Journal: Stem cell research & therapy

Article Title: Exosomes from glioma cells induce a tumor-like phenotype in mesenchymal stem cells by activating glycolysis.

doi: 10.1186/s13287-019-1149-5

Figure Lengend Snippet: Fig. 2 U251 cell-exosomes promote the proliferation of hBMSCs. a Cellular internalization of Dil-labeled U251 cell-derived exosomes into hBMSCs. b hBMSCs were treated with different concentrations of U251 cell-derived exosomes for different time points (24, 48, and 72 h), and then cell viability was measured using the CCK-8 analyses. The results showed U251 cell-derived exosomes could promote hBMSC proliferation in a time- and dose-dependent manner with respect to control cells (*P < 0.05). c Representative images of Ki67 immunostaining in hBMSCs treated with or without U251 cell- derived exosomes for 48 h. Green fluorescence indicated the Ki67- positive cells. d Protein expression of PCNA and C-myc was determined by Western blotting analysis in hBMSCs treated U251 cell-derived exosomes for 48 h. e Bar graphs showing the relative protein expression level of PCNA and C-myc in hBMSCs treated with U251 cell-derived exosomes for 48 h. The protein levels of GAPDH served as loading controls (*P < 0.05). f The hBMSCs exposed with exosomes for 48 h showed an increase in the S phase of the cell cycle compared to the controls. g Treatment of exosomes caused a S cell cycle arrest in a dose-dependent manner, the arresting rate increased more rapid in hBMSCs exposed with exosomes than control cells for 48 h (*P < 0.05, **P < 0.01). h Real-time PCR analysis of P21 and P16 mRNA expression in hBMSCs treated with U251 cell- derived exosomes for 48 h (*P < 0.05, **P < 0.01)

Article Snippet: Antibodies against HSP70, CD9, MMP-2, MMP-9, PCNA, C-myc, and GAPDH were obtained from Proteintech Group Inc. (Chicago, IL, USA).

Techniques: Labeling, Derivative Assay, CCK-8 Assay, Control, Immunostaining, Fluorescence, Expressing, Western Blot, Real-time Polymerase Chain Reaction

Journal: Stem cell research & therapy

Article Title: Exosomes from glioma cells induce a tumor-like phenotype in mesenchymal stem cells by activating glycolysis.

doi: 10.1186/s13287-019-1149-5

Figure Lengend Snippet: Fig. 3 U251 cell-derived exosomes contribute to the promotion of the migration and invasion of hBMSCs. a The movement of hBMSCs was observed by wound healing assay at 0 h and 48 h after exosomes treatment. b The wound closure rate of hBMSCs was quantified by measuring wound closure areas at 0 h and 48 h (*P < 0.05, **P < 0.01). c The migration and invasion abilities of hBMSCs were examined by Transwell assay after treatment with U251 cell- derived exosomes for 48 h. The photos represented cell invasion into the underside of the Transwell membrane under a microscope at × 200 magnification field. d The hBMSC number that crossed the Transwell chamber of each group was counted and averaged after treatment with U251 cell-derived exosomes for 48 h (*P < 0.05, **P < 0.01). e hBMSCs were treated with 300 μg/mL U251 cell- derived exosomes for 48 h. MMP-2 and MMP-9 proteins in hBMSCs were detected by Western blotting. The ratios of MMP-2 and MMP-9 were relative to GAPDH and standardized to the control group (*P < 0.05). f The mRNA expression levels of MMP- 2 and MMP-9 in hBMSCs treated with U251 cell-derived exosomes for 48 h (*P < 0.05)

Article Snippet: Antibodies against HSP70, CD9, MMP-2, MMP-9, PCNA, C-myc, and GAPDH were obtained from Proteintech Group Inc. (Chicago, IL, USA).

Techniques: Derivative Assay, Migration, Wound Healing Assay, Transwell Assay, Membrane, Microscopy, Western Blot, Control, Expressing

Journal: Stem cell research & therapy

Article Title: Exosomes from glioma cells induce a tumor-like phenotype in mesenchymal stem cells by activating glycolysis.

doi: 10.1186/s13287-019-1149-5

Figure Lengend Snippet: Fig. 11 2-DG reversed the tumor-like phenotype transformation of hBMSCs under the treatment of U251 cell-derived exosomes. The hBMSCs were incubated with 300 μg/mL U251 cell-derived exosomes or 2.5 mmol/L 2-DG for 48 h. a 2-DG could suppress the proliferation of hBMSCs under the treatment of U251 cell- derived exosomes for 24 h, 48 h, and 72 h (*P < 0.05). b, c The effects of 2-DG on hBMSCs migration capability were assessed by the Transwell assay with or without Matrigel under the treatment of U251 cell-derived exosomes. The number of migrated or invaded cells was quantified by counting the number of cells from five random fields at a × 200 magnification (*P < 0.05). d, e Protein levels of MMP-2 and MMP-9 were examined by Western blotting under the treatment of U251 cell- derived exosome or 2.5 mmol/L 2-DG. GAPDH was an internal control (*P < 0.05)

Article Snippet: Antibodies against HSP70, CD9, MMP-2, MMP-9, PCNA, C-myc, and GAPDH were obtained from Proteintech Group Inc. (Chicago, IL, USA).

Techniques: Transformation Assay, Derivative Assay, Incubation, Migration, Transwell Assay, Western Blot, Control

Journal: Stem cell research & therapy

Article Title: Exosomes from glioma cells induce a tumor-like phenotype in mesenchymal stem cells by activating glycolysis.

doi: 10.1186/s13287-019-1149-5

Figure Lengend Snippet: Fig. 12 2-DG reverses the tumor-associated factors of hBMSCs under the treatment of U251 cell-derived exosomes. The hBMSCs were incubated with 300 μg/mL U251 cell-derived exosomes or 2.5 mmol/L 2-DG for 48 h. a The protein levels of PCNA, C-myc, CD133, Nestin, and GFAP were examined by Western blotting. b–d Bar graphs showing the relative protein expression level of PCNA, C-myc, CD133, Nestin, and GFAP. The protein levels of GAPDH served as loading controls (*P < 0.05)

Article Snippet: Antibodies against HSP70, CD9, MMP-2, MMP-9, PCNA, C-myc, and GAPDH were obtained from Proteintech Group Inc. (Chicago, IL, USA).

Techniques: Derivative Assay, Incubation, Western Blot, Expressing

Journal: American Journal of Cancer Research

Article Title: Repression of GCN5 expression or activity attenuates c-MYC expression in non-small cell lung cancer

doi:

Figure Lengend Snippet: GCN5 and c-MYC proteins are upregulated and positively associated in NSCLC cell lines. Steady state levels of GCN5, c-MYC, and ADA2B proteins were analyzed in (A) murine and (B) human immortalized epithelial and fibroblast lung controls and NSCLC cell lines. Immunoblotting was performed using anti-GCN5, anti-c-MYC, and anti-ADA2B antibodies. β-Actin served as a loading control. GCN5 and c-MYC expression levels were normalized to β-Actin expression and compared between immortalized lung controls and NSCLC cell lines. Steady state levels of histone H3 K9 acetylation were detected by immunoblotting using anti-H3K9 acetylation and anti-H3 antibodies in (C) murine and (D) human cell lines. Total H3 protein served as a loading control. Stable reduction of endogenous c-MYC expression by independent shRNAs introduced into murine ED1 (E) and human H1355 (F) lung cancer cells decreased endogenous GCN5 protein levels. Immunoblotting was performed using anti-GCN5 and anti-c-MYC antibodies. β-Actin served as a loading control.

Article Snippet: Antibodies used for immunoblot analyses were: anti-ADA2B (Abcam, #57953), anti-β-Actin (Santa Cruz, #sc47778), anti-c-Myc (Cell Signaling Technology, #5605 and #9402), anti-GCN5 (Cell Signaling Technology, #3305), anti-H3 (Abcam, #1791), anti-H3-K9Ac (Abcam, #32129), and anti-PCAF (Cell Signaling Technology, #3378).

Techniques: Western Blot, Expressing

Journal: American Journal of Cancer Research

Article Title: Repression of GCN5 expression or activity attenuates c-MYC expression in non-small cell lung cancer

doi:

Figure Lengend Snippet: Reduction of GCN5 protein represses cell proliferation, increases the percentage of necrotic cells, and decreases c-MYC protein expression. A. Proliferation of murine ED1 and human H1355 NSCLC cell lines with stable knockdown of GCN5 by two independent shRNAs was monitored over 4 days. Proliferation rate was normalized to day 1. Representative immunoblots confirming GCN5 knockdown are shown in panel 3C. B. Apoptosis assays were performed in the murine ED1 NSCLC cell line with stable knockdown of GCN5 by shRNAs 3 days after seeding cells. Percentage of live, apoptotic and necrotic cells are shown. C. Stable reduction of endogenous GCN5 expression by two independent shRNAs introduced into murine and human lung cancer cells decreased endogenous c-MYC protein. Immunoblotting was performed using anti-GCN5 and anti-c-MYC antibodies. β-Actin served as a loading control. Symbols refer to *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.

Article Snippet: Antibodies used for immunoblot analyses were: anti-ADA2B (Abcam, #57953), anti-β-Actin (Santa Cruz, #sc47778), anti-c-Myc (Cell Signaling Technology, #5605 and #9402), anti-GCN5 (Cell Signaling Technology, #3305), anti-H3 (Abcam, #1791), anti-H3-K9Ac (Abcam, #32129), and anti-PCAF (Cell Signaling Technology, #3378).

Techniques: Expressing, Western Blot