Journal: Heliyon

Article Title: Calotropis gigantea stem bark extract activates HepG2 cell apoptosis through ROS and its effect on cytochrome P450

doi: 10.1016/j.heliyon.2023.e16375

Figure Lengend Snippet: Apoptotic induction in HepG2 cells treated with CGDCM at 200, 400, and 800 μg/mL and 0.8% DMSO as the vehicle control for 24 h. (A) Flow cytometry detection of apoptosis using double staining with annexin-V and PI. (B) The percentage of apoptotic cells is presented in the histogram. (C) Western blot image of the expression of Bcl-2 and (D) the relative expression level of Bcl-2/β-actin. The original uncropped western blots of Bcl-2 and β-actin are presented in the Supplementary Fig. S1. (E) The expression of cleaved caspase-3 was measured by immunofluorescence with Hoechst 33342-stained nuclei. (F) MTT analysis of IMR-90 cell viability after CGDCM treatment. Data are presented as the mean ± SD from at least three replicates, n = 3, and statistics were analyzed using one-way ANOVA with Tukey's HSD test. *p < 0.05 relative to the vehicle.

Article Snippet: Next, the membranes were incubated overnight at 4 °C with primary antibodies, including rabbit polyclonal β-cell lymphoma 2 (Bcl-2) (PA1-30411, Thermo Fisher Scientific, USA), rabbit polyclonal SOD2/MnSOD (AF5144, Affinity bioscience, USA), rabbit polyclonal catalase (DF7545, Affinity bioscience, USA), and mouse monoclonal β-actin (8H10D10, Cell Signaling Technology, USA).

Techniques: Flow Cytometry, Double Staining, Western Blot, Expressing, Immunofluorescence, Staining

Journal: Heliyon

Article Title: Calotropis gigantea stem bark extract activates HepG2 cell apoptosis through ROS and its effect on cytochrome P450

doi: 10.1016/j.heliyon.2023.e16375

Figure Lengend Snippet: The effect of CGDCM on the percentage of (A) fatty acids, (B) ATP, and (C) ROS levels in HepG2 cells. Cells were treated for 24 h with 200, 400, and 800 μg/mL CGDCM and 0.8% DMSO as the vehicle control. (D) ROS were analyzed by fluorescence staining with H2DCFDA and then observed with a fluorescence microscope, Bars = 100 μm. (E) SOD2 and catalase expression determined by western blotting. (F) The relative expression levels of SOD2/β-actin and (G) catalase/β-actin. The original uncropped western blots of SOD2, catalase, and β-actin are presented in the Supplementary Fig. S2. Data are presented as the mean ± SD from at least three replicates, n = 3, and statistics were analyzed with one-way ANOVA with Tukey's HSD test. *p < 0.05 vs. the vehicle.

Article Snippet: Next, the membranes were incubated overnight at 4 °C with primary antibodies, including rabbit polyclonal β-cell lymphoma 2 (Bcl-2) (PA1-30411, Thermo Fisher Scientific, USA), rabbit polyclonal SOD2/MnSOD (AF5144, Affinity bioscience, USA), rabbit polyclonal catalase (DF7545, Affinity bioscience, USA), and mouse monoclonal β-actin (8H10D10, Cell Signaling Technology, USA).

Techniques: Fluorescence, Staining, Microscopy, Expressing, Western Blot

Journal: Pharmaceuticals

Article Title: Resveratrol Enhances Cytotoxic Effects of Cisplatin by Inducing Cell Cycle Arrest and Apoptosis in Ovarian Adenocarcinoma SKOV-3 Cells through Activating the p38 MAPK and Suppressing AKT

doi: 10.3390/ph16050755

Figure Lengend Snippet: RES enhances cisplatin-induced SKOV-3 apoptosis. SKOV-3 cells were treated with RES (25, 50 and 100 μM) alone or in combination with cisplatin (20 μM) for 24 h or 48 h. ( A ) Annexin V/PI double staining and flow cytometry were performed for apoptosis analysis. ( B ) The right graphs show the quantification (%) of total, early and late apoptosis according to treatment at each time point; values represent mean ± SD of three independent experiments. # p < 0.05 vs. cisplatin-treated cells. ( C ) Western blot analysis, detecting caspase-9, caspase-3, PARP, and their cleaved forms, was performed as described in the ‘Materials and Methods’; β-actin was used to normalize protein expression. ( D ) Protein quantification via densitometry; values represent mean ± SD of three independent experiments. * p < 0.05 vs. untreated cells; # p < 0.05 vs. cisplatin-treated cells.

Article Snippet: Rabbit anti-phospho p38 antibody, rabbit anti p38 antibody, rabbit anti-phospho ERK1/2 antibody, mouse anti-ERK1/2 antibody, rabbit anti-phospho JNK antibody, rabbit anti-JNK antibody, rabbit anti-phospho AKT (Ser473) antibody, mouse anti-AKT antibody, rabbit anti-cyclin A2 antibody, rabbit anti-cyclin B1 antibody, rabbit anti-cyclin E1 antibody, rabbit anti-caspase-9 antibody, rabbit anti-caspase-3 antibody, rabbit anti-PARP antibody, mouse anti-β-actin antibody, and DAPI (4, 6-diamidino-2-phenylindole, dihydrochloride) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Double Staining, Flow Cytometry, Western Blot, Expressing

Journal: Pharmaceuticals

Article Title: Resveratrol Enhances Cytotoxic Effects of Cisplatin by Inducing Cell Cycle Arrest and Apoptosis in Ovarian Adenocarcinoma SKOV-3 Cells through Activating the p38 MAPK and Suppressing AKT

doi: 10.3390/ph16050755

Figure Lengend Snippet: Effects of RES on cell cycle distribution of SKOV-3 cells. SKOV-3 cells were treated with RES (25, 50, and 100 µM) alone or in combination with cisplatin (20 µM) for 24 h and 48 h. ( A ) Cell cycle analysis was performed via flow cytometry as described in the . ( B ) The right graph shows the percent of the cell population at each stage of the cell cycle; values represent mean ± SD of three independent experiments. * p < 0.05 vs. untreated cells; # p < 0.05 vs. cisplatin-treated cells. ( C ) Western blot analysis of cyclin A2, cyclin B1, and cyclin E1 expression was performed as described in the ‘Materials and Methods’; β-actin was used to normalize protein expression. ( D ) The quantification of cyclin A2 protein expression assayed via densitometry. ( E ) The quantification of cyclin B1 protein expression assayed via densitometry. ( F ) The quantification of cyclin E1 protein expression assayed via densitometry. Values represent mean ± SD of three independent experiments. * p < 0.05 vs. untreated cells; # p < 0.05 vs. cisplatin-treated cells.

Article Snippet: Rabbit anti-phospho p38 antibody, rabbit anti p38 antibody, rabbit anti-phospho ERK1/2 antibody, mouse anti-ERK1/2 antibody, rabbit anti-phospho JNK antibody, rabbit anti-JNK antibody, rabbit anti-phospho AKT (Ser473) antibody, mouse anti-AKT antibody, rabbit anti-cyclin A2 antibody, rabbit anti-cyclin B1 antibody, rabbit anti-cyclin E1 antibody, rabbit anti-caspase-9 antibody, rabbit anti-caspase-3 antibody, rabbit anti-PARP antibody, mouse anti-β-actin antibody, and DAPI (4, 6-diamidino-2-phenylindole, dihydrochloride) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Cell Cycle Assay, Flow Cytometry, Western Blot, Expressing

Journal: Pharmaceuticals

Article Title: Resveratrol Enhances Cytotoxic Effects of Cisplatin by Inducing Cell Cycle Arrest and Apoptosis in Ovarian Adenocarcinoma SKOV-3 Cells through Activating the p38 MAPK and Suppressing AKT

doi: 10.3390/ph16050755

Figure Lengend Snippet: Effects of RES on regulating the MAPKs and PI3K/AKT pathways in SKOV-3 cells. ( A ) Western blot analysis of p-p38, total p38, p-ERK1/2, total ERK1/2, p-JNK, total JNK, p-AKT and total AKT in cells treated with 25 µM of RES at various time points (0–240 min). β-actin was shown as a loading control. Quantification of p-p38 ( B ), p-ERK1/2 ( C ), p-JNK ( D ) and p-AKT ( E ) upon treatment with 25 µM of RES at various time points (0–240 min). Values represent mean ± SD of three independent experiments. * p < 0.05 vs. RES-treated cells at 0 min. Immunofluorescence study of intracellular p-p38 (green) ( F ), p-ERK1/2 (red) ( G ), p-JNK (green) ( H ) and p-AKT (green) ( I ) in RES (25 µM)-treated cells. Nuclei were counterstained with DAPI (blue) (100× magnification).

Article Snippet: Rabbit anti-phospho p38 antibody, rabbit anti p38 antibody, rabbit anti-phospho ERK1/2 antibody, mouse anti-ERK1/2 antibody, rabbit anti-phospho JNK antibody, rabbit anti-JNK antibody, rabbit anti-phospho AKT (Ser473) antibody, mouse anti-AKT antibody, rabbit anti-cyclin A2 antibody, rabbit anti-cyclin B1 antibody, rabbit anti-cyclin E1 antibody, rabbit anti-caspase-9 antibody, rabbit anti-caspase-3 antibody, rabbit anti-PARP antibody, mouse anti-β-actin antibody, and DAPI (4, 6-diamidino-2-phenylindole, dihydrochloride) were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Western Blot, Immunofluorescence