Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin inhibits the proliferation of ovarian cancer cells. EdU assays demonstrate that metformin reduces proliferation of (A) SKOV3, (B) A2780 and (C) ES2 cells. Representative fluorescence microscopy images of EdU incorporation and Hoechst 33342 staining of nuclei (magnification, ×100). The proliferation rates of SKOV3, A2780, and ES2 cells were calculated. Data are presented as the mean ± standard deviation. * P<0.05; ** P<0.01; *** P<0.001. Met, metformin; Glu, glucose; EdU, 5-ethynyl-20-deoxyuridine; NC, negative control.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Fluorescence, Microscopy, Staining, Standard Deviation, Negative Control

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin promotes the apoptosis of ovarian cancer cells. Percentage of apoptotic (A) SKOV3, (B) A2780 and (C) ES2 cells were detected by flow cytometry analysis using Annexin V-fluorescein isothiocyanate/PI staining. Apoptotic cells includes Annexin V(-)/PI(+), Annexin V(+)/PI(−) and Annexin V(+)/PI(+) cells. Data are presented the mean ± standard deviation. * P<0.05; ** P<0.01. PI, propidium iodide; Met, metformin; Glu, glucose; NC, negative control.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Flow Cytometry, Staining, Standard Deviation, Negative Control

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin inhibits migration of ovarian cancer cells. Migration of (A) SKOV3, (B) A2780 and (C) ES2 cells were measured by scratch wound healing assay. Representative bright-field images (magnification, ×100) and quantification of recovered distance at 24 h after 10 mM metformin treatment. Data are presented the mean ± standard deviation. * P<0.05; ** P<0.01; *** P<0.001. Met, metformin; Glu, glucose; NC, negative control.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Migration, Wound Healing Assay, Standard Deviation, Negative Control

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin inhibits migration of ovarian cancer cells. Migration of (A) SKOV3, (B) A2780 and (C) ES2 cells were measured by Transwell migration assay. Representative bright-field images of the cell migration (magnification, ×100) and quantification of migrated cells at 24 h after 10 mM metformin treatment are shown. Data are presented the mean ± standard deviation. * P<0.05; ** P<0.01; *** P<0.001. Met, metformin; Glu, glucose; NC, negative control.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Migration, Transwell Migration Assay, Standard Deviation, Negative Control

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin decreases H3K27me3 and expression of polycomb repressor complex 2 in ovarian cancer cells. The protein expression of (A) H3K27me3 and (B) EZH2, SUZ12 and EED in A2780, SKOV3 and ES2 cells were assayed by western blot analysis. Met, metformin; Glu, glucose; H3K27me3, histone H3 lysine 27 trimethylated; H3, histone H3; EZH2, histone-lysine N-methyltransferase EZH2; SUZ12, polycomb protein SUZ12; EED, polycomb protein EED.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Expressing, Western Blot

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin decreases PRC2 of ovarian cancer cells. mRNA expression of EZH2, SUZ12 and EED in (A) SKOV3, (B) A2780 and (C) ES2 cells determined by reverse transcription-quantitative polymerase chain reaction analysis. Data are presented the mean ± standard deviation. * P<0.05; ** P<0.01; *** P<0.001. Met, metformin; Glu, glucose; EZH2, histone-lysine N-methyltransferase EZH2; SUZ12, polycomb protein SUZ12; EED, polycomb protein EED.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Standard Deviation

Journal: International Journal of Oncology

Article Title: Metformin inhibits ovarian cancer via decreasing H3K27 trimethylation

doi: 10.3892/ijo.2018.4343

Figure Lengend Snippet: Metformin inhibits H3K27me3 through the AMPK pathway. (A) Metformin activates AMPK of ovarian cancer cells in normoglycemic condition. (B) 2-DG activates AMPK and suppresses H3K27me3, PRC2 of ovarian cancer cells in normoglycemic condition. (C) Compound C inhibits the effect of metformin on PRC2 and H3K27me3 when ovarian cancer cells were cultured in normoglycemic condition. The protein expression of p-AMPKα, AMPKα, H3K27me3, EZH2, SUZ12 and EED in A2780, SKOV3 and ES2 cells were assayed by western blot analysis. The expression was normalized to β-actin. Met, metformin; Glu, glucose; p-AMPKα, phospho-AMPKα; AMPKα, AMP-activated protein kinase; 2-DG, 2-deoxy-D-glucose; EZH2, histone-lysine N-methyltransferase EZH2;SUZ12, polycomb protein SUZ12; EED, polycomb protein EED; 2-DG, dorsomorphin 2HCl; H3K27me3, histone H3 lysine 27 trimethylation.

Article Snippet: Human epithelial ovarian cancer cell lines SKOV3 (ovarian adenocarcinoma), ES2 (ovarian clear cell carcinoma), and A2780 (ovarian carcinoma) were purchased from China Center for Type Culture Collection (Wuhan University, Wuhan, China) and were cultured in Dulbecco's modified Eagle's medium (DMEM) (HyClone; GE Healthcare Life Sciences, Logan, UT, USA) containing 25 mM glucose (mimicking hyperglycemia) or 5.5 mM glucose (mimicking normoglycemia) supplemented with 10% fetal bovine serum (v/v; Hangzhou Sijiqing Biological Engineering Materials Co., Ltd., Hangzhou, China) at 37°C in 95% air and 5% CO 2 .

Techniques: Cell Culture, Expressing, Western Blot

Journal: British Journal of Cancer

Article Title: ABCB1 (MDR1) induction defines a common resistance mechanism in paclitaxel- and olaparib-resistant ovarian cancer cells

doi: 10.1038/bjc.2016.203

Figure Lengend Snippet: Cross-resistance of paclitaxel- and olaparib-resistant ovarian cancer cell lines. Novel ovarian cancer A2780-derived paclitaxel (A2780pacR) and olaparib (A2780olapR) resistant cell lines were created as described in Materials and Methods section. MTT chemosensitivity assays were used to compare cell viability following exposure to serial dilutions of ( A and D ) paclitaxel and ( B and C ) olaparib in A2780 and A2780pacR ( A and C ) and A2780olapR ( B and D ) cells. Chemosensitivity values obtained from triplicate replicate experiments, expressed as IC 50 values, are summarised in panel E .

Article Snippet: A2780, A2780pacR or A2780olapR cells for immunohistochemical analysis were grown in 10 cm 2 dishes, harvested by scraping, centrifuged (1000 r.p.m., 5 min, RT) and re-suspended in 50 μ l Normal Pooled Plasma (CCN-10, Alpha Laboratories, Eastleigh, UK).

Techniques: Derivative Assay

Journal: British Journal of Cancer

Article Title: ABCB1 (MDR1) induction defines a common resistance mechanism in paclitaxel- and olaparib-resistant ovarian cancer cells

doi: 10.1038/bjc.2016.203

Figure Lengend Snippet: ABCB1 (P-glycoprotein) expression is increased in A2780pacR and A2780olapR cells. ABCB1 ( A ) mRNA expression, ( B ) copy number and ( C and D ) P-glycoprotein expression was compared in A2780, A2780pacR and A2780olapR cells by qRT-PCR analysis, Taqman-based copy number analysis, western blotting and immunohistochemical analysis, respectively, as described in Materials and Methods section.

Article Snippet: A2780, A2780pacR or A2780olapR cells for immunohistochemical analysis were grown in 10 cm 2 dishes, harvested by scraping, centrifuged (1000 r.p.m., 5 min, RT) and re-suspended in 50 μ l Normal Pooled Plasma (CCN-10, Alpha Laboratories, Eastleigh, UK).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Immunohistochemical staining

Journal: British Journal of Cancer

Article Title: ABCB1 (MDR1) induction defines a common resistance mechanism in paclitaxel- and olaparib-resistant ovarian cancer cells

doi: 10.1038/bjc.2016.203

Figure Lengend Snippet: Paclitaxel and olaparib resistance are reversible following combination treatment with the P-glycoprotein inhibitors verapamil and elacridar. MTT chemosensitivity assays were used as described in Materials and Methods section to compare cell viability following exposure to serial dilutions of paclitaxel and olaparib in A2780, A2780pacR and A2780olapR cells in the presence and absence of combination treatments (5 μ M and 10 μ M) with the P-glycoprotein inhibitors ( A – C ) verapamil and ( D – F ) elacridar.

Article Snippet: A2780, A2780pacR or A2780olapR cells for immunohistochemical analysis were grown in 10 cm 2 dishes, harvested by scraping, centrifuged (1000 r.p.m., 5 min, RT) and re-suspended in 50 μ l Normal Pooled Plasma (CCN-10, Alpha Laboratories, Eastleigh, UK).

Techniques:

Journal: British Journal of Cancer

Article Title: ABCB1 (MDR1) induction defines a common resistance mechanism in paclitaxel- and olaparib-resistant ovarian cancer cells

doi: 10.1038/bjc.2016.203

Figure Lengend Snippet: P-glycoprotein-mediated drug efflux is altered in paclitaxel- and olaparib-resistant cells. ( A ) Vybrant Multidrug Resistance Assays were used, as described in Materials and Methods section, to compare P-glycoprotein-dependent efflux of the model substrate Calcein-AM in untreated A2780, A2780pacR and A2780olapR cells and ( B ) in the presence of 0–25 μ M verapamil. Results were confirmed ( C ) using the P-glycoprotein-Glo Assay System, as described in Materials and Methods section, to demonstrate dose-dependent increases in luminescence following the incubation of paclitaxel or olaparib with P-glycoprotein-containing membranes.

Article Snippet: A2780, A2780pacR or A2780olapR cells for immunohistochemical analysis were grown in 10 cm 2 dishes, harvested by scraping, centrifuged (1000 r.p.m., 5 min, RT) and re-suspended in 50 μ l Normal Pooled Plasma (CCN-10, Alpha Laboratories, Eastleigh, UK).

Techniques: Glo Assay, Incubation

Journal: British Journal of Cancer

Article Title: ABCB1 (MDR1) induction defines a common resistance mechanism in paclitaxel- and olaparib-resistant ovarian cancer cells

doi: 10.1038/bjc.2016.203

Figure Lengend Snippet: Paclitaxel- and olaparib-resistant cells are cross-resistant to doxorubicin and rucaparib but not to AZD2461 or veliparib. MTT chemosensitivity assays were used as described in Materials and Methods section to compare cell viability in A2780, A2780pacR and A2780olapR cells following exposure to serial dilutions of ( A and B ) doxorubicin, ( C and D ) AZD2461, ( E and F ) veliparib and ( G and H ) rucaparib. Chemosensitivity values obtained from triplicate replicate experiments, expressed as IC 50 values, are summarised in Panel I .

Article Snippet: A2780, A2780pacR or A2780olapR cells for immunohistochemical analysis were grown in 10 cm 2 dishes, harvested by scraping, centrifuged (1000 r.p.m., 5 min, RT) and re-suspended in 50 μ l Normal Pooled Plasma (CCN-10, Alpha Laboratories, Eastleigh, UK).

Techniques:

Journal: Journal of Translational Medicine

Article Title: Kynurenine in IDO1 high cancer cell-derived extracellular vesicles promotes angiogenesis by inducing endothelial mitophagy in ovarian cancer

doi: 10.1186/s12967-024-05054-5

Figure Lengend Snippet: OC cell-derived EVs induced endothelial mitophagy related to tumor angiogenesis and progression in vivo. a Schematic illustration showing EVs isolation and purification process from the culture medium of normal ovarian epithelial cells (IOSE80) and ovarian cancer (OC) cells (A2780, SKOV3). b Representative images of EVs obtained from IOSE80 and A2780 cells by transmission electron microscope (TEM). Scale bar:100 nm. c Diameter ranges of EVs produced from IOSE80 and A2780 cells by NanoFCM. d Schematic diagram of orthotopic OC murine model educated with IOSE80-EVs or A2780-EVs (n = 5). e Representative images of the primary ovary tumor size in orthotopic OC model education with IOSE80-EVs or A2780-EVs. bar = 1 cm. (n = 5, Mean ± SD, t-test, *** P < 0.001). f Numbers of peritoneal metastases validated by HE staining in orthotopic OC model education with IOSE80-EVs or A2780-EVs. Scale bar:50 μm. (n = 5, Mean ± SD, t-test, * P < 0.05). g Immunohistochemistry (IHC) analysis of CD31 expression in primary tumor tissues of the orthotopic OC model education with IOSE80-EVs or A2780-EVs.Scale bar:50 μm. (n = 3, Mean ± SD, t-test, * P < 0.05). h Representative images of LC3, TOMM20and CD31 in primary tumor tissue sections of orthotopic OC model educated with IOSE80-EVs or A2780-EVs by immunofluorescence staining. Scale bar:20 μm

Article Snippet: Scale bar:100 nm. c Diameter ranges of EVs produced from IOSE80 and A2780 cells by NanoFCM. d Schematic diagram of orthotopic OC murine model educated with IOSE80-EVs or A2780-EVs (n = 5). e Representative images of the primary ovary tumor size in orthotopic OC model education with IOSE80-EVs or A2780-EVs. bar = 1 cm. (n = 5, Mean ± SD, t-test, *** P < 0.001). f Numbers of peritoneal metastases validated by HE staining in orthotopic OC model education with IOSE80-EVs or A2780-EVs.

Techniques: Derivative Assay, In Vivo, Isolation, Purification, Transmission Assay, Microscopy, Produced, Staining, Immunohistochemistry, Expressing, Immunofluorescence

Journal: Journal of Translational Medicine

Article Title: Kynurenine in IDO1 high cancer cell-derived extracellular vesicles promotes angiogenesis by inducing endothelial mitophagy in ovarian cancer

doi: 10.1186/s12967-024-05054-5

Figure Lengend Snippet: Enrichment of L-kyn in EVs acquired from OC patients and OC cells. a Representative images of EVs obtained from plasma with non-OC or OC patients by TEM. Scale bar: 100 nm. b Diameter ranges of EVs acquired from plasma with non-OC or OC patients by NanoFCM. c Principal component analysis (PCA) of total metabolite profiles with positive ion mode. d Pie chart depicting metabolite classification in positive ion mode. e Matchstick diagram showing top20 significant fold change of differential metabolites with positive ion mode between OC and non-OC plasma derived-EVs. f KEGG pathway enrichment analysis based on differentially accumulated metabolites with positive ion mode between OC and non-OC plasma derived-EVs. g Violin diagram showing level of L-kyn in plasma and plasma derived-EVs of non-OC and OC patients determined by ELISA (n = 40, two-way ANOVA, * P < 0.05,*** P < 0.001). h Level of L-kyn in tissue supernatant and tissue derived-EVs of OC tissues and the corresponding para cancer tissues (n = 4, Mean ± SD, two-way ANOVA, *** P < 0.001). i Level of L-kyn in cell culture supernatant and cell derived-EVs of normal ovarian epithelial cells (IOSE80) and ovarian cancer cells (A2780) (n = 5, Mean ± SD, two-way ANOVA, *** P < 0.001). j Level of L-kyn in the endothelial cells treated with EVs obtained from normal ovarian epithelial cells (IOSE80) and ovarian cancer cells (A2780) (n = 5, Mean ± SD, t-test, *** P < 0.001). For c – f , all n = 6 plasma-derived EVs from patients with OC or non-OC were performed for analysis

Article Snippet: Scale bar:100 nm. c Diameter ranges of EVs produced from IOSE80 and A2780 cells by NanoFCM. d Schematic diagram of orthotopic OC murine model educated with IOSE80-EVs or A2780-EVs (n = 5). e Representative images of the primary ovary tumor size in orthotopic OC model education with IOSE80-EVs or A2780-EVs. bar = 1 cm. (n = 5, Mean ± SD, t-test, *** P < 0.001). f Numbers of peritoneal metastases validated by HE staining in orthotopic OC model education with IOSE80-EVs or A2780-EVs.

Techniques: Clinical Proteomics, Derivative Assay, Enzyme-linked Immunosorbent Assay, Cell Culture

Journal: Journal of Translational Medicine

Article Title: Kynurenine in IDO1 high cancer cell-derived extracellular vesicles promotes angiogenesis by inducing endothelial mitophagy in ovarian cancer

doi: 10.1186/s12967-024-05054-5

Figure Lengend Snippet: Aberrant IDO1 upregulation involved in tryptophan metabolic reprogramming of OC cells. a Dot plot showing IDO1 expression of OC and non-OC samples. b Visualization of IDO1 expression in major cell clusters of OC and non-OC ovarian samples. c Visualization of IDO1 expression in epithelial cell subclusters with OC and non-OC ovarian samples. d Heatmap showing IDO1 expression in OC samples from OCGA_OV dataset and non-OC samples from GETx dataset. e Heatmap showing IDO1 expression in OC samples from GSE140082 dataset and non-OC samples from GETx dataset. f Immunohistochemistry analysis of IDO1 protein expression between OC and non-OC ovarian tissues from the human protein atlas (HPA)database. Scale bar:200 μm. g Immunohistochemistry detecting of IDO1 protein expression in tissue microarray composed of OC (n = 115) and non-OC (n = 80) ovarian tissues from our hospital. Scale bar:200 μm. (u-test, *** P < 0.001). h IDO1 mRNA expression between non-OC cells (IOSE80) and OC cells (A2780, SKOV3) was detected by qRT-PCR (n = 3, Mean ± SD, t-test, ** P < 0.01, *** P < 0.001). i IDO1 protein expression between non-OC cells (IOSE80) and OC cells (A2780, SKOV3) was detected by Western blot (n = 3, Mean ± SD, t-test, *** P < 0.001). For a – c , all n = 7 OC and n = 5 non-OC samples obtained from GSE184880 dataset were used for analysis

Article Snippet: Scale bar:100 nm. c Diameter ranges of EVs produced from IOSE80 and A2780 cells by NanoFCM. d Schematic diagram of orthotopic OC murine model educated with IOSE80-EVs or A2780-EVs (n = 5). e Representative images of the primary ovary tumor size in orthotopic OC model education with IOSE80-EVs or A2780-EVs. bar = 1 cm. (n = 5, Mean ± SD, t-test, *** P < 0.001). f Numbers of peritoneal metastases validated by HE staining in orthotopic OC model education with IOSE80-EVs or A2780-EVs.

Techniques: Expressing, Immunohistochemistry, Microarray, Quantitative RT-PCR, Western Blot