Fig 1: Increased stromal EFNB1 and EFNB3 is associated with enhanced proliferation of BPH1 cells and stromal remodeling in vivo. (a) Hematoxylin and eosin (H&E) stained sections of the tumors resulting from grafts of BPH1 with engineered BHPrS1 cell lines. Grafts of BPH1 with BHPrS1EFNB1 and BHPrS1EFNB3 show pronounced inflammatory infiltrates compared to BHPrS1EV. (b) Immunohistochemical (IHC) staining showing pronounced Ki67 expression in BPH1 tumors with BHPrS1EFNB1 and BHPrS1EFNB3 and reduced Ki67 expression in BHPrS1EFNB2 compared to BHPrS1EV. Dot plot showing Ki67 expression quantification (* p < 0.05, one-way ANOVA). (c) Higher collagen deposition in BPH1 tumor grafts with BHPrS1EFNB1 and BHPrS1EFNB3 compared to BHPrS1EV as shown by picrosirius red staining. (d) ECM remodeling marker Tenascin-C (TN-C) is highly expressed in tumor grafts of BPH1 with BHPrS1EFNB1 and BHPrS1EFNB3 compared to BHPrS1EV. Grafts of BPH1 with BHPrS1EFNB2 and BHPrS1EV has relatively lower expressions of TN-C. k in the figure represent Kidney. Scale bars in yellow and black lines in the figures represent that pictures were taken at the same magnification.
Fig 2: Overexpression of Ephrin B ligands in normal prostate fibroblasts induce the activation of cancer associated fibroblasts (CAF) markers. (a) Expression of EFNB1, EFNB2, and EFNB3 ligand proteins in lentivirus transduced BHPrS1 cells was validated by Western blot. The bands were quantified and normalized to β-actin and presented as fold change in protein expression compared to BHPrS1EV (n = 3 independent experiments). (b) The protein levels of Vimentin and putative CAF markers, Tenascin-C (TN-C), and alpha-smooth muscle actin (α-SMA) were evaluated in Ephrin-generated cell lines (BHPrS1EFNB1, BHPrS1EFNB2, BHPrS1EFNB3) by Western blot. The bands were quantified and normalized to β-actin and presented as the mean (* p < 0.05, one-way ANOVA, n = 3 independent experiments).
Supplier Page from DNASU for EFNB1 (Homo sapiens) in pDONR221 (Gateway donor/master vector)