Fig. 4
Loss of DDR2 abolishes cell invasion, Snail expression, and tumor growth by PAPP-A. a Immunoblot of indicated markers in MCF-7PAPP-A CTL and MCF-7PAPP-A DDR2−/− cells. b Representative images of 48 h Transwell in vitro invasion assays of MCF-7 CTL, MCF-7 DDR2−/−, MCF-7PAPP-A CTL, and MCF-7PAPP-A DDR2−/− cells. Experiments repeated in technical triplicate. Scale bar 100 μm. Quantification is shown, unpaired t test with Welch’s correction. *p < 0.05. c Immunoblot of indicated markers in MCF-7 CTL or MCF-7 DDR2−/− cells treated with or without PAPP-A media for 24 h. d Relative tumor growth of MCF-7, MCF-7PAPP-A, and MCF-7PAPP-A DDR2−/− xenografts in a Matrigel-collagen mixture. n = 5 mice per group, ten tumors each. Each point represents the average of five mice, mean ± SEM. Unpaired t test at end point (MCF-7PAPP-A vs. MCF-7PAPP-A DDR2−/−). *p < 0.05 e Representative images of Snail IHC on MCF-7, MCF-7PAPP-A, and MCF-7PAPP-A DDR2−/− xenograft tumors in Matrigel-collagen mixture. n = 3 mice, six tumors total per group. Scale bar 100 μm. f Representative images of Masson’s trichrome collagen stain (blue) and second-harmonic generation (SHG) imaging of collagen (lower panels). Yellow dotted lines indicate the mammary tumor borders. Corresponding regions of interest (ROI) on magnified histological sections of MCF-7, MCF-7PAPP-A, or MCF-7PAPP-A DDR2−/− xenograft tumors in a Matrigel-collagen mixture. n = 3 mice per group. Scale bar 100 μm. g Quantification of TACS3 per total curvelets analyzed in MCF-7, MCF-7PAPP-A, or MCF-7PAPP-A DDR2−/− xenograft tumors in a Matrigel-collagen mixture using the CurveAlign software. TACS3 characterized as curvelet angles 60–90 relative to the ductal border. n = 3 mice, six tumors total per group. Mean + SEM, unpaired t test with Welch’s correction (comparisons between TACS3 group only, MCF-7PAPP-A vs. MCF-7PAPP-A DDR2−/−). *p < 0.05. h Immunoblot of indicated markers in MCF-7PAPP-A cells treated for 48 h at the indicated concentrations of PQ401 and imatinib. i Proliferation assay of MCF-7 and MCF-7PAPP-A cells treated with imatinib at the indicated concentrations for 48 h. j Proliferation assay of MCF-7 and MCF-7PAPP-A cells treated with PQ401 at the indicated concentrations for 48 h. k Proliferation assay of MCF-7 and MCF-7PAPP-A cells treated with PQ401 at 7.5 μM in addition to the indicated concentrations of imatinib for 48 h