Additional file 2: of The phospholipase DDHD1 as a new target in colorectal cancer therapy

Figure S1. DDHD1 silencing. To evaluate DDHD1 silencing a. Real-time PCR and b. Western blot analysis were performed on SW480, HCT116, HS5 and HUVEC transfected for 48 or 72 h with scrambled siRNA or DDHD1 siRNA. (TIFF 6629 kb

Additional file 3: of The phospholipase DDHD1 as a new target in colorectal cancer therapy

Table S1. Data from SWATH-MS Gene Ontology analysis. (XLSX 740 kb

Additional file 4: of The phospholipase DDHD1 as a new target in colorectal cancer therapy

Figure S2. Effects of DDHD1-expressing cells conditioned medium on DDHD1-silenced cell growth. Cell viability was measured by MTT assay on DDHD1-silenced SW480 cells in the presence of the conditioned medium (CM) of mock cells and DDHD1 overexpressing cells. (TIFF 3275 kb

Additional file 1: of CD90+ liver cancer cells modulate endothelial cell phenotype through the release of exosomes containing H19 lncRNA

(a) Characterization of isolated exosomes. Left panel: DSL for exosomes released by SKHep Middle panel: Western blot forTsg101 and HSC70 in SkHep cells and their relative exosomes. Right panel: Confocal microscopy analysis on HUVECs treated for 1, 3 and 6 hours with 5 mg/ml of SKHep-derived exosomes. HUVECs were stained with phalloidin Alexa Fluor (green), nuclear counterstaining was performed using DAPI (blue), exosomes were labelled with PKH26 (red). (b) Target analysis. Real time-PCR analysis on HUVECs treated for 18 h with 5 mg/ml of SkHep-derived exosomes. Normalized for b-actin the DDct were indicated as fold of induction respect to control (untreated cells). *p<0.05. (c) Tubulogenesis of HUVECs after exosomes treatment. Matrigel assay performed on HUVECs cells after 18 hour of 5 mg/ml SkHep-derived exosomes.  Left panel: phase contrast, magnification 20x. Right panel: quantification of matrigel assay expresses as length of cable as arbitrary unit **p<0.01. (d) Adhesion assay of SkHep cells on HUVECs. Left panel: phase contrast, magnification 10X. Right Panel: quantification of Huh7 or SKHep cells adherent on HUVECs, performed by counting the number of CD90+adherent cells (violet) per field ***p<0.001. (e) Analysis on H19 expression in exosomes. Real time-PCR analysis for H19 expression performed on SkHep-derived exosomes respect to Huh7 derived exosomes. Normalized for b-actin the DDct were indicated as fold of induction. **p<0.01 (f) Comparison of H19 expression in HUVECs after exosomes treatment. Real time-PCR analysis for H19 expression on HUVECs treated for 18 h with 5 mg/ml of SkHep or Huh7 exosomes. Normalized for b-actin the DDct were indicated as fold of induction respect to control (untreated cells). (DOCX 855 kb