Journal: Oncotarget

Article Title: Prostate tumor-induced angiogenesis is blocked by exosomes derived from menstrual stem cells through the inhibition of reactive oxygen species

doi: 10.18632/oncotarget.9852

Figure Lengend Snippet: A. PC3 cells were treated with or without exosomes or lysed exosomes and assayed for ROS production by FACS using the fluorescent dye DCFDA. NAC and H 2 O 2 were used as negative and positive control, respectively. Representative histograms show ROS generation in each experimental condition. Graph represents the mean values ± SE of the MFI of DCF. *, respect to PC3. B-C. HUVEC were resuspended in the CM derived from PC3 cells treated with exosomes, exosomes plus H 2 O 2 , lysed exosomes, NAC, or untreated, and seeded in matrigel (B); Representative images show tube formation after 5 hours. Graphs represent the quantitative analysis of the angiogenic potential (C). D-G. A panel of prostate cancer cell lines were incubated in the absence or presence of MenSCs-derived exosomes for 36 hours and their effect on VEGF , tube-like structures formation and ROS was determined. Relative expression levels of VEGF was assessed by qRT-PCR (D); Protein level of VEGF was determined by western-blot (E); Representative images show tube formation after 5 hours. Graphs represent the quantitative analysis of the angiogenic potential (F); ROS generation by FACS using the fluorescent dye DCFDA (G). Data are presented as mean values ± SE. Western blotting results were evaluated by densitometry, corrected with respect to β-actin expression, and expressed relative to the value obtained with the corresponding control (arbitrarily set as 1). Equal protein loading was assessed by anti–β-actin immunoblotting. Scale bar: 200 μm. *, respect to PC3-CM EXO ; √, respect to PC3-CM EXO +H 2 O 2 ; +, respect to PC3-CM+NAC. Abbreviations: MenSCs, menstrual derived mesenchymal stem cells; EXO, exosomes; CM, conditioned medium; LysEXO, lysed exosomes; DMEM, Dulbecco's modified Eagle's medium; EGM, endothelial growth medium; NAC, N-acetylcysteine; ROS, reactive oxygen species; DCFDA, 2′, 7′–dichlorofluorescin diacetate; DCF, 2′, 7′–dichlorofluorescin; FACS, fluorescence-activated cell sorting; MFI, mean fluorescent intensity; SE, standard error.

Article Snippet: Internalization of exosomes in PC3 cells was determined by fluorescence-activated cell sorting (FACS) (FACS Canto II, BD Biosciences, USA) and visualized with confocal microscopy (FV1200 Laser Scanning Microscope in combination with an IX83 automated inverted microscope platform, Olympus).

Techniques: Positive Control, Derivative Assay, Incubation, Expressing, Quantitative RT-PCR, Western Blot, Modification, Fluorescence, FACS

Journal: Oncotarget

Article Title: Prostate tumor-induced angiogenesis is blocked by exosomes derived from menstrual stem cells through the inhibition of reactive oxygen species

doi: 10.18632/oncotarget.9852

Figure Lengend Snippet: A. HUVECs were resuspended in CM from PC3 cells treated with BMSC-derived exosomes or untreated, and seeded in matrigel. Representative images show tube formation after 5 hours. Graphs represent quantitative analysis of the angiogenic potential. Data are presented as mean values ± SE. B. PC3 cells were treated with or without BMSC-derived exosomes or lysed exosomes, and assayed for ROS production by FACS using the fluorescent dye DCFDA. Representative histograms show endogenous ROS production in each experimental condition. Data are presented as mean values ± SE. Scale bar: 200 μm. Abbreviations: BMSCs, bone marrow derived mesenchymal stem cells; EXO, exosomes; CM, conditioned medium; DMEM, Dulbecco's modified Eagle's medium; EGM, endothelial growth medium; ROS, reactive oxygen species; NAC, N-acetylcysteine; DCFDA, 2′, 7′–dichlorofluorescein diacetate; DCF, 2′, 7′–dichlorofluorescein; FACS, fluorescence-activated cell sorting; SE, standard error.

Article Snippet: Internalization of exosomes in PC3 cells was determined by fluorescence-activated cell sorting (FACS) (FACS Canto II, BD Biosciences, USA) and visualized with confocal microscopy (FV1200 Laser Scanning Microscope in combination with an IX83 automated inverted microscope platform, Olympus).

Techniques: Derivative Assay, Modification, Fluorescence, FACS

Journal: Oncotarget

Article Title: Prostate tumor-induced angiogenesis is blocked by exosomes derived from menstrual stem cells through the inhibition of reactive oxygen species

doi: 10.18632/oncotarget.9852

Figure Lengend Snippet: A-B. Breast (MDA-MB-231 and MCF-7) and pancreatic (MIA PaCa-2 and PANC-1) cancer cell lines were incubated in the absence or presence of MenSCs-derived exosomes for 36 hours and their effects on VEGF were determined. Relative expression levels of VEGF was assessed by qRT-PCR (A) and western-blot (B). C. HUVECs were resuspended in CM from tumor cells treated with MenSCs-derived exosomes or untreated, and seeded in matrigel. Representative images show tube formation after 5 hours. Graphs represent quantitative analysis of the angiogenic potential. Data are presented as mean values ± SE. Western blot results were evaluated by densitometry, corrected with respect to β-actin expression, and expressed relative to the value obtained with the corresponding control (arbitrarily set as 1). Equal protein loading was assessed by anti–β-actin immunoblotting. Scale bar: 200 μm. D-E. Cancer cells were treated with or without MenSCs-derived exosomes and assayed for ROS production by FACS using the fluorescent dye DCFDA. Representative histograms show endogenous ROS production in each experimental condition. Abbreviations: EXO, exosomes; CM, conditioned medium; DMEM, Dulbecco's modified Eagle's medium; EGM, endothelial growth medium; ROS, reactive oxygen species; DCFDA, 2′, 7′–dichlorofluorescein diacetate; DCF, 2′, 7′–dichlorofluorescein; FACS, fluorescence-activated cell sorting; ns, non-significant; SE, standard error.

Article Snippet: Internalization of exosomes in PC3 cells was determined by fluorescence-activated cell sorting (FACS) (FACS Canto II, BD Biosciences, USA) and visualized with confocal microscopy (FV1200 Laser Scanning Microscope in combination with an IX83 automated inverted microscope platform, Olympus).

Techniques: Incubation, Derivative Assay, Expressing, Quantitative RT-PCR, Western Blot, Modification, Fluorescence, FACS