Fig 2: DDR1 is expressed in luminal epithelial cells and regulates basal/myoepithelial cells. (A) Immunofluorescence of mammary glands from DDR1-/- mice was conducted by using anti-DDR1 (green) and anti-a-SMA (red) antibodies. DDR1 was detected in the luminal cell compartment but not on myoepithelial cells. (B,C) Carmine red whole-mount staining of representative mammary glands showed that mammary development of DDR1-/- mice was delayed at 5 wk but promoted at 8 wk. (B) The mammary ducts reached the dashed lines at 5 wk. (C) Primary ductal length (in millimeters) (left panel) and frequency of mammary branching (branches per millimeter) (right panel) are shown. Data are shown as mean ± SD. n = 3–4. (*) P < 0.05, unpaired Student's t-test. (D,E) Flow cytometry analysis was conducted using anti-CD24 and anti-CD49f antibodies to examine the relationship between mammary development and epithelial populations. (D) Representative data are shown. (Basal/Myo) Basal/myoepithelial cells; (Lu) luminal cells; (Strm) stromal cells. (E) Basal/myoepithelial (CD24low CD49fhigh) cells increased in 8-wk-old DDR1-/- mammary glands. (*) P < 0.05, one-way ANOVA and unpaired Student's t-test. (F) Traction stresses generated on collagen-coated polyacrylamide gels by luminal (CD24high CD49flow) and basal/myoepithelial (CD24low CD49fhigh) cells isolated from wild-type murine mammary glands by flow cytometry. The results shown represent the maximum traction stress generated by single cells. (*) P < 0.01, unpaired Student's t-test. (G,H) K14- luminal cells and K14+ basal/myoepithelial cells were isolated from K14/DDR1+/+ mice and K14/DDR1-/- mice by flow cytometry (shown in G). K14-/DDR1+/+ and K14-/DDR1-/- were mixed and aggregated with K14+/DDR1+/+ and K14+/DDR1-/- for a three-dimensional (3D) branching assay. (H) K14+/DDR1-/- basal/myoepithelial cells promoted mammary branching. K14+/DDR1+/+ cells did not promote mammary branching of K14-/DDR1-/- luminal cells.

Fig 3: Immunohistochemical staining of DDR1 in cancer tissue microarray cores obtained from radical prostatectomy specimens. Representative image of a core containing normal prostate glands that display weak basolateral DDR1 protein expression (A), with outlined area shown at higher magnification (B) using the Rab-819 antibody. GS 6 (3 + 3) lesions with strong membranous DDR1 immunostaining (C, D). High grade lesion (GS 9 [4 + 5]) with weak cytoplasmic but non-detectable membranous DDR1 staining (E, F)

Fig 4: E0771M-AP NPs mediated tumor microenvironment remodeling through attenuating hypoxia. (A) Representative images of photoacoustic imaging to assess blood oxygen level within tumor. The yellow curve marked non-hypoxic area, the green curve marked as hypoxic area, and the red curve marked as necrotic area. (B) Quantization results of ROI (along the margin of tumor) of tumor photoacoustic images in different groups (n = 5). (C) Flow cytometry was used to analyze the proportion of CD8+ T (PE) cells and CD4+ T (APC) cells in immune cells (FITC anti-CD3) of different body parts (n = 5). (D) Representative tissue immunofluorescence images of the expression of CD8 (green), CD4 (red), and HIF-1a (pink) in tumor tissue of different mouse groups. The yellow curve showed the region of HIF-1a high expression and CD8 low expression, scale bar: 100 µm (E&F) Image J quantization results of CD8, HIF-1a fluorescence in immunofluorescence from left to right successively. (Group: Control, ARS, E0771M-AP NPs, n = 10; group: PD1, n = 8; group: ARS + PD1, n = 7; group: PD1+E0771M-AP NPs, n = 4). (G) Western Blot analysis of the DDR1, HIF-1a, MMP-14, a-SMA expression on tumor tissue in different groups (n = 2). Data are represented as mean ± SD.

Fig 5: Expression of DDR1, BIK and MT1-MMP in basal and luminal breast cancer cell lines. Heat map depicting the relative expression of E-cadherin (CDH1, luminal marker), vimentin (VIM, basal marker), DDR1, DDR2, a2 integrin, a11 integrin, av integrin, COL1A1, MT1-MMP, and BIK in 58 breast cancer cell lines.