pc9 cells (egfr exon 19 deletion/e746-a750)  (BioResource International Inc)

Journal: Scientific Reports

Article Title: MicroRNA-130a-3p regulates osimertinib resistance by targeting runt-related transcription factor 3 in lung adenocarcinoma

doi: 10.1038/s41598-024-76196-1

Figure Lengend Snippet: Characteristics of EVs secreted by PC9 and PC9OR cells in the medium ( a ) TEM images of isolated EVs from PC9 and PC9OR cells. The black sphere image shows 40-nm gold colloids bound to the CD9 antibody. Scale bars:100 nm. ( b ) Protein concentrations of CD9 and CD81 (exosomal markers) in isolated EVs from PC9 and PC9OR cells. Original blots are presented in Supplementary Figure S4a. These images show that EVs were isolated ( a , b ). ( c ) Size distribution of isolated EVs from PC9 and PC9OR cells. ( d ) Protein concentrations of isolated total EVs from PC9 and PC9OR cells. ( e ) Number of CD63- and CD9-positive EVs from PC9 and PC9OR cells. The release of EVs differed in the PC9 and PC9OR cells ( c–e ). Data are presented as mean ± SEM ( n = 3–4) ( d , e ). * P < 0.05, compared with PC9 cells. P-values were determined using Student’s t-test. EVs, extracellular vesicles; TEM, transmission electron microscopy.

Article Snippet: PC9 cells ( EGFR exon 19 deletion/E746-A750) and H1975 cells ( EGFR L858R/T790M), which are human lung adenocarcinoma cell lines, were purchased from the Riken BioResource Center (Tsukuba, Japan) and American Type Culture Collection (ATCC, VA), respectively.

Techniques: Isolation, Transmission Assay, Electron Microscopy

Journal: Scientific Reports

Article Title: MicroRNA-130a-3p regulates osimertinib resistance by targeting runt-related transcription factor 3 in lung adenocarcinoma

doi: 10.1038/s41598-024-76196-1

Figure Lengend Snippet: Effects of PC9OR-EV- and PC9OR-EV-derived miRNAs on osimertinib resistance in PC9 cells ( a ) Fluorescence images of PC9 cells after exposure to red-labeled EVs from PC9 (PC9-EVs) and PC9OR cells (PC9OR-EVs). Scale bars: 50 μm. ( b ) Viability of PC9 cells treated with 5 µg of PC9-EVs and PC9OR-EVs in the absence or presence of 1 µM of osimertinib for 72 h. The controls are PC9 cells treated with PBS (vehicle). ( c ) The total concentration of miRNA derived from PC9-EVs and PC9OR-EVs. ( d ) Viability of PC9 cells transfected with 6–12 nM of miRNAs derived from PC9OR-EVs in the absence or presence of 1 µM osimertinib for 72 h. Data are presented as mean ± SEM ( n = 3–6) ( b–d ). * P < 0.05, compared with control or PC9 cells. P-values were determined using Student’s t-test ( c ) or ANOVA with Tukey–Kramer ( b , d ). ns , no significant difference. EVs, extracellular vesicles; ANOVA, analysis of variance.

Article Snippet: PC9 cells ( EGFR exon 19 deletion/E746-A750) and H1975 cells ( EGFR L858R/T790M), which are human lung adenocarcinoma cell lines, were purchased from the Riken BioResource Center (Tsukuba, Japan) and American Type Culture Collection (ATCC, VA), respectively.

Techniques: Derivative Assay, Fluorescence, Labeling, Concentration Assay, Transfection, Control

Journal: Scientific Reports

Article Title: MicroRNA-130a-3p regulates osimertinib resistance by targeting runt-related transcription factor 3 in lung adenocarcinoma

doi: 10.1038/s41598-024-76196-1

Figure Lengend Snippet: Identification of EV-miRNAs associated with osimertinib resistance in lung adenocarcinoma cell lines ( a ) Scatter plot for the miRNA array analysis of isolated EVs from PC9 and PC9OR cells. ( b ) A heat map illustrating the expression of 22 EV-miRNAs and demonstrating a 3.5-fold change between PC9 and PC9OR cells. ( c ) Relative EV-miRNA expression levels in PC9 and PC9OR cells. ( d ) Relative EV-miRNA expression levels in H1975 and H1975OR cells. ( e ) Relative intracellular miRNA levels in PC9 and PC9OR cells. miRNA levels were normalized using U6 levels determined by RT-qPCR ( c–e ). Data are presented as mean ± SEM ( n = 5–6). * P < 0.05, compared with PC9 or H1975 cells. P-values were determined using Student’s t-test. EVs, extracellular vesicles; RT-qPCR, reverse transcription-quantitative polymerase chain reaction.

Article Snippet: PC9 cells ( EGFR exon 19 deletion/E746-A750) and H1975 cells ( EGFR L858R/T790M), which are human lung adenocarcinoma cell lines, were purchased from the Riken BioResource Center (Tsukuba, Japan) and American Type Culture Collection (ATCC, VA), respectively.

Techniques: Isolation, Expressing, Quantitative RT-PCR, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Scientific Reports

Article Title: MicroRNA-130a-3p regulates osimertinib resistance by targeting runt-related transcription factor 3 in lung adenocarcinoma

doi: 10.1038/s41598-024-76196-1

Figure Lengend Snippet: Effect of miR-130a-3p on osimertinib resistance in lung adenocarcinoma cell lines ( a , b ) Relative miR-130a-3p levels in PC9 and H1975 cells transfected with a negative control (NC) and an miR-130a-3p mimic (130a mimic). The miR-130a-3p concentrations were normalized using U6 levels relative to the levels in the cells transfected with the NC. ( c , d ) Viability of PC9 and H1975 cells transfected with the NC and miR-130a-3p mimic after exposure to various concentrations of osimertinib for 72 h. ( e , f ) Apoptotic rate in PC9 and H1975 cells after transfection with the NC and miR-130a-3p mimic at 24, 48, and 72 h in the absence or presence of 1 µM osimertinib. ( g ) Relative miR-130a-3p concentrations in PC9 and PC9OR cells transfected with the NC and miR-130a-3p inhibitor (130a inhibitor). The miR-130a-3p concentrations were normalized using U6 levels relative to the levels in PC9 cells transfected with the NC. ( h ) Cell viability in PC9 and PC9OR cells transfected with the NC and 130a inhibitor in the absence or presence of 1 µM osimertinib for 72 h. Cell viability is the percentage of viable cells relative to PC9 cells transfected with the NC in the absence of osimertinib. When the standard errors of the means are small, they are contained within the symbols. Data are presented as mean ± SEM ( n = 3–12). * P < 0.05, compared with NC. P-values were determined using Student’s t-test ( a , b ) or ANOVA with Tukey–Kramer ( c–h ). RLUs, relative luminescence units; ANOVA, analysis of variance.

Article Snippet: PC9 cells ( EGFR exon 19 deletion/E746-A750) and H1975 cells ( EGFR L858R/T790M), which are human lung adenocarcinoma cell lines, were purchased from the Riken BioResource Center (Tsukuba, Japan) and American Type Culture Collection (ATCC, VA), respectively.

Techniques: Transfection, Negative Control

Journal: Scientific Reports

Article Title: MicroRNA-130a-3p regulates osimertinib resistance by targeting runt-related transcription factor 3 in lung adenocarcinoma

doi: 10.1038/s41598-024-76196-1

Figure Lengend Snippet: Effect of RUNX3 on osimertinib resistance in PC9 cells ( a ) Identification of the target genes of miR-130a-3p using miRDB, TargetScan, and miRTarBase databases. ( b , c ) Protein concentrations of RUNX3 in PC9 and PC9OR cells normalized with β-actin. ( d ) Protein concentrations of RUNX3 and β-actin in PC9 cells transfected with the negative control (NC) and the miR-130a-3p mimic (130a mimic). ( e ) Relative mRNA expressions of RUNX3 in PC9 cells transfected with NC and miR-130a-3p inhibitor (130a inhibitor). ( f ) Protein concentrations of RUNX3 and β-actin in PC9 and PC9OR cells transfected with the NC and the 130a inhibitor. ( g ) Relative mRNA expression of RUNX3 in PC9 cells transfected with NC and si-RUNX3. The RUNX3 mRNA levels were normalized using the glyceraldehyde-3-phosphate dehydrogenase concentrations relative to the levels in PC9 cells transfected with the NC ( e , g ). ( h ) Protein concentrations of RUNX3 and β-actin in PC9 cells transfected with si-RUNX3. ( i ) Viability of PC9 cells transfected with si-RUNX3 in the absence or presence of 1 µM osimertinib for 72 h. Data are presented as mean ± SEM ( n = 3–6). * P < 0.05, compared with PC9 cells or NC. P-values were determined using Student’s t-test ( c , e , g ) or ANOVA with Tukey–Kramer ( i ). Original blots are presented in Supplementary Figures S4b-d. ANOVA, analysis of variance.

Article Snippet: PC9 cells ( EGFR exon 19 deletion/E746-A750) and H1975 cells ( EGFR L858R/T790M), which are human lung adenocarcinoma cell lines, were purchased from the Riken BioResource Center (Tsukuba, Japan) and American Type Culture Collection (ATCC, VA), respectively.

Techniques: Transfection, Negative Control, Expressing

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: Morphology of cells after treatment with gefitinib in PC9 and PC9/N cells. Cells were seeded in six-well culture plates, and after 24 hrs, the cells were treated with gefitinib (0, 0.1, or 1 µM) for 48 hrs. Cells were photographed under a phase contrast inverted microscope. Cells were stained with Wright and Giemsa (original magnification 200×). These results are representative of technical triplicate.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Inverted Microscopy, Staining

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: Cell viability assay for PC9 and PC9/N cells cultured in the presence of gefitinib. Cell growth inhibition in response to gefitinib was evaluated by using the MTT assay. Cells were treated with the indicated concentrations of gefitinib and cell viability was determined 48 hrs later. There are mean of independent triplicate experiments. The data are presented as means ± SEM from technical triplicate. PC9/N: nicotine (1 µM) expose for 3 months in PC9 cells. *, P<0.05; **, P<0.01 and ***, P<0.001 compared with PC9 + gefitinib 0 µM. ## , P<0.01 and ### , P<0.001 compared with PC9/N + gefitinib 0 µM. $ , P<0.05 between two cells with gefitinib 0.01 µM. Data are presented as mean ± standard error of the mean (SEM). SEM, standard error.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Viability Assay, Cell Culture, Inhibition, MTT Assay

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: The mRNA expression of cells after treatment with gefitinib in PC9 and PC9/N cells. Cells were treated with gefitinib (0, 0.1, or 1 µM) for 48 hrs, and the mRNA expressions of α1-nAchR, and PD-L1 were examined by quantitative reverse transcription (qRT)-PCR (A) and RT-PCR (B). These results are representative of technical triplicate. *, P<0.05 and ***, P<0.001 compared with PC9 cells. ### , P<0.001 compared with PC9 cells + gefitinib 0 µM and $$$ , P<0.001 compared with PC9/N + gefitinib 0 µM. nAchR, nicotinic acetylcholine receptors; PD-L1, programmed death ligand 1.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Expressing, Reverse Transcription, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: The protein expression levels of cells after treatment with gefitinib in PC9 and PC9/N cells. The cells were cultured with gefitinib (0, 0.1, or 1 µM) for 48 hrs, and EGFR, mTOR, AKT, PD-L1 and α1-nAchR was detected by Western blot. β-actin was used as an internal control. Western blot was quantified by densitometry and ImageJ. These results are representative of technical triplicate. EGFR, epidermal growth factor receptor; PD-L1, programmed death ligand 1; nAchR, nicotinic acetylcholine receptors.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Expressing, Cell Culture, Western Blot, Control

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: Immunofluorescence staining of PD-L1 and phosphorylation of EGFR in PC9 and PC9/N cells treated with 0.1 µM gefitinib. The localizations of PD-L1 and p-EGFR (green signal; Alexa488) in PC9 and PC9/N cells were shown by immunofluorescence counterstained with DAPI (blue signal) and analyzed by confocal microscopy (original magnification 400×). These results are representative of technical triplicate. DAPI, 4',6-diamidino-2-phenylindole; EGFR, epidermal growth factor receptor.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Immunofluorescence, Staining, Phospho-proteomics, Confocal Microscopy

Journal: Translational Cancer Research

Article Title: Chronic nicotine exposure affects programmed death-ligand 1 expression and sensitivity to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer

doi: 10.21037/tcr.2019.05.02

Figure Lengend Snippet: Overall proportion of PD-L1 TPS expression according to pack-year in NSCLC patients harboring activating EGFR mutation. Heavy smokers ( ≥ 30 PY) tended to have higher expression ( ≥ 50% PD-L1 TPS) than never and light smokers, however, Fisher exact test showed no statistical significance (P value =0.628). PD-L1, programmed death ligand 1; TPS, tumor proportion score; NSCLC, non-small cell lung cancer; EGFR, epidermal growth factor receptor; PY, pack-year.

Article Snippet: Human NSCLC cell line PC9 (EGFR exon 19 deletion) was purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Expressing, Mutagenesis