Journal: International Journal of Molecular Sciences

Article Title: Reactivation of P53 Antiproliferative and Pro-Apoptotic Pathways by Resveratrol in Mutant P53 Cancer Cell Lines

doi: 10.3390/ijms27104481

Figure Lengend Snippet: Dose-dependent effect of resveratrol (RES) on cellular metabolic activity in cancer cell lines with different p53 status. Cells were treated with increasing concentrations of RES (50–250 µM) for 48 h. Cellular metabolic activity was assessed by means of MTT assay and normalized to that of vehicle-treated controls (EtOH 0.3% v / v ). Data represent mean ± SD of at least four independent experiments. IC50 values are indicated for each cell line (presented in box).

Article Snippet: The human cervical cancer cell line C33A ( p53 mutation R273C) was purchased from the American Type Culture Collection (ATCC cat. no. HTB-31 [C-33A]; Manassas, VA, USA).

Techniques: Activity Assay, MTT Assay

Journal: International Journal of Molecular Sciences

Article Title: Reactivation of P53 Antiproliferative and Pro-Apoptotic Pathways by Resveratrol in Mutant P53 Cancer Cell Lines

doi: 10.3390/ijms27104481

Figure Lengend Snippet: Resveratrol induces apoptosis in cancer cell lines with mutant or null p53 . ( A ) C33A, SK-BR-3, and Saos-2 cells were treated with vehicle (ethanol) or resveratrol at their respective IC50s for 48 h. Apoptosis was assessed by means of Annexin V/propidium iodide (PI) double staining followed by flow cytometry analysis. Representative dot plots show viable cells in the lower left quadrant (Annexin V − /PI − ), early apoptotic cells in the lower right quadrant (Annexin V + /PI − ), and late apoptotic cells in the upper right quadrant (Annexin V + /PI + ). ( B ) Quantifications of total apoptotic cells (early + late apoptosis) are shown as the mean ± SD from three independent experiments. ** p < 0.01; *** p < 0.001 versus EtOH. RES, resveratrol; EtOH, control with ethanol.

Article Snippet: The human cervical cancer cell line C33A ( p53 mutation R273C) was purchased from the American Type Culture Collection (ATCC cat. no. HTB-31 [C-33A]; Manassas, VA, USA).

Techniques: Mutagenesis, Double Staining, Flow Cytometry, Control

Journal: International Journal of Molecular Sciences

Article Title: Reactivation of P53 Antiproliferative and Pro-Apoptotic Pathways by Resveratrol in Mutant P53 Cancer Cell Lines

doi: 10.3390/ijms27104481

Figure Lengend Snippet: Resveratrol induces the expression of p53 target genes across different p53 backgrounds. Gene expression analysis was performed in ( A ) C33A ( p53 R273C), ( B ) SK-BR-3 ( p53 R175H), ( C ) MCF-7 ( p53 WT), and ( D ) Saos-2 ( p53 null) cells following treatment with vehicle (EtOH) or resveratrol (RES) at their respective IC50s for 48 h. Relative mRNA levels of p53 target genes were determined by means of RT-qPCR and expressed as fold changes relative to vehicle-treated controls, normalized to the housekeeping gene β2-microglobulin (β2M). Data are presented as the mean ± SD of at least three independent experiments. Statistical significance was evaluated using one-way ANOVA followed by appropriate post hoc multiple comparison tests. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001 versus control.

Article Snippet: The human cervical cancer cell line C33A ( p53 mutation R273C) was purchased from the American Type Culture Collection (ATCC cat. no. HTB-31 [C-33A]; Manassas, VA, USA).

Techniques: Expressing, Gene Expression, Quantitative RT-PCR, Comparison, Control

Journal: International Journal of Molecular Sciences

Article Title: Reactivation of P53 Antiproliferative and Pro-Apoptotic Pathways by Resveratrol in Mutant P53 Cancer Cell Lines

doi: 10.3390/ijms27104481

Figure Lengend Snippet: Resveratrol enhances cisplatin sensitivity in cancer cell lines with mutant or null p53 . ( A ) C33A, ( B ) SK-BR-3, and ( C ) Saos-2 cells were treated with increasing concentrations of cisplatin (CDDP; 2.5–20 μM) in the presence or absence of resveratrol (RES), at the corresponding IC 50 for each cell line, for 48 h. Cellular metabolic activity was assessed via MTT assay and normalized to that of vehicle-treated controls. Data represent the mean ± SD of at least four independent experiments. ( D ) IC50 values for CDDP were calculated using non-linear regression analysis (log [CDDP] vs. normalized response, variable slope model) using GraphPad Prism 6.01. * Previously published results from our working group.

Article Snippet: The human cervical cancer cell line C33A ( p53 mutation R273C) was purchased from the American Type Culture Collection (ATCC cat. no. HTB-31 [C-33A]; Manassas, VA, USA).

Techniques: Mutagenesis, Activity Assay, MTT Assay

Journal: Current Issues in Molecular Biology

Article Title: The Therapeutic Effect of EZH2 Inhibitors in Targeting Human Papillomavirus Associated Cervical Cancer

doi: 10.3390/cimb47120990

Figure Lengend Snippet: MTT assay results for cytotoxicity of drug treatments. Drug dose–response curves after 48 h for HPV- C33a, HPV+ CaSki, and normal epithelial cell line CRL1790: ( A ) cisplatin, ( B ) EPZ6438, and ( C ) ZLD1039. Data is expressed as log of mean ± SD (n = 3). IC50 values, pinpointed on the graph, were calculated using GraphPad prism. ( D ) Table of mean IC50 value data (n = 3). The IC50 value for EPZ6438 could not be determined within the 0–80 µM range.

Article Snippet: According to human protein atlas [ ], the C33a cell line exhibits no expression of CDH1 gene, which encodes E-cadherin; however, mass spectrometry proteomics data could further confirm its protein expression.

Techniques: MTT Assay

Journal: Current Issues in Molecular Biology

Article Title: The Therapeutic Effect of EZH2 Inhibitors in Targeting Human Papillomavirus Associated Cervical Cancer

doi: 10.3390/cimb47120990

Figure Lengend Snippet: Migration rate assessment following the scratch wound–healing assay. ( A ) Representative images from the in vitro scratch wound–healing assays demonstrating cell migration into the cell-free region (outlined by black lines) following 48 h treatment (scale = 100 µm). ( B ) Summary plots showing the migration rates by C33a and CaSki cells after treatment (Mean ± SD, n = 3).

Article Snippet: According to human protein atlas [ ], the C33a cell line exhibits no expression of CDH1 gene, which encodes E-cadherin; however, mass spectrometry proteomics data could further confirm its protein expression.

Techniques: Migration, Wound Healing Assay, In Vitro

Journal: Current Issues in Molecular Biology

Article Title: The Therapeutic Effect of EZH2 Inhibitors in Targeting Human Papillomavirus Associated Cervical Cancer

doi: 10.3390/cimb47120990

Figure Lengend Snippet: Effect of EZH2 inhibitors on protein and mRNA expression of ZO-1 and E-cadherin on cervical cancer cells after 48 h treatment. ( A ) Immunofluorescence staining of ZO-1 and E-cadherin protein expression (pink) and nuclei (blue) under 400× microscope magnification. Scale = 50 µm. Mean fluorescence intensity of immunocytochemistry results were normalised to control cells (right) (n = 3). ( B ) Representative Western blots’ results of protein bands ZO-1 (250 kDa) and β-actin (42 kDa) for C33a and CaSki and E-cadherin (110 kDa) for CaSki are shown on the left. Bar charts on the right show the Western blot quantification of these protein expression levels following treatment. Values are expressed as a fold change of control (n = 3). ( C ) RT-qPCR analysis of relative ZO-1 (C33a and CaSki) and E-cadherin (CaSki) mRNA expression levels following 48 h treatment. Values are expressed as fold change of control values. Data is presented as mean ± SD (n = 3). * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001. Bold asterisk above a single sample bar indicates the significant difference of that sample from the rest of the treatments.

Article Snippet: According to human protein atlas [ ], the C33a cell line exhibits no expression of CDH1 gene, which encodes E-cadherin; however, mass spectrometry proteomics data could further confirm its protein expression.

Techniques: Expressing, Immunofluorescence, Staining, Microscopy, Fluorescence, Immunocytochemistry, Control, Western Blot, Quantitative RT-PCR

Journal: Current Issues in Molecular Biology

Article Title: The Therapeutic Effect of EZH2 Inhibitors in Targeting Human Papillomavirus Associated Cervical Cancer

doi: 10.3390/cimb47120990

Figure Lengend Snippet: Immunohistochemical staining results of EZH2, p53, ZO-1, β-catenin, and HPV16 E6 from CAM assay tissue sections after 48 h treatment with control (DMSO 0.1%), cisplatin, and EPZ6438. Immunohistochemical staining of ( A ) C33a and ( B ) CaSki for expression of EZH2, p53, ZO-1, β-catenin, and HPV16 E6 from the CAM tissue model (n = 1). Scale = 50 µm. Chorionic (CE) and allantoic (AE) epithelial layers with sub-epithelial capillary network (SEC), tumour cells (TC), mesoderm (M), and blood vessels (BV) are displayed.

Article Snippet: According to human protein atlas [ ], the C33a cell line exhibits no expression of CDH1 gene, which encodes E-cadherin; however, mass spectrometry proteomics data could further confirm its protein expression.

Techniques: Immunohistochemical staining, Staining, Chick Chorioallantoic Membrane Assay, Control, Expressing