Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: Effect of JNK inhibition on A549 cell viability as a function of cisplatin concentration. (a, b) 7.5 μg/mL concentration of cisplatin becomes lethal in combination with JNK inhibitor SP600125. However, at higher cisplatin concentrations, the inhibitor either protects against cisplatin-induced cell death (a) or does not have any effect (b). (c) Similar effects are observed in the colon cancer cell line DLD-1. (d–g) SP600125 potentiates the appearance of cells with apoptotic morphology in 7.5 μg/mL cisplatin-treated A549 cells (d, e), in contrast to 30 μg/mL cisplatin-treated A549 cells (f, g). Relative cell viability is cell viability after 72 h treatment normalized by initial viability (measured by the MTT method). Representative test results (all measurements were performed in quadruplicate) from more than five experiments are presented. Fluorescent pictures were obtained with a mixture of AO/EB dyes as described in the section. SP—JNK inhibitor SP600125 (20 μM).

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Inhibition, Concentration Assay

Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: JNK inhibitor SP600125 reduces JNK target transcription factor c-Jun phosphorylation in cisplatin-treated A549 cells. Different concentrations of cisplatin were used. (a) Representative Western blots from 20 and 40 h of treatments are shown. Total protein Coomassie-stained polyacrylamide gels serve as loading controls. (b) Repeated addition (after 24 h) of SP potentiates suppression of c-Jun phosphorylation in response to cisplatin treatment. (c) Repeated addition of SP strengthens the protective effect of JNK inhibition at 15−30 μg/mL concentrations of cisplatin in A549 cells. SP—JNK inhibitor SP600125 (20 μM). p *** < 0.0005, N = 3.

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Phospho-proteomics, Western Blot, Staining, Inhibition

Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: Cisplatin concentration-dependent effect of different JNK inhibitors on A549 cell viability. Different JNK inhibitors show the same dependence on cisplatin concentration on cell viability. Statistically significant reduction in cell viability is observed at 7.5 μg/mL cisplatin in combination with 10 μM AS601245 (a), 10 μM bentamapimod (b), 2 μM JNK inhibitor IX (c), and 5 μM XG-102 (d). Representative test results (all measurements were performed in quadruplicate) from more than five experiments are presented. (e) Reduction in c-Jun phosphorylation upon the addition of JNK inhibitor XG-102, as determined by Western blot. Representative Western blots are shown. Total protein Coomassie-stained polyacrylamide gels serve as loading controls.

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Concentration Assay, Phospho-proteomics, Western Blot, Staining

Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: Activation of TP53 in A549 cells in response to different concentrations of cisplatin. (a, b) Expression and phosphorylation at serine-6 is induced by cisplatin and is maximal at 15 μg/mL (6 h of cisplatin treatment). (c, d) Prolonged and increasing expression and phosphorylation of TP53 in cells treated with either 7.5 μg/mL or 30 μg/mL of cisplatin. (e) TP53 activator nutlin-3a potentiates cisplatin-induced decrease in cell viability. (f) Nutlin-3a reduces viability of cisplatin + SP600125-treated cells. C—control without cisplatin; Nut—nutlin-3a (10 μM); SP—SP600125 (20 μM). Representative test results (all measurements were performed in quadruplicate) from more than five experiments are presented, p ** < 0.005, p *** < 0.0005, N = 4. (g) Nutlin induces expression of TP53. Representative Western blots are shown. Total protein Coomassie-stained polyacrylamide gels serve as loading controls.

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Activation Assay, Expressing, Phospho-proteomics, Control, Western Blot, Staining

Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: Involvement of AKT signaling pathway in cisplatin-induced A549 cell death. (a) AKT protein level does not depend on cisplatin concentration. 6-h-long exposure is presented in the Western blot picture. (b) Phosphorylation/activation of AKT is cisplatin concentration-dependent. 6-h-long exposure is presented in the Western blot picture. (c) AKT protein level does not change during 40 h of cisplatin treatment. (d) Dynamics of AKT activation following cisplatin (7.5 or 30 μg/mL) treatment. (e, f) AKT inhibitor capivasertib enhances cell death at all concentrations of cisplatin used both in the absence (e) and presence (f) of JNK inhibitor SP600125. (g, h) GSK3-β inhibitor SB415286 protects A549 cells from cisplatin both in the absence (g) and presence (h) of JNK inhibition. Representative test results (all measurements were performed in quadruplicate) from more than five experiments are presented. p*<0.05, p**<0.005, p***<0.0005, N = 4. (i) Capivasertib inhibits AKT activity as shown by the inhibition of AKT molecular target GSK3-β phosphorylation at serine-9. Representative Western blots are shown. Total protein Coomassie-stained polyacrylamide gels serve as loading controls. C—control without cisplatin; CAP—capivasertib (10 μM); SP—SP600125 (20 μM), SB—GSK3 inhibitor SB415286 (15 μM).

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Concentration Assay, Western Blot, Phospho-proteomics, Activation Assay, Inhibition, Activity Assay, Staining, Control

Journal: ACS Omega

Article Title: The Role and Efficacy of JNK Inhibition in Inducing Lung Cancer Cell Death Depend on the Concentration of Cisplatin

doi: 10.1021/acsomega.4c01950

Figure Lengend Snippet: Opposite changes in TP53 and AKT phosphorylation following SP600125 treatment of A549 cells exposed to different concentrations of cisplatin. (a) Expression of TP53 is increased in response to the combination of SP + 7.5 μg/mL cisplatin, in contrast to the combination of SP + 30 μg/mL cisplatin. (b) Phosphorylation of TP53 (serine-6) is increased in response to the combination of SP + 7.5 μg/mL cisplatin, in contrast to the combination of SP + 30 μg/mL cisplatin. (c) Phosphorylation of AKT (threonine-308) is decreased in response to the combination of SP + 7.5 μg/mL cisplatin, in contrast to the combination of SP + 30 μg/mL cisplatin. Representative Western blots are shown. Total protein Coomassie-stained polyacrylamide gels serve as a loading control. SP—JNK inhibitor SP600125 (20 μM); 20 h of treatment.

Article Snippet: Human non-small-cell lung carcinoma A549 cells were purchased from Cell Lines Service GmbH, Eppelheim, Germany.

Techniques: Phospho-proteomics, Expressing, Western Blot, Staining, Control

Journal: Analytical methods : advancing methods and applications

Article Title: Nonspecific nuclear uptake of anti-MUC1 aptamers by dead cells: the role of cell viability monitoring in aptamer targeting of membrane-bound protein cancer biomarkers.

doi: 10.1039/d0ay01878c

Figure Lengend Snippet: Fig. 1 Aptamer and RND1 control binding to the MUC1 peptide and cell line lysates. (A) The ELONA response obtained for 5TR1, 5TR4, 5TRG2, S22 aptamer and RND1 control binding to MUC1 peptide (MUC1) and milk powder control (MP) layers. Significant binding by the biotin-labelled aptamers to wells containing the MUC1 peptide in comparison to the milk powder control layer only is denoted with an asterisk (*, p < 0.05). Error bars represent the standard deviation of the mean (n ¼ 4). Two-way ANOVA tests were performed using the Tukey–Kramer post hoc test, with results from the two-way ANOVA annotated in the graph. (B) SDS-PAGE and corresponding south- western blots of MCF-7, Hs578T, A549, SW480, and SW620 cell lysates using biotin-labelled 5TR1 aptamer, RND1 and no aptamer controls. Southwestern blots of cancer cell line lysates by the 5TR4, 5TRG2, and S22 aptamers shown in ESI as Fig. S1A–C.†

Article Snippet: Cell lines and tissue culture Adherent human breast carcinoma cell lines: MCF-7 (ATCC: HTB-22) and Hs578T (ATCC: HTB-126), and the human lung carcinoma cell line A549 (ATCC: CCL-185) were sourced from This journal is © The Royal Society of Chemistry 2021 American Type Culture Collection (ATCC, USA).

Techniques: Control, Binding Assay, Comparison, Standard Deviation, SDS Page, Western Blot

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: Characterization of A549 and A549/DDP cells. (A) A549 and A549/DDP cells were exposed to different concentrations of DDP (1–32 µg/ml) for 24 h, and cell viability was determined by Cell Counting Kit-8 assay. The protein expression levels of (B) E-cadherin, vimentin, α-SMA and (C) CIP2A, p-AKT, AKT, p-mTOR were measured using western blotting; E, epithelial; SMA, smooth muscle actin; p-mTOR, phosphorylated mammalian target of rapamycin; AKT, protein kinase B; DDP, cisplatin; CIP2A, cancerous inhibitor of protein phosphatase 2A; IC 50 , half maximal inhibitory concentration.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Cell Counting, Expressing, Western Blot, Concentration Assay

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII induce cytotoxicity in A549 and A549/DDP cells. (A) Chemical structure of PPI and PPVII. (B) A549 and A549/DDP cells were treated with different dose of PPI (0.5–8 µg/ml) and PPVII (0.5–8 µg/ml) for 24 h, respectively. In A549 cells, the cell inhibition rate of 0.5 µg/ml PPI was significantly different from other concentration groups (**P<0.01); 1.0 µg/ml was significantly different from 4.0 and 8.0 µg/ml (**P<0.01), and 2.0 µg/ml was significantly different from 8.0 µg/ml (**P<0.01). As for A549/DDP cells, the cell inhibition rates of 0.5 µg/ml PPI and 1.0 µg/ml were both significantly different from other concentration groups ( ## P<0.01), and 2.0 µg/ml was significantly different from 8.0 µg/ml ( ## P<0.01). In A549 cells, there were significant differences in cell inhibition rates of PPVII among different concentration groups (**P<0.01); as for A549/DDP cells, there were significant differences among different concentration groups except for 4.0 and 8.0 µg/ml ( ## P<0.01). PPI, Polyphyllin I; PPVII, polyphyllin VII; DDP, cisplatin.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Inhibition, Concentration Assay

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII possess chemo-sensitizing effects on A549/DDP cells. (A) A549/DDP cells treated with indicated doses of DDP, or DDP plus either 0.375 µg/ml PPI or 0.75 µg/ml PPI, respectively for 24 h. (B) A549/DDP cells treated with indicated doses of DDP, or DDP plus either 0.375 µg/ml PPVII or 0.75 µg/ml PPVII, respectively for 24 h, cell viability was measured with Cell Counting Kit-8 assay. Data were presented as the mean ± standard deviation. *P<0.05, **P<0.01 and ***P<0.001 vs. DDP treated group. # P<0.05 and ## P<0.01. PPI, Polyphyllin I; PPVII, polyphyllin VII; DDP, cisplatin.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Cell Counting, Standard Deviation

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII reduce the IC 50 value of cisplatin in A549/DDP cells

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques:

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII enhance DDP-induced apoptosis. A549/DDP cells were treated with DDP (6 µg/ml), PPI (0.75 µg/ml), PPVII (0.75 µg/ml), or DDP (6 µg/ml) in combination with either PPI (0.75 µg/ml) or PPVII (0.75 µg/ml) for 24 h, respectively. Apoptosis of cells was detected by Annexin V-FITC/PI staining and flow cytometry. The apoptotic percentage is shown as a bar graph. Data represents the mean ± standard deviation of three independent experiments. **P<0.01 and ***P<0.001. PPI, Polyphyllin I; PPVII, polyphyllin VII; DDP, cisplatin; FITC, fluorescein isothiocyanate; PI, propidium iodide.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Staining, Flow Cytometry, Standard Deviation

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII enhance DDP-induced apoptosis through the P53 pathway and caspases-dependent pathway. A549/DDP cells were treated with DDP (6 µg/ml), PPI (0.75 µg/ml), PPVII (0.75 µg/ml), or DDP (6 µg/ml) in combination with either PPI (0.75 µg/ml) or PPVII (0.75 µg/ml) for 24 h, respectively. The protein expression levels of P53, Bax, Bcl-2 (A), PARP, C-PARP, pro-Caspase-3, C-Caspase-3 (B) were measured using western blotting. PPI, Polyphyllin I; PPVII, polyphyllin VII; DDP, cisplatin; C-PARP, cleaved-poly (ADP-ribose) polymerase 1.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Expressing, Western Blot

Journal: Oncology Letters

Article Title: Polyphyllin I and VII potentiate the chemosensitivity of A549/DDP cells to cisplatin by enhancing apoptosis, reversing EMT and suppressing the CIP2A/AKT/mTOR signaling axis

doi: 10.3892/ol.2019.10895

Figure Lengend Snippet: PPI and PPVII reverse EMT and suppress the CIP2A/AKT/mTOR pathway. A549/DDP cells were incubated with DDP (6 µg/ml), PPI (0.75 µg/ml), PPVII (0.75 µg/ml), or DDP (6 µg/ml) in combination with either PPI (0.75 µg/ml) or PPVII (0.75 µg/ml) for 24 h, then cells were harvested for further western blotting analysis. (A) The protein expression levels of E-cadherin, vimentin, α-SMA in A549/DDP cells were measured. (B) The protein expression levels of CIP2A, p-AKT, AKT, p-mTOR, m-TOR were measured. PPI, Polyphyllin I; PPVII, polyphyllin VII; DDP, cisplatin; p-AKT, phosphorylated-protein kinase B; mTOR, mammalian target of rapamycin; CIP2A, cancerous inhibitor of protein phosphatase 2A; SMA, smooth muscle actin; E, epithelial.

Article Snippet: Human NSCLC cell lines A549 and DDP-resistant A549/DDP cells (OriGene Technologies, Inc.) were cultured in RPMI-1640 (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% fetal bovine serum (Gibco; Thermo Fisher Scientific, Inc.) and penicillin (100 U/ml)-streptomycin (100 mg/ml) at 37°C, with 5% CO 2 .

Techniques: Incubation, Western Blot, Expressing