditioned medium from 67NR and 66cl4 (n = 3) measured by ELISA. Results are shown as mean ± SEM. (f) Q-PCR analysis of Bmp4 mRNA expression in 67NR and 66cl4 cell lines in vitro (n = 3). Fold change is relative to Actb and Tbp. Results are shown as mean ± SEM. Student’s t-test, **0.001 < P < 0.01. (g) Representative BMP4 immunoblot (n = 4) of whole lysates of the 67NR and 66cl4 cell lines without or with PTI for 6 h