nsclc cells (pc9 hcc827  (Procell Inc)

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Ubiquitin‐specific peptidase 22 (USP22) was highly expressed in non‐small cell lung cancer (NSCLC) tissues and cells. (a) USP22 expression in NSCLC tissues was evaluated by immunohistochemistry (IHC) assay. (b, c) USP22 mRNA and protein levels in NSCLC tissues and adjacent normal tissues were measured by qRT‐PCR and western blot assays. (d, e) The mRNA and protein levels of USP22 in 16HBE, PC9, and HCC827 cells were determined by qRT‐PCR assay and western blot assay, respectively. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

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

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Deficiency of ubiquitin‐specific peptidase 22 (USP22) repressed non‐small cell lung cancer (NSCLC) cell migration and aggravated apoptosis and ferroptosis. (a) The protein level of USP22 in sh‐NC or sh‐USP22‐transfected PC9 and HCC827 cells was measured by western blot. (b) The migration of PC9 and HCC827 cells with sh‐NC or sh‐USP22 transfection was evaluated by transwell assay. (c) The apoptosis of PC9 and HCC827 cells transfected with sh‐NC or sh‐USP22 was analyzed by flow cytometry analysis. (d–g) The levels of reactive oxygen species (ROS), MDA, Fe 2+ , and GSH in sh‐NC or sh‐USP22 transfected PC9 and HCC827 cells were examined by relevant commercial kits. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: Migration, Transfection, Western Blot, Transwell Assay, Flow Cytometry

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Ferroptosis promoted the effect of gefitinib on non‐small cell lung cancer (NSCLC) cells. (a) The migration ability of PC9 and HCC827 cells treated with DMSO or 2.5‐μM gefitinib was assessed by transwell assay. (b) The apoptosis of DMSO or 2.5‐μM gefitinib‐treated PC9 and HCC827 cells was analyzed by flow cytometry analysis. (c, d) After PC9 and HCC827 cells were treated with DMSO, gefitinib, Fer‐1, or gefitinib+Fer‐1, the oxidized C11‐BODIPY fluorescence intensity was examined by C11‐BODIPY staining. (e–g) After PC9 and HCC827 cells were treated with dimethyl sulfoxide (DMSO), gefitinib, Fer‐1, or gefitinib+Fer‐1, the levels of MDA, Fe 2+ , and glutathione (GSH) were examined by relevant commercial kits. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: Migration, Transwell Assay, Flow Cytometry, Fluorescence, Staining

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Ubiquitin‐specific peptidase 22 (USP22) overexpression promoted gefitinib resistance and repressed ferroptosis in gefitinib‐treated non‐small cell lung cancer (NSCLC) cells. (a) USP22 protein level in gefitinib or dimethyl sulfoxide (DMSO)‐treated PC9 and HCC827 cells was measured by western blot. (b) The effect of USP22 on gefitinib resistance was assessed by CCK‐8 assay. (c–i) PC9 and HCC827 cells were treated with DMSO+pcDNA, gefitinib+pcDNA, or gefitinib+USP22. (c, d) The migration and apoptosis of PC9 and HCC827 cells were evaluated by transwell assay and flow cytometry analysis, respectively. (e, f) The oxidized C11‐BODIPY fluorescence intensity in PC9 and HCC827 cells was examined by C11‐BODIPY staining. (g–i) The levels of MDA, Fe 2+ , and glutathione (GSH )in PC9 and HCC827 cells were examined by relevant commercial kits. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: Over Expression, Western Blot, CCK-8 Assay, Migration, Transwell Assay, Flow Cytometry, Fluorescence, Staining

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Ubiquitin‐specific peptidase 22 (USP22) stabilized murine double minute 2 (MDM2) and modulated MDM2 expression. (a) Ubibrowser software predicted the relation between USP22 and MDM2. (b) Pull‐down assay was conducted to analyze the level of MDM2 ubiquitination. (c) Co‐IP assay was performed to explore the interaction between USP22 and MDM2. (d) The expression of MDM2 in non‐small cell lung cancer (NSCLC) tissues and normal tissues was estimated through IHC assay. (e, f) The mRNA and protein levels of MDM2 in NSCLC tissues and normal tissues were measured by qRT‐PCR and western blot, respectively. (g, h) The mRNA and protein levels of MDM2 in 16HBE, PC9, and HCC827 cells were examined by qRT‐PCR and western blot, respectively. (i) After PC9 and HCC827 cells were treated with DMSO, gefitinib, gefitinib+USP22, or gefitinib+USP22 + sh‐MDM2, MDM2 protein level was measured by western blot. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: Expressing, Software, Pull Down Assay, Co-Immunoprecipitation Assay, Quantitative RT-PCR, Western Blot

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Murine double minute 2 (MDM2) knockdown reversed the effects of Ubiquitin‐specific peptidase 22 (USP22) overexpression on gefitinib sensitivity and ferroptosis in gefitinib‐treated NSCLC cells. (a) IC50 of gefitinib in PC9 and HCC827 cells transfected with pcDNA, USP22, or USP22 + sh‐MDM2 was estimated through CCK‐8 assay. (b–h) PC9 and HCC827 cells were treated with dimethyl sulfoxide (DMSO), gefitinib, gefitinib+USP22, gefitinib+USP22 + sh‐NC, or gefitinib+USP22 + sh‐MDM2. (b, c) The migration and apoptosis of PC9 and HCC827 cells were explored by transwell assay and flow cytometry analysis. (d, e) The oxidized C11‐BODIPY fluorescence intensity in PC9 and HCC827 cells was examined by C11‐BODIPY staining. (f–h) The levels of MDA, Fe 2+ , and glutathione (GSH) in PC9 and HCC827 cells were determined with commercial kits. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: Knockdown, Over Expression, Transfection, CCK-8 Assay, Migration, Transwell Assay, Flow Cytometry, Fluorescence, Staining

Journal: Thoracic Cancer

Article Title: USP22 promotes gefitinib resistance and inhibits ferroptosis in non‐small cell lung cancer by deubiquitination of MDM2

doi: 10.1111/1759-7714.15439

Figure Lengend Snippet: Ubiquitin‐specific peptidase 22 (USP22) enhanced gefitinib resistance of non‐small cell lung cancer (NSCLC) cells in vivo. (a) The protein levels of USP22 and murine double minute 2 (MDM2) in PBS, gefitinib, and gefitinib+USP22 treated PC9 cells were measured by western blot. (b) Xenograft tumor volume was monitored every 5 days. (c) Xenograft tumor weight was examined after 25 days. (d) The expression of USP22 and MDM2 in xenograft tumors was examined through immunohistochemistry (IHC) assay. * p < 0.05.

Article Snippet: NSCLC cells (PC9 and HCC827) were purchased from Procell (Wuhan, China).

Techniques: In Vivo, Western Blot, Expressing, Immunohistochemistry

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: The expression of circCCDC134 in non‐small cell lung cancer (NSCLC) tissues and cells. (a) The circCCDC134 expression was measured by quantitative real‐time PCR (qRT‐PCR) in NSCLC tumor tissues and adjacent normal tissues. (b) The circCCDC134 expression in the tumor tissues of NSCLC patients with different stages was detected by qRT‐PCR. (c) QRT‐PCR was used to determine the circCCDC134 expression in NSCLC cells and BEAS‐2B cells. Subcellular localization analysis (d, e), RNase R assay (f, g), and ActD assay (h–i) were used to assess the circular features of circCCDC134. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

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

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: The regulation of circCCDC134 knockdown on non‐small cell lung cancer (NSCLC) cell functions. PC9 and A549 cells were transfected with sh‐NC or sh‐circCCDC134#1/#2. (a) The circCCDC134 expression was assessed by qRT‐PCR. Colony formation assay (b), 5‐ethynyl‐2′‐deoxyuridine (EdU) assay (c), transwell assay (d, e), wound healing assay (f) and flow cytometry (g) were used to measure cell proliferation, migration, invasion and apoptosis. (h, i) Protein expression was detected by western blot (WB) analysis. (j–l) Glucose consumption, lactate production and ATP level were measured to assess cell glycolysis ability. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Knockdown, Transfection, Expressing, Quantitative RT-PCR, Colony Assay, EdU Assay, Transwell Assay, Wound Healing Assay, Flow Cytometry, Migration, Western Blot

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: CircCCDC134 sponged miR‐625‐5p. (a) The binding sites between circCCDC134 and miR‐625‐5p are shown. (b) The transfection efficiency of miR‐625‐5p mimic and inhibitor was confirmed by quantitative real‐time PCR (qRT‐PCR). Dual‐luciferase reporter assay (c, d) and RNA immunoprecipitation (RIP) assay (e, f) were used to assess RNA interaction. (g) The miR‐625‐5p expression was measured by qRT‐PCR in non‐small cell lung cancer (NSCLC) tumor tissues and adjacent normal tissues. (h) QRT‐PCR was used to determine the miR‐625‐5p expression in NSCLC cells and BEAS‐2B cells. (i) The miR‐625‐5p expression was detected by qRT‐PCR in NSCLC cells transfected with sh‐circCCDC134#1 and in‐miR‐625‐5p. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Binding Assay, Transfection, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Luciferase, Reporter Assay, RNA Immunoprecipitation, Expressing

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: The regulation of sh‐circCCDC134#1 and in‐miR‐625‐5p on non‐small cell lung cancer (NSCLC) cell behaviors. PC9 and A549 cells were transfected with sh‐circCCDC134#1 and in‐miR‐625‐5p. Colony formation assay (a), 5‐ethynyl‐2′‐deoxyuridine (EdU) assay (b), transwell assay (c, d), wound healing assay (e) and flow cytometry (f) were performed to detect cell proliferation, migration, invasion, and apoptosis. (g, h) Western blot (WB) analysis was used to test protein expression. (i–k) Cell glycolysis ability was measured by detecting glucose consumption, lactate production and ATP level. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Transfection, Colony Assay, EdU Assay, Transwell Assay, Wound Healing Assay, Flow Cytometry, Migration, Western Blot, Expressing

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: MiR‐625‐5p could target NFAT5. (a) The binding sites between NFAT5 3'UTR and miR‐625‐5p are shown. (b, c) Dual‐luciferase reporter assay was used to assess RNA interaction. (d, e) The NFAT5 mRNA and protein expression in non‐small cell lung cancer (NSCLC) tumor tissues and adjacent normal tissues was measured by qRT‐PCR and western blot (WB) analysis. (f) WB analysis was used to determine the NFAT5 protein expression in NSCLC cells and BEAS‐2B cells. (g, h) The NFAT5 protein expression was detected by WB analysis under the specified transfection conditions. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Binding Assay, Luciferase, Reporter Assay, Expressing, Quantitative RT-PCR, Western Blot, Transfection

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: The regulation of miR‐625‐5p and NFAT5 on non‐small cell lung cancer (NSCLC) cell behaviors. PC9 and A549 cells were transfected with miR‐625‐5p mimic and pcDNA NFAT5 overexpression vector. Cell proliferation, migration, invasion and apoptosis were determined using colony formation assay (a), 5‐ethynyl‐2′‐deoxyuridine (EdU) assay (b), transwell assay (c, d), wound healing assay (e) and flow cytometry (f). (g, h) Western blot (WB) analysis was performed to examine protein expression. (i–k) Glucose consumption, lactate production, and ATP level were analyzed to evaluate cell glycolysis ability. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Transfection, Over Expression, Plasmid Preparation, Migration, Colony Assay, EdU Assay, Transwell Assay, Wound Healing Assay, Flow Cytometry, Western Blot, Expressing

Journal: Thoracic Cancer

Article Title: A novel molecular mechanism mediated by circ CCDC134 regulates non‐small cell lung cancer progression

doi: 10.1111/1759-7714.14942

Figure Lengend Snippet: The regulation of circCCDC134 knockdown on non‐small cell lung cancer (NSCLC) tumor growth. A549 cells transfected with sh‐circCCDC134#1/sh‐NC were injected into nude mice. Tumor volume (a) and weight (b) were detected. (c, d) The circCCDC134 and miR‐625‐5p expression was examined by quantitative real‐time PCR (qRT‐PCR). (e) The NFAT5 protein expression was tested by western blot (WB) analysis. (f) Immunohistochemical (IHC) staining was used to assess NFAT5 positive cells. * p < 0.05.

Article Snippet: Human NSCLC cells (H1299, HCC827, PC9, and A549) and a normal bronchial epithelial cell line (BEAS‐2B) were purchased from Biovector NTCC (Beijing, China).

Techniques: Knockdown, Transfection, Injection, Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Immunohistochemical staining, Immunohistochemistry

Journal: Journal of Clinical Oncology

Article Title: ROS1 Rearrangements Define a Unique Molecular Class of Lung Cancers

doi: 10.1200/JCO.2011.35.6345

Figure Lengend Snippet: (A) Dose-response cell survival curves of ROS1-rearranged cell line (HCC78) and ROS1-negative cell lines in response to crizotinib (nM); ALK-positive lines H3122 and MGH006 are positive controls for crizotinib response, and the PC-9, HCC827, and crizotinib-resistant H3122 CR lines are negative controls. (B) Western blot reveals a three-fold reduction of phospho-ROS1 in HCC78 cells at the same concentration (300 nmol/L) of crizotinib that results in near-complete reduction of phospho-ALK in H3122 cells. Total ROS1 and ALK, as well as actin, are shown as controls. The concentration of crizotinib is indicated above each lane (in nM).

Article Snippet: Cell Line Crizotinib Sensitivity Testing Human NSCLC lines PC9, HCC827, MGH006, NCI-H3122, and HCC-78 cells (obtained from DSMZ, Braunschweig, Germany) were cultured in RPMI 1640 supplemented with 10% fetal bovine serum (RPMI 1640 growth medium).

Techniques: Western Blot, Concentration Assay