cisplatin Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Millipore cisplatin cisplatin
    <t>Cisplatin</t> and radiation resistance A. Graphs of non-viable (apoptotic+necrotic) cells measured by Annexin V/PI staining after cisplatin (after 24 hours) or radiation (after 48 hours) treatment. Orospheres are more resistant to both treatments. *P
    Cisplatin Cisplatin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin cisplatin/product/Millipore
    Average 99 stars, based on 42 article reviews
    Price from $9.99 to $1999.99
    cisplatin cisplatin - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    85
    Bristol Myers cisplatin platinol
    Effect of bortezomib on tumor accumulation of <t>cisplatin</t> in vivo . A , scattergram of nagogram platinum per milligram tumor wet weight as a function of tumor wet weight +, no prior bortezomib; +, 1mg/kg bortezomib i.p. 4 h prior to injection of cisplatin by the i.p. route. B , mean nanogram platinum per milligram tumor wet weight. Vertical bars, SE.
    Cisplatin Platinol, supplied by Bristol Myers, used in various techniques. Bioz Stars score: 85/100, based on 59 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin platinol/product/Bristol Myers
    Average 85 stars, based on 59 article reviews
    Price from $9.99 to $1999.99
    cisplatin platinol - by Bioz Stars, 2020-08
    85/100 stars
      Buy from Supplier

    94
    Abcam cisplatin
    Antioxidant effect of NY and PS on <t>cisplatin-induced</t> cell death in HK-2 cells. Cells were treated with 10 μM cisplatin for 30 m and the level of ROS was detected by microplate reader ( a ). Cells were exposed to cisplatin for 24 h with or without PS or NY ( b ). The measures of JC-1 fluorescence intensity ( c and d ). Cells were treated to bind Annexin V and then to undergo PI staining; exposed to cisplatin for 24 h with or without PS or NY. HK-2 cells were treated with cisplatin for 24 h to measure apoptosis ( e ). Cell lysates were assayed for caspase-9, 3 activity. The statistical significance (* p
    Cisplatin, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 84 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin/product/Abcam
    Average 94 stars, based on 84 article reviews
    Price from $9.99 to $1999.99
    cisplatin - by Bioz Stars, 2020-08
    94/100 stars
      Buy from Supplier

    93
    Selleck Chemicals cisplatin
    Phendione inhibits the proliferation of melanoma with intrinsic resistance to MAPK inhibitors. ( A–E ) Panels show growth inhibition curves and IC50 values. Primary patient-derived melanoma cell lines with BRAF V600E , BRAF V600K , and NRAS Q61R mutations ( A–C ), and human melanoma cell line SK-MEL28 ( D ) and its BRAF inhibitor (dabrafenib)-resistant line SK-MEL28R ( E ) were treated with phendione or inhibitors of the MAPK pathway for 96 h. Data are presented as mean ± SD of three independent experiments. ( F ) Cell cycle analysis for A375 BRAFi-resistant cells. Cells were treated with 2.5 µM <t>cisplatin,</t> 0.2 µM phendione, or vehicle for the indicated time. Graph shows the percentage of cells in each phase of the cell cycle. Data shown are average ± SD of three independent experiments. ( G ) Comet assay to measure DNA damage. Representative images are shown at the left and quantification results are shown at the right . Data are presented as mean ± SD of three independent experiments. ( H ) Immunoblots of whole-cell lysate collected from comet assay. ( I , J ) Quantification of reactive oxygen species (ROS) production measured by flow cytometry in A375 ( I ) and A375 BRAFi-resistant ( J ) cells. Hydroxyl peroxide (H 2 O 2 ; 200 µM) was used as positive control. Data are presented as means ± SEMs of three independent experiments. (*) P
    Cisplatin, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 93/100, based on 632 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin/product/Selleck Chemicals
    Average 93 stars, based on 632 article reviews
    Price from $9.99 to $1999.99
    cisplatin - by Bioz Stars, 2020-08
    93/100 stars
      Buy from Supplier

    95
    Tocris cisplatin
    Collagen promoted the proliferation and migration of head and neck squamous cell carcinoma (HNSCC) cells and suppressed the response to <t>cisplatin.</t> ( A ) Collagen significantly increased the growth of the HNSCC cell lines, SCC040, SCC154, VU040T and VU147T. ( B ) Pre-treatment of cells with collagen significantly enhanced the migration of cells through fibronectin-coated membranes. ( C ) Cells pre-treated with collagen were significantly less sensitive to cisplatin, as determined by Annexin V-FITC/Propidium Iodide staining following treatment with 50 μM cisplatin for 72 hours. Results shown are mean +/− standard deviation values of triplicates. *, **, ***, **** denote p
    Cisplatin, supplied by Tocris, used in various techniques. Bioz Stars score: 95/100, based on 148 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin/product/Tocris
    Average 95 stars, based on 148 article reviews
    Price from $9.99 to $1999.99
    cisplatin - by Bioz Stars, 2020-08
    95/100 stars
      Buy from Supplier

    92
    LKT Laboratories cisplatin
    SETD8 inhibition preferentially sensitises LUSC cell lines to chemotherapy. a Dose–response curves were derived by treating NSCLC cell lines with SETD8 inhibitor NSC663284. 1000 cells were seeded and allowed to recover for 24 h. The inhibitor was then added at increasing concentrations to LUSC (red) and LUAD (blue) cells and Cell Titre (see Methods) assay was performed after 72 h. b IC 50 values were derived from the dose–response assay indicating LUSC cells are significantly more responsive to SETD8 inhibition than LUAD cells. c Dose–response curves were derived by treating NSCLC cell lines with <t>cisplatin</t> as above. d IC 50 values were derived from the dose–response assay indicating cisplatin effects LUSC and LUAD cells equally. e Dose–response curves derived from treating NSCLC cell lines with cisplatin and NSC663284 IC 50 concentration for each cell line as above. f IC 50 values were derived from the dose–response assay indicating SETD8 inhibition preferentially enhances cisplatin efficacy in LUSC cells. The whiskers indicate the range of the data and the line represents the median. Data presented as mean ± s.d. (LUSC n = 5 and LUAD n = 6). Student’s t -test performed, * p
    Cisplatin, supplied by LKT Laboratories, used in various techniques. Bioz Stars score: 92/100, based on 20 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin/product/LKT Laboratories
    Average 92 stars, based on 20 article reviews
    Price from $9.99 to $1999.99
    cisplatin - by Bioz Stars, 2020-08
    92/100 stars
      Buy from Supplier

    88
    Corden platinol
    SETD8 inhibition preferentially sensitises LUSC cell lines to chemotherapy. a Dose–response curves were derived by treating NSCLC cell lines with SETD8 inhibitor NSC663284. 1000 cells were seeded and allowed to recover for 24 h. The inhibitor was then added at increasing concentrations to LUSC (red) and LUAD (blue) cells and Cell Titre (see Methods) assay was performed after 72 h. b IC 50 values were derived from the dose–response assay indicating LUSC cells are significantly more responsive to SETD8 inhibition than LUAD cells. c Dose–response curves were derived by treating NSCLC cell lines with <t>cisplatin</t> as above. d IC 50 values were derived from the dose–response assay indicating cisplatin effects LUSC and LUAD cells equally. e Dose–response curves derived from treating NSCLC cell lines with cisplatin and NSC663284 IC 50 concentration for each cell line as above. f IC 50 values were derived from the dose–response assay indicating SETD8 inhibition preferentially enhances cisplatin efficacy in LUSC cells. The whiskers indicate the range of the data and the line represents the median. Data presented as mean ± s.d. (LUSC n = 5 and LUAD n = 6). Student’s t -test performed, * p
    Platinol, supplied by Corden, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/platinol/product/Corden
    Average 88 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    platinol - by Bioz Stars, 2020-08
    88/100 stars
      Buy from Supplier

    99
    Millipore cisplatin
    mtFoxO3A is involved in cancer cell response to metabolic stress. a HCT116-FoxO3A +/+ and HCT116-FoxO3A −/− cells were subjected to different treatments: glucose restriction (LG, 0.75 mM glucose, 24 h), metformin (MET, 10 μM, 72 h), <t>cisplatin</t> (CDDP, 30 μM, 48 h), irinotecan (CPT-11, 30 μM, 24 h), 5-fluorouracil (5-FU, 2 μM, 24 h) and etoposide (VP-13, 40 μM, 24 h). Relative cell viability and relative cell death were calculated. b Correlation between LG-resistance (days) and mitochondrial FoxO3A (mtFoxO3A) protein levels in different human cell lines (HCT116 and HT29 colorectal cancer cells, HEK293 embryonic kidney cell, DU145 prostate cancer cells, A549 lung cancer cells, MDA-MB-468 breast cancer cells and OVCAR3 ovarian cancer cells). a.u. arbitrary units. c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids (48 h) and subjected to LG (24 h). Upper panel: relative cell viability and relative cell death. Lower panel: immunoblot analysis of total proteins. β-actin: loading control. d Transcription analysis of selected mitochondrial ( ND6 and COX1 ) and nuclear ( BIM ) genes by RT-PCR in HCT116-FoxO3A −/− cells transfected with the indicated plasmids (48 h) and subjected to LG (24 h). e HCT116-FoxO3A −/− cells, transfected with the indicated plasmids (48 h), were subjected to metabolic stress with 2-DG (1 mM, 6 h). The graph reflects the quantification of tetramethylrhodamine ethyl ester (TMRE) fluorescence of active mitochondria in transfected cells. f Clonogenic assay on HCT116-FoxO3A +/+ cells cultured in LG (24 h) and treated with increasing concentrations of trametinib and/or compound C, as indicated, for 24 h. Cell growth percent inhibition at each drug concentration is presented. g Left panel: immunoblot analysis of total and mitochondrial proteins isolated from tumors ( n ≥ 7 for each group) derived from HCT116-xenografted nude mice subjected to 2-DG treatment (100 mg/kg, 6 days). β-actin and HSP60 were used as total and mitochondrial loading control, respectively. Right panel: densitometric analysis of full-length and cleaved FoxO3A normalized against the mitochondrial loading control and the results of the densitometric analysis of the phosphorylated-AMPK and ERK normalized against total AMPK and ERK, respectively, and the loading control. Data are presented as mean ± SEM and significance was calculated with Student’s t test; * p
    Cisplatin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 10270 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/cisplatin/product/Millipore
    Average 99 stars, based on 10270 article reviews
    Price from $9.99 to $1999.99
    cisplatin - by Bioz Stars, 2020-08
    99/100 stars
      Buy from Supplier

    Image Search Results


    Cisplatin and radiation resistance A. Graphs of non-viable (apoptotic+necrotic) cells measured by Annexin V/PI staining after cisplatin (after 24 hours) or radiation (after 48 hours) treatment. Orospheres are more resistant to both treatments. *P

    Journal: Oncotarget

    Article Title: Increased fucosylation has a pivotal role in invasive and metastatic properties of head and neck cancer stem cells

    doi:

    Figure Lengend Snippet: Cisplatin and radiation resistance A. Graphs of non-viable (apoptotic+necrotic) cells measured by Annexin V/PI staining after cisplatin (after 24 hours) or radiation (after 48 hours) treatment. Orospheres are more resistant to both treatments. *P

    Article Snippet: In vitro treatment with cisplatin Cisplatin (cis-diammineplatinum (II) dischloride, DDP) (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in sterile 0.9% NaCl to achieve a stock concentration of 5 mM.

    Techniques: Staining

    HCMV infection blocks cisplatin A mediated activation of Bak.

    Journal: Journal of molecular biochemistry

    Article Title: HCMV activation of ERK-MAPK drives a multi-factorial response promoting the survival of infected myeloid progenitors

    doi:

    Figure Lengend Snippet: HCMV infection blocks cisplatin A mediated activation of Bak.

    Article Snippet: Cisplatin A (25uM-250uM; SIGMA-Aldrich, St Louis, MO) or 0.1% DMSO (mock) control was added for 4 hours to trigger the apoptotic pathway.

    Techniques: Infection, Activation Assay

    Effect of bortezomib on tumor accumulation of cisplatin in vivo . A , scattergram of nagogram platinum per milligram tumor wet weight as a function of tumor wet weight +, no prior bortezomib; +, 1mg/kg bortezomib i.p. 4 h prior to injection of cisplatin by the i.p. route. B , mean nanogram platinum per milligram tumor wet weight. Vertical bars, SE.

    Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

    Article Title: Enhanced Delivery of Cisplatin to Intraperitoneal Ovarian Carcinomas Mediated by the Effects of Bortezomib on the Human Copper Transporter 1

    doi: 10.1158/1078-0432.CCR-08-2081

    Figure Lengend Snippet: Effect of bortezomib on tumor accumulation of cisplatin in vivo . A , scattergram of nagogram platinum per milligram tumor wet weight as a function of tumor wet weight +, no prior bortezomib; +, 1mg/kg bortezomib i.p. 4 h prior to injection of cisplatin by the i.p. route. B , mean nanogram platinum per milligram tumor wet weight. Vertical bars, SE.

    Article Snippet: Cisplatin (Platinol) was a gift from Bristol-Myers Squibb.

    Techniques: In Vivo, Injection

    Establishment of txr cells A–C. Cell viability in response to taxol, cisplatin, and vincristine. D–F. Apoptotic sub-G1 cells in response to chemotherapeutic drugs. G–I. Caspase activation in response to taxol, cisplatin, and vincristine. The experiments were performed in triplicate (* p

    Journal: Oncotarget

    Article Title: Integrative transcriptomics-based identification of cryptic drivers of taxol-resistance genes in ovarian carcinoma cells: Analysis of the androgen receptor

    doi:

    Figure Lengend Snippet: Establishment of txr cells A–C. Cell viability in response to taxol, cisplatin, and vincristine. D–F. Apoptotic sub-G1 cells in response to chemotherapeutic drugs. G–I. Caspase activation in response to taxol, cisplatin, and vincristine. The experiments were performed in triplicate (* p

    Article Snippet: The chemotherapeutic drugs used included vincristine, taxol (paclitaxel), and cisplatin (Bristol-Myers Squibb, New York, NY, USA).

    Techniques: Activation Assay

    Antioxidant effect of NY and PS on cisplatin-induced cell death in HK-2 cells. Cells were treated with 10 μM cisplatin for 30 m and the level of ROS was detected by microplate reader ( a ). Cells were exposed to cisplatin for 24 h with or without PS or NY ( b ). The measures of JC-1 fluorescence intensity ( c and d ). Cells were treated to bind Annexin V and then to undergo PI staining; exposed to cisplatin for 24 h with or without PS or NY. HK-2 cells were treated with cisplatin for 24 h to measure apoptosis ( e ). Cell lysates were assayed for caspase-9, 3 activity. The statistical significance (* p

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: Antioxidant effect of NY and PS on cisplatin-induced cell death in HK-2 cells. Cells were treated with 10 μM cisplatin for 30 m and the level of ROS was detected by microplate reader ( a ). Cells were exposed to cisplatin for 24 h with or without PS or NY ( b ). The measures of JC-1 fluorescence intensity ( c and d ). Cells were treated to bind Annexin V and then to undergo PI staining; exposed to cisplatin for 24 h with or without PS or NY. HK-2 cells were treated with cisplatin for 24 h to measure apoptosis ( e ). Cell lysates were assayed for caspase-9, 3 activity. The statistical significance (* p

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques: Fluorescence, Staining, Activity Assay

    Herbal medicines ameliorated cisplatin-induced kidney injury markers. Mice were given an intraperitoneal injection (i.p.) of cisplatin (20 mg/kg) for 3 days. After Nelumbo nymphaea (NY) , Paeonia suffruticosa (PS) given an orally administration (300 mg/kg) for 3 days. Concentration of NGAL and KIM-1 were measured in urine and serum ( a – d ). All the values are expressed as mean ± S.E.M., n = 3 in each group. *, ** and *** statistically different from cisplatin at p

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: Herbal medicines ameliorated cisplatin-induced kidney injury markers. Mice were given an intraperitoneal injection (i.p.) of cisplatin (20 mg/kg) for 3 days. After Nelumbo nymphaea (NY) , Paeonia suffruticosa (PS) given an orally administration (300 mg/kg) for 3 days. Concentration of NGAL and KIM-1 were measured in urine and serum ( a – d ). All the values are expressed as mean ± S.E.M., n = 3 in each group. *, ** and *** statistically different from cisplatin at p

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques: Mouse Assay, Injection, Concentration Assay

    Cisplatin was induced up to cell death and kidney injury biomarkers in HK2 cells. Cell viability of HK-2 cells was treated with cisplatin low dose-long time manner for 24 h ( a ), and high dose-short time treatments for 2 to 6 h ( b ). Release of LDH ( c ) and HMGB1 ( d ) were treated with cisplatin (400 μM) for 2 to 6 h. Secretion levels of NGAL ( e ), KIM-1 ( f , g ) and HMGB1 ( h ) for 6 to 48 h. The statistical significance (* p

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: Cisplatin was induced up to cell death and kidney injury biomarkers in HK2 cells. Cell viability of HK-2 cells was treated with cisplatin low dose-long time manner for 24 h ( a ), and high dose-short time treatments for 2 to 6 h ( b ). Release of LDH ( c ) and HMGB1 ( d ) were treated with cisplatin (400 μM) for 2 to 6 h. Secretion levels of NGAL ( e ), KIM-1 ( f , g ) and HMGB1 ( h ) for 6 to 48 h. The statistical significance (* p

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques:

    Herbal medicines regulated NGAL and KIM-1 biomarkers. Pretreatment with NAC 10 mM for 1 h and stimulated with cisplatin (10 μM) for 24 h ( a ). Cytotoxicity was measured by MTT assay treatment with herbal medicine and cisplatin for 24 h ( b ). NGAL and KIM-1 were measured by ELISA ( c and d ). Lane 1:; Artemisia capillaris , Lane 2:; Houttuynia cordata , Lane 3:; Leonurus japonicas , Lane 4:; Nelumbo nymphaea , Lane 5:; Schisandra chinensis , Lane 6:; Akebia quinata , Lane 7:; Ligustrum japonicus , Lane 8:; Paeonia suffruticosa , Lane 9:; Phellodendron amurense , Lane 10:; Trichosanthes kirilowii . The statistical significance (* p

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: Herbal medicines regulated NGAL and KIM-1 biomarkers. Pretreatment with NAC 10 mM for 1 h and stimulated with cisplatin (10 μM) for 24 h ( a ). Cytotoxicity was measured by MTT assay treatment with herbal medicine and cisplatin for 24 h ( b ). NGAL and KIM-1 were measured by ELISA ( c and d ). Lane 1:; Artemisia capillaris , Lane 2:; Houttuynia cordata , Lane 3:; Leonurus japonicas , Lane 4:; Nelumbo nymphaea , Lane 5:; Schisandra chinensis , Lane 6:; Akebia quinata , Lane 7:; Ligustrum japonicus , Lane 8:; Paeonia suffruticosa , Lane 9:; Phellodendron amurense , Lane 10:; Trichosanthes kirilowii . The statistical significance (* p

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques: MTT Assay, Enzyme-linked Immunosorbent Assay

    Fold-change of kidney injury biomarkers after high- and low-dose cisplatin treatment. HMGB1 ( a ), NGAL ( b ) and KIM-1 ( c ) levels are shown for cells treated with 400 μM cisplatin for a short exposure time (2 to 6 h). HMGB ( d ), NGAL ( e ) and KIM-1 ( f ) levels were measured in cells treated with 10 μM cisplatin and a longer exposure time (6 to 48 h). The data were compared with the control condition

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: Fold-change of kidney injury biomarkers after high- and low-dose cisplatin treatment. HMGB1 ( a ), NGAL ( b ) and KIM-1 ( c ) levels are shown for cells treated with 400 μM cisplatin for a short exposure time (2 to 6 h). HMGB ( d ), NGAL ( e ) and KIM-1 ( f ) levels were measured in cells treated with 10 μM cisplatin and a longer exposure time (6 to 48 h). The data were compared with the control condition

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques:

    HMGB1 was regulated by herbal medicines with high dose-treated cisplatin in HK-2 cells. HK-2 cells were treated with high – dose of cisplatin (400 μM) for 6 h. Herbal medicines decreased LDH ( a ), and HMGB1 ( b ). The levels of LDH, HMGB1 were measured by ELISA assay kits

    Journal: BMC Complementary and Alternative Medicine

    Article Title: Assessing the recovery from prerenal and renal acute kidney injury after treatment with single herbal medicine via activity of the biomarkers HMGB1, NGAL and KIM-1 in kidney proximal tubular cells treated by cisplatin with different doses and exposure times

    doi: 10.1186/s12906-017-2055-y

    Figure Lengend Snippet: HMGB1 was regulated by herbal medicines with high dose-treated cisplatin in HK-2 cells. HK-2 cells were treated with high – dose of cisplatin (400 μM) for 6 h. Herbal medicines decreased LDH ( a ), and HMGB1 ( b ). The levels of LDH, HMGB1 were measured by ELISA assay kits

    Article Snippet: ROS, JC-1 monomers detection, caspase activation and Annexin V binding PI staining HK-2 cells were seeded in 96 well plate (1 × 104 cells/well) overnight and treated with cisplatin, cisplatin + Nelumbo nymphaea (NY), Paeonia suffruticosa (PS) for 30 min. ROS production assays was stained by DCFDA cellular ROS detection assay kit according to the manufacture instruction (Abcam, Cambridge, USA, ab113851).

    Techniques: Enzyme-linked Immunosorbent Assay

    miR-449 is promoted by cisplatin treatment and successfully inhibited by its sponge in NRK-52E cells. qPCR shows that cisplatin treatment significantly elevates the expression of miR-449 compared to untreated cells, and that miR-449 is significantly down-regulated by its sponge transfection compared to the transfection control. qPCR detection is performed at 48 h after transfection (n=5). Control=NRK-52E cells without any treatment. Cisplatin=cells treated with cisplatin. Cisplatin + blank vector=cells treated with cisplatin and then transfected with blank vector as a transfection control. Cisplatin + miR-449 sponge=cells treated with cisplatin and then transfected with miR-449 sponge. * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Inhibiting microRNA-449 Attenuates Cisplatin-Induced Injury in NRK-52E Cells Possibly via Regulating the SIRT1/P53/BAX Pathway

    doi: 10.12659/MSM.897187

    Figure Lengend Snippet: miR-449 is promoted by cisplatin treatment and successfully inhibited by its sponge in NRK-52E cells. qPCR shows that cisplatin treatment significantly elevates the expression of miR-449 compared to untreated cells, and that miR-449 is significantly down-regulated by its sponge transfection compared to the transfection control. qPCR detection is performed at 48 h after transfection (n=5). Control=NRK-52E cells without any treatment. Cisplatin=cells treated with cisplatin. Cisplatin + blank vector=cells treated with cisplatin and then transfected with blank vector as a transfection control. Cisplatin + miR-449 sponge=cells treated with cisplatin and then transfected with miR-449 sponge. * P

    Article Snippet: Cells of the cisplatin group were treated with 20 μg/mL cisplatin (Abcam, Cambridge, UK) for 24 h before cell transfection.

    Techniques: Real-time Polymerase Chain Reaction, Expressing, Transfection, Plasmid Preparation

    miR-449 regulates SIRT1, p53 acetylation, and BAX in cisplatin-treated NRK-52E cells. Cisplatin induces the down-regulation of SIRT1 and the up-regulation of acetylated p53 and BAX, while inhibiting miR-449 in the treated cells promotes SIRT1 expression and suppresses acetylated p53 and BAX levels. Western blot analysis was performed to detect protein levels, with GAPDH as an internal control ( A ), and a histogram is generated based on triplicate Western blot experiments ( B ). ** P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Inhibiting microRNA-449 Attenuates Cisplatin-Induced Injury in NRK-52E Cells Possibly via Regulating the SIRT1/P53/BAX Pathway

    doi: 10.12659/MSM.897187

    Figure Lengend Snippet: miR-449 regulates SIRT1, p53 acetylation, and BAX in cisplatin-treated NRK-52E cells. Cisplatin induces the down-regulation of SIRT1 and the up-regulation of acetylated p53 and BAX, while inhibiting miR-449 in the treated cells promotes SIRT1 expression and suppresses acetylated p53 and BAX levels. Western blot analysis was performed to detect protein levels, with GAPDH as an internal control ( A ), and a histogram is generated based on triplicate Western blot experiments ( B ). ** P

    Article Snippet: Cells of the cisplatin group were treated with 20 μg/mL cisplatin (Abcam, Cambridge, UK) for 24 h before cell transfection.

    Techniques: Expressing, Western Blot, Generated

    Inhibiting miR-449 promotes NRK-52E cell viability and suppresses cell apoptosis induced by cisplatin. MTT and flow cytometry were used to detect cell viability ( A ) and cell apoptosis ( B ), respectively, at 48 h after transfection (n=5). Cisplatin treatment inhibits cell viability and promotes cell apoptosis, which is abrogated by miR-449 sponge. * P

    Journal: Medical Science Monitor : International Medical Journal of Experimental and Clinical Research

    Article Title: Inhibiting microRNA-449 Attenuates Cisplatin-Induced Injury in NRK-52E Cells Possibly via Regulating the SIRT1/P53/BAX Pathway

    doi: 10.12659/MSM.897187

    Figure Lengend Snippet: Inhibiting miR-449 promotes NRK-52E cell viability and suppresses cell apoptosis induced by cisplatin. MTT and flow cytometry were used to detect cell viability ( A ) and cell apoptosis ( B ), respectively, at 48 h after transfection (n=5). Cisplatin treatment inhibits cell viability and promotes cell apoptosis, which is abrogated by miR-449 sponge. * P

    Article Snippet: Cells of the cisplatin group were treated with 20 μg/mL cisplatin (Abcam, Cambridge, UK) for 24 h before cell transfection.

    Techniques: MTT Assay, Flow Cytometry, Cytometry, Transfection

    Phendione inhibits the proliferation of melanoma with intrinsic resistance to MAPK inhibitors. ( A–E ) Panels show growth inhibition curves and IC50 values. Primary patient-derived melanoma cell lines with BRAF V600E , BRAF V600K , and NRAS Q61R mutations ( A–C ), and human melanoma cell line SK-MEL28 ( D ) and its BRAF inhibitor (dabrafenib)-resistant line SK-MEL28R ( E ) were treated with phendione or inhibitors of the MAPK pathway for 96 h. Data are presented as mean ± SD of three independent experiments. ( F ) Cell cycle analysis for A375 BRAFi-resistant cells. Cells were treated with 2.5 µM cisplatin, 0.2 µM phendione, or vehicle for the indicated time. Graph shows the percentage of cells in each phase of the cell cycle. Data shown are average ± SD of three independent experiments. ( G ) Comet assay to measure DNA damage. Representative images are shown at the left and quantification results are shown at the right . Data are presented as mean ± SD of three independent experiments. ( H ) Immunoblots of whole-cell lysate collected from comet assay. ( I , J ) Quantification of reactive oxygen species (ROS) production measured by flow cytometry in A375 ( I ) and A375 BRAFi-resistant ( J ) cells. Hydroxyl peroxide (H 2 O 2 ; 200 µM) was used as positive control. Data are presented as means ± SEMs of three independent experiments. (*) P

    Journal: Genes & Development

    Article Title: Targeted chemotherapy overcomes drug resistance in melanoma

    doi: 10.1101/gad.333864.119

    Figure Lengend Snippet: Phendione inhibits the proliferation of melanoma with intrinsic resistance to MAPK inhibitors. ( A–E ) Panels show growth inhibition curves and IC50 values. Primary patient-derived melanoma cell lines with BRAF V600E , BRAF V600K , and NRAS Q61R mutations ( A–C ), and human melanoma cell line SK-MEL28 ( D ) and its BRAF inhibitor (dabrafenib)-resistant line SK-MEL28R ( E ) were treated with phendione or inhibitors of the MAPK pathway for 96 h. Data are presented as mean ± SD of three independent experiments. ( F ) Cell cycle analysis for A375 BRAFi-resistant cells. Cells were treated with 2.5 µM cisplatin, 0.2 µM phendione, or vehicle for the indicated time. Graph shows the percentage of cells in each phase of the cell cycle. Data shown are average ± SD of three independent experiments. ( G ) Comet assay to measure DNA damage. Representative images are shown at the left and quantification results are shown at the right . Data are presented as mean ± SD of three independent experiments. ( H ) Immunoblots of whole-cell lysate collected from comet assay. ( I , J ) Quantification of reactive oxygen species (ROS) production measured by flow cytometry in A375 ( I ) and A375 BRAFi-resistant ( J ) cells. Hydroxyl peroxide (H 2 O 2 ; 200 µM) was used as positive control. Data are presented as means ± SEMs of three independent experiments. (*) P

    Article Snippet: Vemurafenib (S1267), trametinib (S2673), PD0325901 (S1036), SCH772984 (S7101), etoposide (S1225), and cisplatin (S1166) were purchased from Selleck Chemicals.

    Techniques: Inhibition, Derivative Assay, Cell Cycle Assay, Single Cell Gel Electrophoresis, Western Blot, Flow Cytometry, Positive Control

    Effect of CDDP on the proportion of cells in each stage of the cell cycle in (A) untreated SP, (B) CDDP-treated SP, (C) untreated non-SP and (D) CDDP-treated non-SP cells, as detected by flow cytometry. ‘Dip’ refers to the ratio of cells during each cell cycle phase. SP, side population; G 1 , gap 1; G 2 , gap 2; S, synthesis; CDDP, cisplatin.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Isolation and characterization of side population cells from the human ovarian cancer cell line SK-OV-3

    doi: 10.3892/etm.2015.2836

    Figure Lengend Snippet: Effect of CDDP on the proportion of cells in each stage of the cell cycle in (A) untreated SP, (B) CDDP-treated SP, (C) untreated non-SP and (D) CDDP-treated non-SP cells, as detected by flow cytometry. ‘Dip’ refers to the ratio of cells during each cell cycle phase. SP, side population; G 1 , gap 1; G 2 , gap 2; S, synthesis; CDDP, cisplatin.

    Article Snippet: Cisplatin (CDDP) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: Flow Cytometry, Cytometry

    Effect of various CDDP doses on viability of SP and non-SP cells. SP, side population; CDDP, cisplatin.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Isolation and characterization of side population cells from the human ovarian cancer cell line SK-OV-3

    doi: 10.3892/etm.2015.2836

    Figure Lengend Snippet: Effect of various CDDP doses on viability of SP and non-SP cells. SP, side population; CDDP, cisplatin.

    Article Snippet: Cisplatin (CDDP) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques:

    Autophagy activation promotes hair cell survival after cisplatin-induced damage in zebrafish lateral line. (A) Hair cell counts obtained from CDDP exposure for 12 or 24 h with or without RA or SRT1720 treatment ( n = 6 zebrafish larvae). Scale bar, 10 μm. (B) The hair cells were counted at 12 h after CDDP exposure. (C) The hair cells were counted 24 h after CDDP exposure. Data represent the mean ± SEM. *** p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: Autophagy activation promotes hair cell survival after cisplatin-induced damage in zebrafish lateral line. (A) Hair cell counts obtained from CDDP exposure for 12 or 24 h with or without RA or SRT1720 treatment ( n = 6 zebrafish larvae). Scale bar, 10 μm. (B) The hair cells were counted at 12 h after CDDP exposure. (C) The hair cells were counted 24 h after CDDP exposure. Data represent the mean ± SEM. *** p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: Activation Assay

    SIRT1 activation attenuates cisplatin-induced hair cell loss and hearing loss in C57BL/6 mice. (A) ABR thresholds decreased with SRT1720 (100 mg/kg, intragastric administration pre-12 h, pre-1 h, post-12 h) treatment mice following CDDP (16 mg/kg, intraperitoneal injection) exposure compared with the CDDP group at 4, 8, 16 and 32 kHz ( n = 6 mice). (B,C) Surface preparations were stained with DAPI. Hair cell counts obtained from the CDDP and CDDP+SRT1720 group ( n = the right cochlea of six mice). Scale bar, 10 μm. Data represent the mean ± SEM. * p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: SIRT1 activation attenuates cisplatin-induced hair cell loss and hearing loss in C57BL/6 mice. (A) ABR thresholds decreased with SRT1720 (100 mg/kg, intragastric administration pre-12 h, pre-1 h, post-12 h) treatment mice following CDDP (16 mg/kg, intraperitoneal injection) exposure compared with the CDDP group at 4, 8, 16 and 32 kHz ( n = 6 mice). (B,C) Surface preparations were stained with DAPI. Hair cell counts obtained from the CDDP and CDDP+SRT1720 group ( n = the right cochlea of six mice). Scale bar, 10 μm. Data represent the mean ± SEM. * p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: Activation Assay, Mouse Assay, Injection, Staining

    Rapamycin promotes HEI-OC1 cell survival after cisplatin-induced damage. (A) The CCK8 assay was performed to examine cell viability of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3 individual experiments). (B) The image of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3 individual experiments). (C,D) Western blots and densitometry analysis for autophagy marker LC3-II and p62 in CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3). Data represent the mean ± SEM. * p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: Rapamycin promotes HEI-OC1 cell survival after cisplatin-induced damage. (A) The CCK8 assay was performed to examine cell viability of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3 individual experiments). (B) The image of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3 individual experiments). (C,D) Western blots and densitometry analysis for autophagy marker LC3-II and p62 in CDDP (20 μM) exposure for 24 h with or without RA (0.5 μM; n = 3). Data represent the mean ± SEM. * p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: CCK-8 Assay, Western Blot, Marker

    SIRT1 activates autophagy and promotes HEI-OC1 cell survival after cisplatin-induced damage. (A) The CCK8 assay was performed to examine cell viability of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 3 individual experiments). (B,C) The fluorescence image of green fluorescent protein (GFP)-LC3 HEI-OC1 cells after CDDP (20 μM) exposure with or without SRT1720 (0.5 μM) and CQ (10 μM). Scale bar, 10 μm. Quantity analysis of green puncta was detected in five cells/experiment ( n = 3 individual experiments). (D,E) Western blots and densitometry analysis for SIRT1 and autophagy marker LC3-II and p62 in CDDP (20 μM) exposure for 24 h with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 3 individual experiments). Data represent the mean ± SEM. * p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: SIRT1 activates autophagy and promotes HEI-OC1 cell survival after cisplatin-induced damage. (A) The CCK8 assay was performed to examine cell viability of HEI-OC1 cells following CDDP (20 μM) exposure for 24 h with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 3 individual experiments). (B,C) The fluorescence image of green fluorescent protein (GFP)-LC3 HEI-OC1 cells after CDDP (20 μM) exposure with or without SRT1720 (0.5 μM) and CQ (10 μM). Scale bar, 10 μm. Quantity analysis of green puncta was detected in five cells/experiment ( n = 3 individual experiments). (D,E) Western blots and densitometry analysis for SIRT1 and autophagy marker LC3-II and p62 in CDDP (20 μM) exposure for 24 h with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 3 individual experiments). Data represent the mean ± SEM. * p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: CCK-8 Assay, Fluorescence, Western Blot, Marker

    SIRT1 protects against cisplatin-induced cell death via autophagy in HEI-OC1 cells. The CCK8 assay was performed to examine cell viability of HEI-OC1 cells in CDDP (20 μM) exposure for 24 h combined with 3-MA (5 mM) treatment with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 4 individual experiments). Data represent the mean ± SEM. *** p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: SIRT1 protects against cisplatin-induced cell death via autophagy in HEI-OC1 cells. The CCK8 assay was performed to examine cell viability of HEI-OC1 cells in CDDP (20 μM) exposure for 24 h combined with 3-MA (5 mM) treatment with or without SRT1720 pre-treatment for 24 h (0.5 μM; n = 4 individual experiments). Data represent the mean ± SEM. *** p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: CCK-8 Assay

    Autophagy activation attenuates cisplatin-induced hair cell loss and hearing loss in C57BL/6 mice. (A) Auditory brainstem response (ABR) thresholds decreased with RA (7.5 mg/kg, intraperitoneal injection pre-24 h, pre-1 h, post-24 h) treatment mice in CDDP (16 mg/kg, intraperitoneal injection) exposure compared with the CDDP groups at 4, 8, 16 and 32 kHz ( n = 6 mice). (B,C) Surface preparations were stained with 4′,6-diamidino-2-phenylindole (DAPI). Hair cell counts obtained for the CDDP and CDDP+RA group ( n = the right cochlea of six mice). Scale bar, 10 μm. Data represent the mean ± SEM. * p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: Sirtuin 1 and Autophagy Attenuate Cisplatin-Induced Hair Cell Death in the Mouse Cochlea and Zebrafish Lateral Line

    doi: 10.3389/fncel.2018.00515

    Figure Lengend Snippet: Autophagy activation attenuates cisplatin-induced hair cell loss and hearing loss in C57BL/6 mice. (A) Auditory brainstem response (ABR) thresholds decreased with RA (7.5 mg/kg, intraperitoneal injection pre-24 h, pre-1 h, post-24 h) treatment mice in CDDP (16 mg/kg, intraperitoneal injection) exposure compared with the CDDP groups at 4, 8, 16 and 32 kHz ( n = 6 mice). (B,C) Surface preparations were stained with 4′,6-diamidino-2-phenylindole (DAPI). Hair cell counts obtained for the CDDP and CDDP+RA group ( n = the right cochlea of six mice). Scale bar, 10 μm. Data represent the mean ± SEM. * p

    Article Snippet: Materials Cisplatin (CDDP, Selleck, S1166, Huston, TX, USA), Rapamycin (RA, Selleck, S1039, TX, USA), 3-Methyladenine (3-MA, S2767, Selleck, Huston, TX, USA), SRT1720 (SRT1720, S1129, Selleck, Huston, TX, USA).

    Techniques: Activation Assay, Mouse Assay, Injection, Staining

    Activation of NF- κ B signaling by BST2 is required for cisplatin resistance in NPC cells. ( a ) NF- κ B activity assay. NPC cells were transfected with BST2 siRNAs (si1, si2) or cDNA (BST2) for 24 h followed by a luciferase reporter assay, as described in Materials and Methods. ( b ) WB assays for total I κ B α , phosphor-I κ B α (p-I κ B α ) and nuclear p65 levels (GAPDH and histone H3 were used as loading controls) in CNE2 cells transfected BST2 siRNAs or S16 cells transfected with BST2 cDNA for 48 h and treated with 10 μ M cisplatin for 24 h. ( c ) Effects of WT BST2 and BST2 mutants (ΔGPI and Y6,8A) on cisplatin resistance were investigated by MTT assays in S16 cells stably overexpressing WT BST2 or BST2 mutants (left, WB assay for BST2 expression; middle, representative dose-dependent cell viability curves; right, relative resistant factors). ( d ) inhibition of I κ B α phosphorylation reverses BST2-induced anti-apoptosis. After 24 h of treatment with 10 μ M cisplatin plus 10 μ M BAY11-7085 (DMSO was used as a vehicle control), apoptosis was analyzed by Annexin-V/PI dual staining (left, representative dot plots for flow cytometry; right, the percentages of apoptotic cells) in S16 cells stably expressing BST2. NC, negative control siRNA; si1, si2, BST2 siRNAs; Vec, empty vector-tranfected; BST2, BST2 overexpression;. * P

    Journal: Cell Death & Disease

    Article Title: BST2 confers cisplatin resistance via NF-κB signaling in nasopharyngeal cancer

    doi: 10.1038/cddis.2017.271

    Figure Lengend Snippet: Activation of NF- κ B signaling by BST2 is required for cisplatin resistance in NPC cells. ( a ) NF- κ B activity assay. NPC cells were transfected with BST2 siRNAs (si1, si2) or cDNA (BST2) for 24 h followed by a luciferase reporter assay, as described in Materials and Methods. ( b ) WB assays for total I κ B α , phosphor-I κ B α (p-I κ B α ) and nuclear p65 levels (GAPDH and histone H3 were used as loading controls) in CNE2 cells transfected BST2 siRNAs or S16 cells transfected with BST2 cDNA for 48 h and treated with 10 μ M cisplatin for 24 h. ( c ) Effects of WT BST2 and BST2 mutants (ΔGPI and Y6,8A) on cisplatin resistance were investigated by MTT assays in S16 cells stably overexpressing WT BST2 or BST2 mutants (left, WB assay for BST2 expression; middle, representative dose-dependent cell viability curves; right, relative resistant factors). ( d ) inhibition of I κ B α phosphorylation reverses BST2-induced anti-apoptosis. After 24 h of treatment with 10 μ M cisplatin plus 10 μ M BAY11-7085 (DMSO was used as a vehicle control), apoptosis was analyzed by Annexin-V/PI dual staining (left, representative dot plots for flow cytometry; right, the percentages of apoptotic cells) in S16 cells stably expressing BST2. NC, negative control siRNA; si1, si2, BST2 siRNAs; Vec, empty vector-tranfected; BST2, BST2 overexpression;. * P

    Article Snippet: For the Annexin-V/PI binding assay, NPC cells transiently transfected with siRNAs or stably overexpressing BST2 were exposed to 10 μ M cisplatin with or without 10 μ M BAY11-7085 (Selleck, Houston, TX, USA) for 24 h; floating and adherent cells were then collected by centrifugation and trypsinization respectively.

    Techniques: Activation Assay, Activity Assay, Transfection, Luciferase, Reporter Assay, Western Blot, MTT Assay, Stable Transfection, Expressing, Inhibition, Staining, Flow Cytometry, Cytometry, Negative Control, Plasmid Preparation, Over Expression

    BST2 knockdown reverses cisplatin resistance of NPC cells in xenograft tumors in nude mice. CNE2 cells stably expressing two specific BST2 shRNAs (sh1, sh2) or scrambled control shRNA (shNC) were subcutaneously injected to generate xenograft tumors in nude mice; cDDP treatment was performed as described in Materials and Methods. Normal saline (n.s.) was used as the treatment control. ( a ) Western blot analysis for BST2 expression in stable cell lines. ( b ) Growth curves of tumor xenografts. ( c ) Images of xenograft tumors harvested at the end of the experiment. ( d ) The weights of tumors are presented as a Cleveland dot plot, and the average±S.D. is included ( n =6/group; ** P

    Journal: Cell Death & Disease

    Article Title: BST2 confers cisplatin resistance via NF-κB signaling in nasopharyngeal cancer

    doi: 10.1038/cddis.2017.271

    Figure Lengend Snippet: BST2 knockdown reverses cisplatin resistance of NPC cells in xenograft tumors in nude mice. CNE2 cells stably expressing two specific BST2 shRNAs (sh1, sh2) or scrambled control shRNA (shNC) were subcutaneously injected to generate xenograft tumors in nude mice; cDDP treatment was performed as described in Materials and Methods. Normal saline (n.s.) was used as the treatment control. ( a ) Western blot analysis for BST2 expression in stable cell lines. ( b ) Growth curves of tumor xenografts. ( c ) Images of xenograft tumors harvested at the end of the experiment. ( d ) The weights of tumors are presented as a Cleveland dot plot, and the average±S.D. is included ( n =6/group; ** P

    Article Snippet: For the Annexin-V/PI binding assay, NPC cells transiently transfected with siRNAs or stably overexpressing BST2 were exposed to 10 μ M cisplatin with or without 10 μ M BAY11-7085 (Selleck, Houston, TX, USA) for 24 h; floating and adherent cells were then collected by centrifugation and trypsinization respectively.

    Techniques: Mouse Assay, Stable Transfection, Expressing, shRNA, Injection, Western Blot

    Effect of BST2 on cisplatin-induced apoptosis. NPC cells transiently transfected with siRNAs for 24 h or cells stably overexpressing BST2 were treated with cisplatin (cDDP) for 24 h, and apoptosis was analyzed. The experiments were independently repeated for 3–4 times. ( a , c ) Annexin-V-FITC/PI dual staining assay (left, representative plots for flow cytometry; right, bar charts indicating the average percentages of apoptotic cells). ( b , d ) TUNEL assays (left, representative immunofluorescent pictures; right, bar charts indicating the average percentages of apoptotic cells). ( e ) WB assays (c-PARP, cleaved PARP; GAPDH, a loading control). NC, negative control siRNA; si1, si2, BST2 siRNAs; Vec, empty vector transfected; BST2, BST2 overexpression; n.s., normal saline. * P

    Journal: Cell Death & Disease

    Article Title: BST2 confers cisplatin resistance via NF-κB signaling in nasopharyngeal cancer

    doi: 10.1038/cddis.2017.271

    Figure Lengend Snippet: Effect of BST2 on cisplatin-induced apoptosis. NPC cells transiently transfected with siRNAs for 24 h or cells stably overexpressing BST2 were treated with cisplatin (cDDP) for 24 h, and apoptosis was analyzed. The experiments were independently repeated for 3–4 times. ( a , c ) Annexin-V-FITC/PI dual staining assay (left, representative plots for flow cytometry; right, bar charts indicating the average percentages of apoptotic cells). ( b , d ) TUNEL assays (left, representative immunofluorescent pictures; right, bar charts indicating the average percentages of apoptotic cells). ( e ) WB assays (c-PARP, cleaved PARP; GAPDH, a loading control). NC, negative control siRNA; si1, si2, BST2 siRNAs; Vec, empty vector transfected; BST2, BST2 overexpression; n.s., normal saline. * P

    Article Snippet: For the Annexin-V/PI binding assay, NPC cells transiently transfected with siRNAs or stably overexpressing BST2 were exposed to 10 μ M cisplatin with or without 10 μ M BAY11-7085 (Selleck, Houston, TX, USA) for 24 h; floating and adherent cells were then collected by centrifugation and trypsinization respectively.

    Techniques: Transfection, Stable Transfection, Staining, Flow Cytometry, Cytometry, TUNEL Assay, Western Blot, Negative Control, Plasmid Preparation, Over Expression

    Contributions of BST2 to cisplatin resistance in nasopharyngeal cancer (NPC) cells. ( a ) S16 cells stably overexpressing BST2 or ( b ) CNE2, ( c ) HONE1 and ( d ) HNE1 cells transfected with BST2 siRNAs for 24 h were subjected to MTT assays, as described in Materials and Methods (Vec: empty vector transfected; BST2: BST2 overexpression; si1, si2, NC: BST2 siRNAs or negative control siRNA transfected). Left: western blotting (WB) analysis of BST2 expression at 48 h after siRNA transfection; actin or GAPDH was used as a loading control. Middle: representative dose-dependent cell viability curves. Right: averages of relative resistant factors (RRFs, the ratios of IC 50 value of experimental cells to that of control cells) in 3–5 independent experiments (** P

    Journal: Cell Death & Disease

    Article Title: BST2 confers cisplatin resistance via NF-κB signaling in nasopharyngeal cancer

    doi: 10.1038/cddis.2017.271

    Figure Lengend Snippet: Contributions of BST2 to cisplatin resistance in nasopharyngeal cancer (NPC) cells. ( a ) S16 cells stably overexpressing BST2 or ( b ) CNE2, ( c ) HONE1 and ( d ) HNE1 cells transfected with BST2 siRNAs for 24 h were subjected to MTT assays, as described in Materials and Methods (Vec: empty vector transfected; BST2: BST2 overexpression; si1, si2, NC: BST2 siRNAs or negative control siRNA transfected). Left: western blotting (WB) analysis of BST2 expression at 48 h after siRNA transfection; actin or GAPDH was used as a loading control. Middle: representative dose-dependent cell viability curves. Right: averages of relative resistant factors (RRFs, the ratios of IC 50 value of experimental cells to that of control cells) in 3–5 independent experiments (** P

    Article Snippet: For the Annexin-V/PI binding assay, NPC cells transiently transfected with siRNAs or stably overexpressing BST2 were exposed to 10 μ M cisplatin with or without 10 μ M BAY11-7085 (Selleck, Houston, TX, USA) for 24 h; floating and adherent cells were then collected by centrifugation and trypsinization respectively.

    Techniques: Stable Transfection, Transfection, MTT Assay, Plasmid Preparation, Over Expression, Negative Control, Western Blot, Expressing

    CAF-derived exosomal miR-98-5p stimulates cisplatin resistance in OC cells by targeting CDKN1A. A2780 cells transfected with oe-CDKN1A or oe-NC were co-cultured with CAF-exo and further treated with cisplatin.  a  Cell proliferation as detected by CCK-8 assay.  b  OC cell colony formation ability as detected by colony formation assay.  c  Statistical analysis for  b .  d  OC cell cycle distribution as detected by flow cytometry.  e  Statistical analysis for  d .  f  OC cell apoptosis as detected by flow cytometry.  g  Statistical analysis for  f . * p

    Journal: Cancer Cell International

    Article Title: Cancer-associated fibroblast-derived exosomal microRNA-98-5p promotes cisplatin resistance in ovarian cancer by targeting CDKN1A

    doi: 10.1186/s12935-019-1051-3

    Figure Lengend Snippet: CAF-derived exosomal miR-98-5p stimulates cisplatin resistance in OC cells by targeting CDKN1A. A2780 cells transfected with oe-CDKN1A or oe-NC were co-cultured with CAF-exo and further treated with cisplatin. a Cell proliferation as detected by CCK-8 assay. b OC cell colony formation ability as detected by colony formation assay. c Statistical analysis for b . d OC cell cycle distribution as detected by flow cytometry. e Statistical analysis for d . f OC cell apoptosis as detected by flow cytometry. g Statistical analysis for f . * p

    Article Snippet: Determination of half maximal inhibitory concentration (IC50) of cisplatin A total of 100 μL cell suspension (1 × 105 cells/mL) was seeded to 96-well plates, followed by treatment with cisplatin at variant concentrations (0, 1, 2, 4, 8 μg/mL; Selleck Chemicals, Houston, TX, USA) for 72 h. The sensitivity of OC cells to cisplatin was then evaluated via the CCK-8 assay, and IC50 was obtained.

    Techniques: Derivative Assay, Transfection, Cell Culture, CCK-8 Assay, Colony Assay, Flow Cytometry, Cytometry

    β-catenin-induced stemness is responsible for PAK1-mediated cisplatin resistance. ( a ) Sphere formation abilities in H23 and PAK1-overexpressing H1355 cells were identified in a low-attachment plate by incubation in serum-free media for 1 week. The photographs show sphere colonies in H23 and PAK1-overexpressing H1355 cells (upper panel). The graphs show the average number of spheres in triplicate samples (lower panel). ( b ) H23 and PAK1-overexpressing H1355 cells were transfected with a β-catenin overexpression plasmid for 24 h, followed by treatment with the MEK/ERK inhibitor AZD6244 for 5 h. The cells were then incubated for 1 week in a low-attachment plate in serum-free media and evaluated for sphere formation abilities. Expression of the cancer stem cell markers OCT4, SOX2, Nanog, and cmyc was identified using western blotting. ( c ) H23 and PAK1-overexpressing H1355 cells were treated with a cell stemness inhibitor (10 μM BBI-608) for 5 h. The inhibitor was then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was then evaluated by MTT assay. ( d ) H1355 and H23 cells were transfected with PAK1 expression vector and shPAK1 for 24 h. The cells were then treated with 0.1% DMSO or 25 μM cisplatin for 24 h and subjected to annexin-V and PI staining, followed by a flow cytometry analysis. The percentages of apoptotic cells in the annexin V+/PI- and the annexin-V+/PI+ populations were determined. ( e ) H23 and PAK1-overexpressing H1355 cells were transfected with β-catenin overexpression plasmid for 24 h, followed by treatment with AZD6244 for 5 h. The inhibitor was then removed and the cells were treated with cisplatin for an additional 48 h. ( f ) H23 and PAK1-overexpressing H1355 cells were treated with a cell stemness inhibitor (10 μM BBI-608) for 5 h. The inhibitor was then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell apoptosis was evaluated by flow cytometry. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars . The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary Figure S3 .

    Journal: Scientific Reports

    Article Title: PAK1 confers chemoresistance and poor outcome in non-small cell lung cancer via β-catenin-mediated stemness

    doi: 10.1038/srep34933

    Figure Lengend Snippet: β-catenin-induced stemness is responsible for PAK1-mediated cisplatin resistance. ( a ) Sphere formation abilities in H23 and PAK1-overexpressing H1355 cells were identified in a low-attachment plate by incubation in serum-free media for 1 week. The photographs show sphere colonies in H23 and PAK1-overexpressing H1355 cells (upper panel). The graphs show the average number of spheres in triplicate samples (lower panel). ( b ) H23 and PAK1-overexpressing H1355 cells were transfected with a β-catenin overexpression plasmid for 24 h, followed by treatment with the MEK/ERK inhibitor AZD6244 for 5 h. The cells were then incubated for 1 week in a low-attachment plate in serum-free media and evaluated for sphere formation abilities. Expression of the cancer stem cell markers OCT4, SOX2, Nanog, and cmyc was identified using western blotting. ( c ) H23 and PAK1-overexpressing H1355 cells were treated with a cell stemness inhibitor (10 μM BBI-608) for 5 h. The inhibitor was then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was then evaluated by MTT assay. ( d ) H1355 and H23 cells were transfected with PAK1 expression vector and shPAK1 for 24 h. The cells were then treated with 0.1% DMSO or 25 μM cisplatin for 24 h and subjected to annexin-V and PI staining, followed by a flow cytometry analysis. The percentages of apoptotic cells in the annexin V+/PI- and the annexin-V+/PI+ populations were determined. ( e ) H23 and PAK1-overexpressing H1355 cells were transfected with β-catenin overexpression plasmid for 24 h, followed by treatment with AZD6244 for 5 h. The inhibitor was then removed and the cells were treated with cisplatin for an additional 48 h. ( f ) H23 and PAK1-overexpressing H1355 cells were treated with a cell stemness inhibitor (10 μM BBI-608) for 5 h. The inhibitor was then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell apoptosis was evaluated by flow cytometry. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars . The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary Figure S3 .

    Article Snippet: Chemicals and antibodies Cisplatin, Perifosine and AZD6244 was obtained from Selleckchem.com (Houston, TX).

    Techniques: Incubation, Transfection, Over Expression, Plasmid Preparation, Expressing, Western Blot, MTT Assay, Staining, Flow Cytometry, Cytometry, Derivative Assay

    MEK/ERK signaling plays a more important role thanPI3K/AKT signaling in mediating PAK1-mediated cisplatin resistance. ( a ) Six lung cancer cell types were treated with four concentrations of cisplatin and the dose response curves were used to calculate the 50% inhibition concentration (IC50) for cisplatin. The expression of PAK1 and its active forms (pS144-PAK1 and pT423-PAK1) in each lung cancer cell type were examined by western blotting. ( b ) Increasing amounts of PAK1 knockdown plasmid were transfected into high-PAK1 expressing (H441 and H23) cell lines. Alternatively, increasing amounts of expression plasmid were transfected into low PAK1 expressing (H358 and H1355) cell lines. The total amount of transfected DNA was kept constant by adding the control vector. After 48 hr, cell lysates were harvested and evaluated by Western blotting for levels of PAK1 and β-actin protein. β-actin was used as a protein loading control. NC: non-specific shRNA control. VC: Vector control. PAK1-knockdown or PAK1-overexpressing lung cancer cells were treated with four doses of cisplatin and the dose response curves were used to calculate the 50% inhibition concentration (IC50). ( c ) PAK1-overexpressing H358 and H1355 cells were treated for 5 h with inhibitors of PI3K/AKT (10 μM perifosine) and ERK (10 μM AZD6244). The inhibitors were then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was evaluated with the MTT assay. ( d ) H23 and PAK1-overexpressing H1355 cells were transfected with β-catenin overexpression plasmid for 24 h. The cells were then treated with AZD6244 for 5 h, the inhibitor was removed, and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was evaluated with the MTT assay. ( e ) H23 and PAK1-overexpressing H1355 cells were treated with AZD6244 for 5 h, followed by treatment with MG132 for an additional 5 h, and then the cell lysates were evaluated for protein expression by western blotting. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars . The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary Figures S2 and S3 .

    Journal: Scientific Reports

    Article Title: PAK1 confers chemoresistance and poor outcome in non-small cell lung cancer via β-catenin-mediated stemness

    doi: 10.1038/srep34933

    Figure Lengend Snippet: MEK/ERK signaling plays a more important role thanPI3K/AKT signaling in mediating PAK1-mediated cisplatin resistance. ( a ) Six lung cancer cell types were treated with four concentrations of cisplatin and the dose response curves were used to calculate the 50% inhibition concentration (IC50) for cisplatin. The expression of PAK1 and its active forms (pS144-PAK1 and pT423-PAK1) in each lung cancer cell type were examined by western blotting. ( b ) Increasing amounts of PAK1 knockdown plasmid were transfected into high-PAK1 expressing (H441 and H23) cell lines. Alternatively, increasing amounts of expression plasmid were transfected into low PAK1 expressing (H358 and H1355) cell lines. The total amount of transfected DNA was kept constant by adding the control vector. After 48 hr, cell lysates were harvested and evaluated by Western blotting for levels of PAK1 and β-actin protein. β-actin was used as a protein loading control. NC: non-specific shRNA control. VC: Vector control. PAK1-knockdown or PAK1-overexpressing lung cancer cells were treated with four doses of cisplatin and the dose response curves were used to calculate the 50% inhibition concentration (IC50). ( c ) PAK1-overexpressing H358 and H1355 cells were treated for 5 h with inhibitors of PI3K/AKT (10 μM perifosine) and ERK (10 μM AZD6244). The inhibitors were then removed and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was evaluated with the MTT assay. ( d ) H23 and PAK1-overexpressing H1355 cells were transfected with β-catenin overexpression plasmid for 24 h. The cells were then treated with AZD6244 for 5 h, the inhibitor was removed, and the cells were treated with 25 μM cisplatin for an additional 48 h. Cell viability was evaluated with the MTT assay. ( e ) H23 and PAK1-overexpressing H1355 cells were treated with AZD6244 for 5 h, followed by treatment with MG132 for an additional 5 h, and then the cell lysates were evaluated for protein expression by western blotting. All experiments were performed three independent times. The mean values and the standard deviations are indicated as columns with error bars . The samples were derived from the same experiment and gels/blots were processed in parallel. Full-length blots are presented in Supplementary Figures S2 and S3 .

    Article Snippet: Chemicals and antibodies Cisplatin, Perifosine and AZD6244 was obtained from Selleckchem.com (Houston, TX).

    Techniques: Inhibition, Concentration Assay, Expressing, Western Blot, Plasmid Preparation, Transfection, shRNA, MTT Assay, Over Expression, Derivative Assay

    The expression of CSF-1R and its ligand influence the resistance to cisplatin treatment ( A ) Cisplatin treatment increases mRNA and protein levels of CSF-1 and CSF-1R. Histograms reporting the mRNA levels of CSF-1 and CSF-1R, in cells harvested at 24 hr (quantitative PCR), and the protein levels of CSF-1 in medium conditioned for 48 hr (ELISA assay), from the indicated cell lines treated with cisplatin at the CC 50 doses. ( B ) The CSF-1R expressing cells survive chemotherapy-induced stress. Upper panel. Representative FACS dot plots showing the percentage of CSF-1R pos in H1299 cells treated with cisplatin at the CC 50 and CC 75 for 72 hrs (gated). Gated cells and isotype controls are further illustrated in Supplementary Figure S1B . Lower panel: histogram bars reporting the percentage of CSF-1R pos cells in the indicated cell lines. In the lower panel, the mean ± SE of two independent experiments is reported. ( C – D ) CSF-1R inhibition affects the clonogenicity and resistance to cisplatin of lung cancer cell lines. (C) Western blotting of CSF-1R immunoprecipitates stained with the indicated antibodies from H1299 cells treated with increasing concentrations of JNJ-40346527. (D) Upper panel. Representative micrographs of the colonies formed by H1299 cells treated with increasing doses of JNJ-40346527. Lower panel. Histograms reporting the mean ± SE of the colonies formed by 4 representative cell lines in three independent experiments. ( E ) CSF-1R inhibition affects the resistance of lung cancer cell lines to cisplatin. Graphs reporting the percentage of colonies formed by A549 and H1299 cells treated with JNJ-40346527 at the CC 25 doses determined in 2D and with the indicated doses of cisplatin. Mean ± SE of three independent experiments. ( F ) JNJ-40346527 treatment modulates the number of CSF-1R pos cells. Histograms reporting the number of CSF-1R pos cells (as assessed by FACS) in the H1299 cells treated for 96 hrs at the previously determined CC 50 dosages for both cisplatin and JNJ-40346527. Mean ± SE of three independent experiments is reported. *= p

    Journal: Oncotarget

    Article Title: Inhibition of the colony-stimulating-factor-1 receptor affects the resistance of lung cancer cells to cisplatin

    doi: 10.18632/oncotarget.10895

    Figure Lengend Snippet: The expression of CSF-1R and its ligand influence the resistance to cisplatin treatment ( A ) Cisplatin treatment increases mRNA and protein levels of CSF-1 and CSF-1R. Histograms reporting the mRNA levels of CSF-1 and CSF-1R, in cells harvested at 24 hr (quantitative PCR), and the protein levels of CSF-1 in medium conditioned for 48 hr (ELISA assay), from the indicated cell lines treated with cisplatin at the CC 50 doses. ( B ) The CSF-1R expressing cells survive chemotherapy-induced stress. Upper panel. Representative FACS dot plots showing the percentage of CSF-1R pos in H1299 cells treated with cisplatin at the CC 50 and CC 75 for 72 hrs (gated). Gated cells and isotype controls are further illustrated in Supplementary Figure S1B . Lower panel: histogram bars reporting the percentage of CSF-1R pos cells in the indicated cell lines. In the lower panel, the mean ± SE of two independent experiments is reported. ( C – D ) CSF-1R inhibition affects the clonogenicity and resistance to cisplatin of lung cancer cell lines. (C) Western blotting of CSF-1R immunoprecipitates stained with the indicated antibodies from H1299 cells treated with increasing concentrations of JNJ-40346527. (D) Upper panel. Representative micrographs of the colonies formed by H1299 cells treated with increasing doses of JNJ-40346527. Lower panel. Histograms reporting the mean ± SE of the colonies formed by 4 representative cell lines in three independent experiments. ( E ) CSF-1R inhibition affects the resistance of lung cancer cell lines to cisplatin. Graphs reporting the percentage of colonies formed by A549 and H1299 cells treated with JNJ-40346527 at the CC 25 doses determined in 2D and with the indicated doses of cisplatin. Mean ± SE of three independent experiments. ( F ) JNJ-40346527 treatment modulates the number of CSF-1R pos cells. Histograms reporting the number of CSF-1R pos cells (as assessed by FACS) in the H1299 cells treated for 96 hrs at the previously determined CC 50 dosages for both cisplatin and JNJ-40346527. Mean ± SE of three independent experiments is reported. *= p

    Article Snippet: Reagents Cisplatin (Selleckchem, Texas, USA) was dissolved according to the manufacturer's instructions.

    Techniques: Expressing, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, FACS, Inhibition, Western Blot, Staining

    The inhibition of CSF-1R affects some protumorigenic features of lung cancer cells ( A ) CSF-1R inhibition affects the propagation of Sphere Forming Cells in 3D cultures. Representative micrographs of H1299 cultures serially propagated in no serum, no adhesion conditions, in the presence of growth factors. The Sphere Forming Efficiency (SFE) was calculated as the percentage of formed spheres/seeded cells scale bar: 100 micrometers. ( B ) Graph reporting the sphere forming efficiency (SFE) of 4 representative cell lines treated with increasing doses of JNJ-40346527. Mean ± SE of two independent experiments. ( C ) Inhibition of CSF-1R affects the levels of cancer related mRNAs. Heat map. Quantitative PCR. mRNA levels of the indicated genes from H1299 and H1975 cells (expressing CSF-1R) and from H460 cells (not expressing the receptor), treated with vehicle or JNJ-40346527 for 24 hrs , alone or in combination with cisplatin(CC 50 ). Please note that at 24 hrs of treatment no significant cell death was observed at the time of harvesting (24 hrs) (data not shown). ( D – F ) Inhibition of CSF-1R affects the number of chemoresistant ALDH bright cells. (D) Representative dot plots of H1299 treated for 72 hr with cisplatin in the presence of vehicle or JNJ-40346527 (upper and lower panel, respectively). The high ALDH expressing cells were defined as the cells that displayed greater fluorescence (right panels) compared with a control staining reaction containing the ALDH inhibitor, DEAB (diethylaminobenzaldehyde) (left panels), upon addition of the synthetic ALDH substrate BAAA. (E) Histogram reporting the number of ALDH bright cells from four representative cell lines after the indicated treatments for 96 hrs. Background staining obtained with DEAB-treated cells was subtracted for each sample analyzed. The mean ± SEM of three independent experiments is reported. (F) Quantitative PCR. Levels of mRNA of the indicated ALDH isoforms in H1299 cells treated with JNJ-40346527(CC 50 ), in presence or absence of cisplatin, for 24 hrs. PP1A was used as internal control. The mean ± SE of two independent experiments is reported. Statistical differences were indicated when not significant (ns). Very similar results were obtained with H1975 cells (data not shown).

    Journal: Oncotarget

    Article Title: Inhibition of the colony-stimulating-factor-1 receptor affects the resistance of lung cancer cells to cisplatin

    doi: 10.18632/oncotarget.10895

    Figure Lengend Snippet: The inhibition of CSF-1R affects some protumorigenic features of lung cancer cells ( A ) CSF-1R inhibition affects the propagation of Sphere Forming Cells in 3D cultures. Representative micrographs of H1299 cultures serially propagated in no serum, no adhesion conditions, in the presence of growth factors. The Sphere Forming Efficiency (SFE) was calculated as the percentage of formed spheres/seeded cells scale bar: 100 micrometers. ( B ) Graph reporting the sphere forming efficiency (SFE) of 4 representative cell lines treated with increasing doses of JNJ-40346527. Mean ± SE of two independent experiments. ( C ) Inhibition of CSF-1R affects the levels of cancer related mRNAs. Heat map. Quantitative PCR. mRNA levels of the indicated genes from H1299 and H1975 cells (expressing CSF-1R) and from H460 cells (not expressing the receptor), treated with vehicle or JNJ-40346527 for 24 hrs , alone or in combination with cisplatin(CC 50 ). Please note that at 24 hrs of treatment no significant cell death was observed at the time of harvesting (24 hrs) (data not shown). ( D – F ) Inhibition of CSF-1R affects the number of chemoresistant ALDH bright cells. (D) Representative dot plots of H1299 treated for 72 hr with cisplatin in the presence of vehicle or JNJ-40346527 (upper and lower panel, respectively). The high ALDH expressing cells were defined as the cells that displayed greater fluorescence (right panels) compared with a control staining reaction containing the ALDH inhibitor, DEAB (diethylaminobenzaldehyde) (left panels), upon addition of the synthetic ALDH substrate BAAA. (E) Histogram reporting the number of ALDH bright cells from four representative cell lines after the indicated treatments for 96 hrs. Background staining obtained with DEAB-treated cells was subtracted for each sample analyzed. The mean ± SEM of three independent experiments is reported. (F) Quantitative PCR. Levels of mRNA of the indicated ALDH isoforms in H1299 cells treated with JNJ-40346527(CC 50 ), in presence or absence of cisplatin, for 24 hrs. PP1A was used as internal control. The mean ± SE of two independent experiments is reported. Statistical differences were indicated when not significant (ns). Very similar results were obtained with H1975 cells (data not shown).

    Article Snippet: Reagents Cisplatin (Selleckchem, Texas, USA) was dissolved according to the manufacturer's instructions.

    Techniques: Inhibition, Real-time Polymerase Chain Reaction, Expressing, Fluorescence, Staining

    Collagen promoted the proliferation and migration of head and neck squamous cell carcinoma (HNSCC) cells and suppressed the response to cisplatin. ( A ) Collagen significantly increased the growth of the HNSCC cell lines, SCC040, SCC154, VU040T and VU147T. ( B ) Pre-treatment of cells with collagen significantly enhanced the migration of cells through fibronectin-coated membranes. ( C ) Cells pre-treated with collagen were significantly less sensitive to cisplatin, as determined by Annexin V-FITC/Propidium Iodide staining following treatment with 50 μM cisplatin for 72 hours. Results shown are mean +/− standard deviation values of triplicates. *, **, ***, **** denote p

    Journal: Cancers

    Article Title: Collagen Induces a More Proliferative, Migratory and Chemoresistant Phenotype in Head and Neck Cancer via DDR1

    doi: 10.3390/cancers11111766

    Figure Lengend Snippet: Collagen promoted the proliferation and migration of head and neck squamous cell carcinoma (HNSCC) cells and suppressed the response to cisplatin. ( A ) Collagen significantly increased the growth of the HNSCC cell lines, SCC040, SCC154, VU040T and VU147T. ( B ) Pre-treatment of cells with collagen significantly enhanced the migration of cells through fibronectin-coated membranes. ( C ) Cells pre-treated with collagen were significantly less sensitive to cisplatin, as determined by Annexin V-FITC/Propidium Iodide staining following treatment with 50 μM cisplatin for 72 hours. Results shown are mean +/− standard deviation values of triplicates. *, **, ***, **** denote p

    Article Snippet: Annexin V-FITC/ Propidium Iodide (PI) Apoptosis Assays Cells treated with Cisplatin (Tocris Bioscience, Minneapolis, MN, USA) with or without collagen (Merck Millipore, Darmstadt, Germany) pre-treatment were collected using Accutase Cell Detachment Reagent (BD Biosciences, San Jose, CA, USA) together with the dead cells in the medium.

    Techniques: Migration, Staining, Standard Deviation

    The effects of collagen are mediated by discoidin domain receptor 1 (DDR1). ( A ) Knockdown of DDR1 following stable transduction of VU040 and VU147T cells with two independent short hairpin RNAs. RT-qPCR showed a marked reduction in DDR1 mRNA (top panel) and total DDR1 protein levels (bottom panel). The expression of DDR1 mRNA in NS (control) cells was normalized to 1. ( B ) Following DDR1 knockdown, VU040 and VU147T cells grew slower compared to NS controls in the presence of collagen. ( C ) Knockdown of DDR1 inhibited the migration of VU040T and VU147T cells pre-treated with collagen in Transwell assays. ( D ) Knockdown of DDR1 inhibited the invasion of VU040T and VU147T cells through matrigel-coated filters in Transwell assays. ( E ) The protective effects of collagen on cisplatin-induced apoptosis were reversed following DDR1 knockdown. Results shown are mean +/- standard deviation values of triplicates. *, **, *** denotes p

    Journal: Cancers

    Article Title: Collagen Induces a More Proliferative, Migratory and Chemoresistant Phenotype in Head and Neck Cancer via DDR1

    doi: 10.3390/cancers11111766

    Figure Lengend Snippet: The effects of collagen are mediated by discoidin domain receptor 1 (DDR1). ( A ) Knockdown of DDR1 following stable transduction of VU040 and VU147T cells with two independent short hairpin RNAs. RT-qPCR showed a marked reduction in DDR1 mRNA (top panel) and total DDR1 protein levels (bottom panel). The expression of DDR1 mRNA in NS (control) cells was normalized to 1. ( B ) Following DDR1 knockdown, VU040 and VU147T cells grew slower compared to NS controls in the presence of collagen. ( C ) Knockdown of DDR1 inhibited the migration of VU040T and VU147T cells pre-treated with collagen in Transwell assays. ( D ) Knockdown of DDR1 inhibited the invasion of VU040T and VU147T cells through matrigel-coated filters in Transwell assays. ( E ) The protective effects of collagen on cisplatin-induced apoptosis were reversed following DDR1 knockdown. Results shown are mean +/- standard deviation values of triplicates. *, **, *** denotes p

    Article Snippet: Annexin V-FITC/ Propidium Iodide (PI) Apoptosis Assays Cells treated with Cisplatin (Tocris Bioscience, Minneapolis, MN, USA) with or without collagen (Merck Millipore, Darmstadt, Germany) pre-treatment were collected using Accutase Cell Detachment Reagent (BD Biosciences, San Jose, CA, USA) together with the dead cells in the medium.

    Techniques: Transduction, Quantitative RT-PCR, Expressing, Migration, Standard Deviation

    PDC/PDX-guided treatment of patients under two independent n = 1 co-clinical trials. a Timeline for patient HN137 from surgery (December), adjuvant chemo-radiation therapy, until tumour recurrence in June. b Graph denoting Log 2 (IC 50 ) values of HN137-Pri, HN137-Pri cisplatin resistant (CR), HN137-Met, HN137-Met cisplatin resistant (CR) cell lines in the presence of gefitinib. c Computed tomography scan (CT-scan) of recurrent responsive metastatic sites (dermal metastasis ( top panel ) and lung metastasis ( bottom panel )) in HN137 patient, before and after treatment with 250 mg per day of gefitinib. Arrows denote sites of tumours before and after treatment. d Dose response of HN177-PDC to erlotinib and olaparib. Cell viability was determined using CellTiter-Glo reagent. Triplicate data, error bars represent mean ± s.d. e Three independent cohorts of mice ( n = 2 for control and n = 3 for treated) bearing HN177-PDX in one flank were treated with vehicle control (Ctrl), 50 mg kg −1 olaparib and 150 mg kg −1 erlotinib. Error bars represent mean ± s.e.m. Two-tail Student’s t test was carried out between olaparib and control group (N.S.: not significant) and between erlotinib and control group ** P value

    Journal: Nature Communications

    Article Title: Phenotype-driven precision oncology as a guide for clinical decisions one patient at a time

    doi: 10.1038/s41467-017-00451-5

    Figure Lengend Snippet: PDC/PDX-guided treatment of patients under two independent n = 1 co-clinical trials. a Timeline for patient HN137 from surgery (December), adjuvant chemo-radiation therapy, until tumour recurrence in June. b Graph denoting Log 2 (IC 50 ) values of HN137-Pri, HN137-Pri cisplatin resistant (CR), HN137-Met, HN137-Met cisplatin resistant (CR) cell lines in the presence of gefitinib. c Computed tomography scan (CT-scan) of recurrent responsive metastatic sites (dermal metastasis ( top panel ) and lung metastasis ( bottom panel )) in HN137 patient, before and after treatment with 250 mg per day of gefitinib. Arrows denote sites of tumours before and after treatment. d Dose response of HN177-PDC to erlotinib and olaparib. Cell viability was determined using CellTiter-Glo reagent. Triplicate data, error bars represent mean ± s.d. e Three independent cohorts of mice ( n = 2 for control and n = 3 for treated) bearing HN177-PDX in one flank were treated with vehicle control (Ctrl), 50 mg kg −1 olaparib and 150 mg kg −1 erlotinib. Error bars represent mean ± s.e.m. Two-tail Student’s t test was carried out between olaparib and control group (N.S.: not significant) and between erlotinib and control group ** P value

    Article Snippet: Compounds Cisplatin was purchased from Tocris (cat. no. 2251), YM155 was purchased from Selleckchem (cat. no. S1130) and gefitinib was purchased from Cayman Chemical (cat. no. 13166).

    Techniques: Computed Tomography, Mouse Assay

    Extracellular vesicles (EV) secreted by hypoxia pre-challenged mesenchymal stem cells (MSCs) promote A549 and H23 cell proliferation, survival, mobility and EMT in vitro. Cells were pre-treated with EVs secreted by naïve (N-EV) or hypoxia pre-challenged (H-EV) MSCs. a and b , CCK-8 cell proliferations assay evaluating viable cell count at different timepoints. c – g , A549 and H23 cell apoptosis under normal, hypoxia challenge (1% O 2 ) or cisplatin challenge (5 μM) after N-EV or H-EV treatment. Hypoxia or cisplatin challenge was performed for 24 h before analysis. h – i , trans-well assay evaluating A549 and H23 cells invasion after N-EV or H-EV treatment. A549 or H23 cells treated with PBS vehicle were used as negative control (NC). Bar in I indicates 20 μm. j – l , western blot detecting epithelial and mesenchymal marker in A549 and H23 cells after treatment with N-EV or H-EV. Statistical analysis results by Student’s t test was marked by # and that by Dunnett’s test were marked by *. * or #, p

    Journal: Journal of Experimental & Clinical Cancer Research : CR

    Article Title: Extracellular vesicles secreted by hypoxia pre-challenged mesenchymal stem cells promote non-small cell lung cancer cell growth and mobility as well as macrophage M2 polarization via miR-21-5p delivery

    doi: 10.1186/s13046-019-1027-0

    Figure Lengend Snippet: Extracellular vesicles (EV) secreted by hypoxia pre-challenged mesenchymal stem cells (MSCs) promote A549 and H23 cell proliferation, survival, mobility and EMT in vitro. Cells were pre-treated with EVs secreted by naïve (N-EV) or hypoxia pre-challenged (H-EV) MSCs. a and b , CCK-8 cell proliferations assay evaluating viable cell count at different timepoints. c – g , A549 and H23 cell apoptosis under normal, hypoxia challenge (1% O 2 ) or cisplatin challenge (5 μM) after N-EV or H-EV treatment. Hypoxia or cisplatin challenge was performed for 24 h before analysis. h – i , trans-well assay evaluating A549 and H23 cells invasion after N-EV or H-EV treatment. A549 or H23 cells treated with PBS vehicle were used as negative control (NC). Bar in I indicates 20 μm. j – l , western blot detecting epithelial and mesenchymal marker in A549 and H23 cells after treatment with N-EV or H-EV. Statistical analysis results by Student’s t test was marked by # and that by Dunnett’s test were marked by *. * or #, p

    Article Snippet: For hypoxia or cisplatin challenge, A549 or H23 cells were pre-treated with different MSC-EV for 24 h, followed by incubation under 1% O2 (hypoxic) conditions or with 5 μM of cisplatin (Tocris Bioscience, Minneapolis, USA) for 24 h, respectively.

    Techniques: In Vitro, CCK-8 Assay, Cell Counting, Negative Control, Western Blot, Marker

    Pharmacological blockade of CB 2 or CB 1 receptors does not alter cisplatin- or paclitaxel-induced neuropathic nociception. Neither the CB 2 antagonist AM630 (3 mg/kg i.p.) nor the CB 1 antagonist AM251 (3 mg/kg i.p.) altered cisplatin- ( A , C ) or paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia. ### P

    Journal: Molecular Pain

    Article Title: The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    doi: 10.1186/1744-8069-8-71

    Figure Lengend Snippet: Pharmacological blockade of CB 2 or CB 1 receptors does not alter cisplatin- or paclitaxel-induced neuropathic nociception. Neither the CB 2 antagonist AM630 (3 mg/kg i.p.) nor the CB 1 antagonist AM251 (3 mg/kg i.p.) altered cisplatin- ( A , C ) or paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia. ### P

    Article Snippet: Drugs and chemicals Cisplatin was purchased from Tocris Bioscience (Ellisville, MO, USA) and was dissolved in saline (0.9% sodium chloride).

    Techniques:

    Time course of development of chemotherapy-induced peripheral neuropathy evoked by cisplatin and paclitaxel treatment. Mechanical ( A , B ) and cold ( C , D ) allodynia developed following cisplatin ( A , C ) or paclitaxel ( B , D ) treatment. Arrows show timing of injections of chemotherapeutic agents. Data are expressed as mean ± SEM (paclitaxel, n = 76; cremophor, n = 5; cisplatin, n = 36; saline, n = 6). *** P

    Journal: Molecular Pain

    Article Title: The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    doi: 10.1186/1744-8069-8-71

    Figure Lengend Snippet: Time course of development of chemotherapy-induced peripheral neuropathy evoked by cisplatin and paclitaxel treatment. Mechanical ( A , B ) and cold ( C , D ) allodynia developed following cisplatin ( A , C ) or paclitaxel ( B , D ) treatment. Arrows show timing of injections of chemotherapeutic agents. Data are expressed as mean ± SEM (paclitaxel, n = 76; cremophor, n = 5; cisplatin, n = 36; saline, n = 6). *** P

    Article Snippet: Drugs and chemicals Cisplatin was purchased from Tocris Bioscience (Ellisville, MO, USA) and was dissolved in saline (0.9% sodium chloride).

    Techniques:

    Effect of AM1710 on chemotherapy-induced mechanical and cold allodynia. AM1710 suppressed both mechanical ( A , B ) and cold ( C , D ) allodynia evoked by cisplatin ( A , C ) or paclitaxel ( B , D ) treatment. Data are expressed as mean ± SEM (n = 5-13 per group). *** P

    Journal: Molecular Pain

    Article Title: The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    doi: 10.1186/1744-8069-8-71

    Figure Lengend Snippet: Effect of AM1710 on chemotherapy-induced mechanical and cold allodynia. AM1710 suppressed both mechanical ( A , B ) and cold ( C , D ) allodynia evoked by cisplatin ( A , C ) or paclitaxel ( B , D ) treatment. Data are expressed as mean ± SEM (n = 5-13 per group). *** P

    Article Snippet: Drugs and chemicals Cisplatin was purchased from Tocris Bioscience (Ellisville, MO, USA) and was dissolved in saline (0.9% sodium chloride).

    Techniques:

    The cannabilactone AM1710 suppresses chemotherapy-induced mechanical and cold allodynia through a CB 2 -specific mechanism. AM1710-induced suppressions of cisplatin- ( A , C ) and paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia were blocked by the CB 2 antagonist AM630 (3 mg/kg i.p.) but not the CB 1 antagonist AM251 (3 mg/kg i.p.). Data are expressed as mean ± SEM (n = 5-6 per group). *** P

    Journal: Molecular Pain

    Article Title: The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    doi: 10.1186/1744-8069-8-71

    Figure Lengend Snippet: The cannabilactone AM1710 suppresses chemotherapy-induced mechanical and cold allodynia through a CB 2 -specific mechanism. AM1710-induced suppressions of cisplatin- ( A , C ) and paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia were blocked by the CB 2 antagonist AM630 (3 mg/kg i.p.) but not the CB 1 antagonist AM251 (3 mg/kg i.p.). Data are expressed as mean ± SEM (n = 5-6 per group). *** P

    Article Snippet: Drugs and chemicals Cisplatin was purchased from Tocris Bioscience (Ellisville, MO, USA) and was dissolved in saline (0.9% sodium chloride).

    Techniques:

    CXCR4 signaling did not contribute to either chemotherapy-induced neuropathy or CB 2 agonist efficacy. The CXCR4 antagonist AMD3100 (10 mg/kg i.p.) failed to suppress the maintenance of cisplatin- ( A , C ) or paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia. The CXCR4 antagonist AMD3100 (10 mg/kg i.p.) did not alter antinociceptive efficacy of the CB 2 agonist AM1710 (5 mg/kg i.p.) in suppressing paclitaxel-induced mechanical ( B ) and cold ( D ) allodynia. *** P

    Journal: Molecular Pain

    Article Title: The maintenance of cisplatin- and paclitaxel-induced mechanical and cold allodynia is suppressed by cannabinoid CB2 receptor activation and independent of CXCR4 signaling in models of chemotherapy-induced peripheral neuropathy

    doi: 10.1186/1744-8069-8-71

    Figure Lengend Snippet: CXCR4 signaling did not contribute to either chemotherapy-induced neuropathy or CB 2 agonist efficacy. The CXCR4 antagonist AMD3100 (10 mg/kg i.p.) failed to suppress the maintenance of cisplatin- ( A , C ) or paclitaxel- ( B , D ) evoked mechanical ( A , B ) and cold ( C , D ) allodynia. The CXCR4 antagonist AMD3100 (10 mg/kg i.p.) did not alter antinociceptive efficacy of the CB 2 agonist AM1710 (5 mg/kg i.p.) in suppressing paclitaxel-induced mechanical ( B ) and cold ( D ) allodynia. *** P

    Article Snippet: Drugs and chemicals Cisplatin was purchased from Tocris Bioscience (Ellisville, MO, USA) and was dissolved in saline (0.9% sodium chloride).

    Techniques:

    SETD8 inhibition preferentially sensitises LUSC cell lines to chemotherapy. a Dose–response curves were derived by treating NSCLC cell lines with SETD8 inhibitor NSC663284. 1000 cells were seeded and allowed to recover for 24 h. The inhibitor was then added at increasing concentrations to LUSC (red) and LUAD (blue) cells and Cell Titre (see Methods) assay was performed after 72 h. b IC 50 values were derived from the dose–response assay indicating LUSC cells are significantly more responsive to SETD8 inhibition than LUAD cells. c Dose–response curves were derived by treating NSCLC cell lines with cisplatin as above. d IC 50 values were derived from the dose–response assay indicating cisplatin effects LUSC and LUAD cells equally. e Dose–response curves derived from treating NSCLC cell lines with cisplatin and NSC663284 IC 50 concentration for each cell line as above. f IC 50 values were derived from the dose–response assay indicating SETD8 inhibition preferentially enhances cisplatin efficacy in LUSC cells. The whiskers indicate the range of the data and the line represents the median. Data presented as mean ± s.d. (LUSC n = 5 and LUAD n = 6). Student’s t -test performed, * p

    Journal: Nature Communications

    Article Title: BCL11A interacts with SOX2 to control the expression of epigenetic regulators in lung squamous carcinoma

    doi: 10.1038/s41467-018-05790-5

    Figure Lengend Snippet: SETD8 inhibition preferentially sensitises LUSC cell lines to chemotherapy. a Dose–response curves were derived by treating NSCLC cell lines with SETD8 inhibitor NSC663284. 1000 cells were seeded and allowed to recover for 24 h. The inhibitor was then added at increasing concentrations to LUSC (red) and LUAD (blue) cells and Cell Titre (see Methods) assay was performed after 72 h. b IC 50 values were derived from the dose–response assay indicating LUSC cells are significantly more responsive to SETD8 inhibition than LUAD cells. c Dose–response curves were derived by treating NSCLC cell lines with cisplatin as above. d IC 50 values were derived from the dose–response assay indicating cisplatin effects LUSC and LUAD cells equally. e Dose–response curves derived from treating NSCLC cell lines with cisplatin and NSC663284 IC 50 concentration for each cell line as above. f IC 50 values were derived from the dose–response assay indicating SETD8 inhibition preferentially enhances cisplatin efficacy in LUSC cells. The whiskers indicate the range of the data and the line represents the median. Data presented as mean ± s.d. (LUSC n = 5 and LUAD n = 6). Student’s t -test performed, * p

    Article Snippet: Cisplatin (LKT Laboratories, C3374) was suspended in 154 mM NaCl at a 3 mM stock concentration.

    Techniques: Inhibition, Derivative Assay, Concentration Assay

    Effects of paclitaxel, cisplatin and oxaliplatin on primary cultured DRG neurons or on co-cultures of Schwann cells and DRG neurons. ( a ) Immunofluorescence staining of primary cultured DRG neurons for MAP2. Primary cultured DRG neurons were treated with vehicle (0.1% DMSO), paclitaxel (0.01 and 0.1 μM), cisplatin (1 and 3 μM) or oxaliplatin (3 and 10 μM) for 48 h. Only higher doses of paclitaxel (0.1 μM), cisplatin (3 μM) and oxaliplatin (10 μM) reduced the number of DRG neurons and associated dendritic trees, and shortened the neuritic processes. Scale bar: 100 μm. ( b ) Co-cultures were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM), or oxaliplatin (3 μM) for 48 h. Scale bar: 200 μm. Each treatment reduced formation of MBP-positive myelin; however, no morphological changes were observed in MAP2-positive DRG neurons. ( c ) MBP- or MAP2-IR in co-cultures. Each column represents the mean ± S.E.M. n = 3. *** p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Effects of paclitaxel, cisplatin and oxaliplatin on primary cultured DRG neurons or on co-cultures of Schwann cells and DRG neurons. ( a ) Immunofluorescence staining of primary cultured DRG neurons for MAP2. Primary cultured DRG neurons were treated with vehicle (0.1% DMSO), paclitaxel (0.01 and 0.1 μM), cisplatin (1 and 3 μM) or oxaliplatin (3 and 10 μM) for 48 h. Only higher doses of paclitaxel (0.1 μM), cisplatin (3 μM) and oxaliplatin (10 μM) reduced the number of DRG neurons and associated dendritic trees, and shortened the neuritic processes. Scale bar: 100 μm. ( b ) Co-cultures were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM), or oxaliplatin (3 μM) for 48 h. Scale bar: 200 μm. Each treatment reduced formation of MBP-positive myelin; however, no morphological changes were observed in MAP2-positive DRG neurons. ( c ) MBP- or MAP2-IR in co-cultures. Each column represents the mean ± S.E.M. n = 3. *** p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: Cell Culture, Immunofluorescence, Staining

    Viability of Schwann cells after exposure to paclitaxel, cisplatin or oxaliplatin. At 2 days after culture in differentiation medium, Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel ( a , d ; 0.001–0.1 μM), cisplatin ( b , e ; 0.1–10 μM) or oxaliplatin ( c , f ; 0.1–10 μM) for 24 h ( a – c ) or 48 h ( d – f ). Cell viability was measured in an MTT assay, and the results were expressed as a percentage relative to vehicle-treated cells. Each column represents the mean ± S.E.M. n = 3–6. * p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Viability of Schwann cells after exposure to paclitaxel, cisplatin or oxaliplatin. At 2 days after culture in differentiation medium, Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel ( a , d ; 0.001–0.1 μM), cisplatin ( b , e ; 0.1–10 μM) or oxaliplatin ( c , f ; 0.1–10 μM) for 24 h ( a – c ) or 48 h ( d – f ). Cell viability was measured in an MTT assay, and the results were expressed as a percentage relative to vehicle-treated cells. Each column represents the mean ± S.E.M. n = 3–6. * p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: MTT Assay

    Effect of paclitaxel, cisplatin and oxaliplatin on expression of galectin-3 in primary cultured Schwann cells. Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM) or cisplatin (1 μM) for 48 h. ( a ) Immunofluorescence staining of primary cultured Schwann cells with DAPI (blue) and an anti-galectin-3 antibody (green). Scale bar: 100 μm. ( b ) Galectin-3-IR in Schwann cells. Each column represents the mean ± S.E.M. n = 3. *** p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Effect of paclitaxel, cisplatin and oxaliplatin on expression of galectin-3 in primary cultured Schwann cells. Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM) or cisplatin (1 μM) for 48 h. ( a ) Immunofluorescence staining of primary cultured Schwann cells with DAPI (blue) and an anti-galectin-3 antibody (green). Scale bar: 100 μm. ( b ) Galectin-3-IR in Schwann cells. Each column represents the mean ± S.E.M. n = 3. *** p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: Expressing, Cell Culture, Immunofluorescence, Staining

    Effects of paclitaxel, cisplatin and oxaliplatin on Schwann cell morphology and MBP expression. ( a ) Phase ( upper ) and fluorescence ( lower ) micrographs of differentiated Schwann cells labeled with an anti-MBP antibody. Differentiated Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. Cisplatin and oxaliplatin reduced the number of MBP-positive Schwann cells, but did not affect cell morphology. Unlike cisplatin and oxaliplatin, paclitaxel induced morphological changes, characterized by retraction of bipolar processes and a rounded shape. Scale bar: 100 μm (25 μm in enlarged images). ( b ) Expression of MBP in Schwann cells 48 h after treatment with vehicle (0.1% DMSO), paclitaxel (0.001 and 0.01 μM) or cisplatin (1 and 3 μM) was analyzed by Western blotting. ( Upper panels ) Representative Western blot showing expression of MBP and actin. ( Lower panels ) Quantification of band intensity. The intensity of each band was normalized to that of actin (loading control). Each column represents the mean ± S.E.M. n = 5–8. * p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Effects of paclitaxel, cisplatin and oxaliplatin on Schwann cell morphology and MBP expression. ( a ) Phase ( upper ) and fluorescence ( lower ) micrographs of differentiated Schwann cells labeled with an anti-MBP antibody. Differentiated Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. Cisplatin and oxaliplatin reduced the number of MBP-positive Schwann cells, but did not affect cell morphology. Unlike cisplatin and oxaliplatin, paclitaxel induced morphological changes, characterized by retraction of bipolar processes and a rounded shape. Scale bar: 100 μm (25 μm in enlarged images). ( b ) Expression of MBP in Schwann cells 48 h after treatment with vehicle (0.1% DMSO), paclitaxel (0.001 and 0.01 μM) or cisplatin (1 and 3 μM) was analyzed by Western blotting. ( Upper panels ) Representative Western blot showing expression of MBP and actin. ( Lower panels ) Quantification of band intensity. The intensity of each band was normalized to that of actin (loading control). Each column represents the mean ± S.E.M. n = 5–8. * p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: Expressing, Fluorescence, Labeling, Western Blot

    Changes in mitochondrial activity in Schwann cells after treatment with paclitaxel, cisplatin or oxaliplatin. ( a ) Fluorescence micrographs of Schwann cells labeled with the MitoTracker probe (red) and DAPI (blue). Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. Scale bar: 50 μm (10 μm in enlarged images) in the vehicle-treated group. ( b ) Number of mitochondria-labeled cells per 100 DAPI-positive cells. The intensity and number of fluorescently labeled mitochondria were reduced after treatment with either cisplatin or oxaliplatin, but not after treatment with paclitaxel. Each column represents the mean ± S.E.M. n = 12. * p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Changes in mitochondrial activity in Schwann cells after treatment with paclitaxel, cisplatin or oxaliplatin. ( a ) Fluorescence micrographs of Schwann cells labeled with the MitoTracker probe (red) and DAPI (blue). Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. Scale bar: 50 μm (10 μm in enlarged images) in the vehicle-treated group. ( b ) Number of mitochondria-labeled cells per 100 DAPI-positive cells. The intensity and number of fluorescently labeled mitochondria were reduced after treatment with either cisplatin or oxaliplatin, but not after treatment with paclitaxel. Each column represents the mean ± S.E.M. n = 12. * p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: Activity Assay, Fluorescence, Labeling

    Effect of paclitaxel, cisplatin and oxaliplatin on expression of p75 in primary cultured Schwann cells. Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. ( a ) Immunofluorescent staining of primary cultured Schwann cells for p75 (red) and GFAP (green). Scale bar: 100 μm. ( b ) p75-IR in Schwann cells. Each column represents the mean ± S.E.M. n = 3. *** p

    Journal: Scientific Reports

    Article Title: Taxanes and platinum derivatives impair Schwann cells via distinct mechanisms

    doi: 10.1038/s41598-017-05784-1

    Figure Lengend Snippet: Effect of paclitaxel, cisplatin and oxaliplatin on expression of p75 in primary cultured Schwann cells. Schwann cells were treated with vehicle (0.1% DMSO), paclitaxel (0.01 μM), cisplatin (1 μM) or oxaliplatin (3 μM) for 48 h. ( a ) Immunofluorescent staining of primary cultured Schwann cells for p75 (red) and GFAP (green). Scale bar: 100 μm. ( b ) p75-IR in Schwann cells. Each column represents the mean ± S.E.M. n = 3. *** p

    Article Snippet: Drugs and chemicals Cisplatin (LKT Laboratories, Inc., St Paul, MN, USA) and oxaliplatin (Wako Pure Chemical Industries, Osaka, Japan) were dissolved in phosphate-buffered saline (PBS, pH 7.4) at a concentration of 1 mM and distilled water at a concentration of 10 mM, respectively, prior to use.

    Techniques: Expressing, Cell Culture, Staining

    Detection of glutathionylated p53 protein in human tumor cells using anti p53-glut antibodies Induction of glut-p53 protein was determined in HCT116 cells after 0.4 mM H 2 O 2 treatment for different times (A), after 20 min incubation with H 2 O 2 (0.4 mM), TBH (0.6 mM) and diamide (0.6 mM) (B), with camptothecin (2 μM, 5h) (C) , and with camptothecin (2 μM), cisplatin (10 μM), and doxorubicin (5 μM) for 6 and 12 h (C). In the last four lanes of (A) and (B) and last lane of (C) the protein samples were treated with DTT before SDS-PAGE to reverse p53 glutathionylation. C, untreated control; CPT, camptothecin.

    Journal: Free radical biology & medicine

    Article Title: Cys-141 glutathionylation of human p53: Studies using specific polyclonal antibodies in cancer samples and cell lines

    doi: 10.1016/j.freeradbiomed.2010.06.020

    Figure Lengend Snippet: Detection of glutathionylated p53 protein in human tumor cells using anti p53-glut antibodies Induction of glut-p53 protein was determined in HCT116 cells after 0.4 mM H 2 O 2 treatment for different times (A), after 20 min incubation with H 2 O 2 (0.4 mM), TBH (0.6 mM) and diamide (0.6 mM) (B), with camptothecin (2 μM, 5h) (C) , and with camptothecin (2 μM), cisplatin (10 μM), and doxorubicin (5 μM) for 6 and 12 h (C). In the last four lanes of (A) and (B) and last lane of (C) the protein samples were treated with DTT before SDS-PAGE to reverse p53 glutathionylation. C, untreated control; CPT, camptothecin.

    Article Snippet: The anticancer drugs cisplatin, doxorubicin and camptothecin were obtained from LKT laboratories (St. Paul, MN).

    Techniques: Incubation, SDS Page, Cycling Probe Technology

    Immunofluorescence staining of glutathionylated p53 in HCT116, HT29, and T47D cells upon treatment with oxidizing and DNA damaging agents Immunofluorescence showing the presence of p53-glut in untreated HCT116 cells (A) ; HCT116 cells treated with 0.6 mM diamide for 30 min (B) ; HCT116 cells treated as in (B) followed by 10 mM DTT treatment for 30 min (C) ; HCT116 cells treated with 0.4 mM H 2 O 2 for 30 min (D); HCT116 cells treated as in (D) and postincubated in H 2 O 2 free medium for 2h (E) ; HCT116 cells treated with 5 μM doxorubicin for 5h (F), and HCT116 cells treated with 10 μM cisplatin for 5h (G) . Immunofluorescence showing the presence of p53-glut in untreated HT29 cells (H); HT29 cells treated with 0.6 mM diamide for 30 min (I); untreated T47D cells (J), and T47D cells treated with 0.6 mM diamide for 30 min (K) . After the respective treatments, cells were fixed using 4% paraformaldehyde and incubated with p53-glut antibody (1:500 dilution) followed by goat anti rabbit Alexa Flour 594 secondary antibody. The cells were photographed using an Olympus IX 81 microscope. Similar patterns of staining were observed in three independent experiments.

    Journal: Free radical biology & medicine

    Article Title: Cys-141 glutathionylation of human p53: Studies using specific polyclonal antibodies in cancer samples and cell lines

    doi: 10.1016/j.freeradbiomed.2010.06.020

    Figure Lengend Snippet: Immunofluorescence staining of glutathionylated p53 in HCT116, HT29, and T47D cells upon treatment with oxidizing and DNA damaging agents Immunofluorescence showing the presence of p53-glut in untreated HCT116 cells (A) ; HCT116 cells treated with 0.6 mM diamide for 30 min (B) ; HCT116 cells treated as in (B) followed by 10 mM DTT treatment for 30 min (C) ; HCT116 cells treated with 0.4 mM H 2 O 2 for 30 min (D); HCT116 cells treated as in (D) and postincubated in H 2 O 2 free medium for 2h (E) ; HCT116 cells treated with 5 μM doxorubicin for 5h (F), and HCT116 cells treated with 10 μM cisplatin for 5h (G) . Immunofluorescence showing the presence of p53-glut in untreated HT29 cells (H); HT29 cells treated with 0.6 mM diamide for 30 min (I); untreated T47D cells (J), and T47D cells treated with 0.6 mM diamide for 30 min (K) . After the respective treatments, cells were fixed using 4% paraformaldehyde and incubated with p53-glut antibody (1:500 dilution) followed by goat anti rabbit Alexa Flour 594 secondary antibody. The cells were photographed using an Olympus IX 81 microscope. Similar patterns of staining were observed in three independent experiments.

    Article Snippet: The anticancer drugs cisplatin, doxorubicin and camptothecin were obtained from LKT laboratories (St. Paul, MN).

    Techniques: Immunofluorescence, Staining, Incubation, Microscopy

    mtFoxO3A is involved in cancer cell response to metabolic stress. a HCT116-FoxO3A +/+ and HCT116-FoxO3A −/− cells were subjected to different treatments: glucose restriction (LG, 0.75 mM glucose, 24 h), metformin (MET, 10 μM, 72 h), cisplatin (CDDP, 30 μM, 48 h), irinotecan (CPT-11, 30 μM, 24 h), 5-fluorouracil (5-FU, 2 μM, 24 h) and etoposide (VP-13, 40 μM, 24 h). Relative cell viability and relative cell death were calculated. b Correlation between LG-resistance (days) and mitochondrial FoxO3A (mtFoxO3A) protein levels in different human cell lines (HCT116 and HT29 colorectal cancer cells, HEK293 embryonic kidney cell, DU145 prostate cancer cells, A549 lung cancer cells, MDA-MB-468 breast cancer cells and OVCAR3 ovarian cancer cells). a.u. arbitrary units. c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids (48 h) and subjected to LG (24 h). Upper panel: relative cell viability and relative cell death. Lower panel: immunoblot analysis of total proteins. β-actin: loading control. d Transcription analysis of selected mitochondrial ( ND6 and COX1 ) and nuclear ( BIM ) genes by RT-PCR in HCT116-FoxO3A −/− cells transfected with the indicated plasmids (48 h) and subjected to LG (24 h). e HCT116-FoxO3A −/− cells, transfected with the indicated plasmids (48 h), were subjected to metabolic stress with 2-DG (1 mM, 6 h). The graph reflects the quantification of tetramethylrhodamine ethyl ester (TMRE) fluorescence of active mitochondria in transfected cells. f Clonogenic assay on HCT116-FoxO3A +/+ cells cultured in LG (24 h) and treated with increasing concentrations of trametinib and/or compound C, as indicated, for 24 h. Cell growth percent inhibition at each drug concentration is presented. g Left panel: immunoblot analysis of total and mitochondrial proteins isolated from tumors ( n ≥ 7 for each group) derived from HCT116-xenografted nude mice subjected to 2-DG treatment (100 mg/kg, 6 days). β-actin and HSP60 were used as total and mitochondrial loading control, respectively. Right panel: densitometric analysis of full-length and cleaved FoxO3A normalized against the mitochondrial loading control and the results of the densitometric analysis of the phosphorylated-AMPK and ERK normalized against total AMPK and ERK, respectively, and the loading control. Data are presented as mean ± SEM and significance was calculated with Student’s t test; * p

    Journal: Cell Death & Disease

    Article Title: Uncoupling FoxO3A mitochondrial and nuclear functions in cancer cells undergoing metabolic stress and chemotherapy

    doi: 10.1038/s41419-018-0336-0

    Figure Lengend Snippet: mtFoxO3A is involved in cancer cell response to metabolic stress. a HCT116-FoxO3A +/+ and HCT116-FoxO3A −/− cells were subjected to different treatments: glucose restriction (LG, 0.75 mM glucose, 24 h), metformin (MET, 10 μM, 72 h), cisplatin (CDDP, 30 μM, 48 h), irinotecan (CPT-11, 30 μM, 24 h), 5-fluorouracil (5-FU, 2 μM, 24 h) and etoposide (VP-13, 40 μM, 24 h). Relative cell viability and relative cell death were calculated. b Correlation between LG-resistance (days) and mitochondrial FoxO3A (mtFoxO3A) protein levels in different human cell lines (HCT116 and HT29 colorectal cancer cells, HEK293 embryonic kidney cell, DU145 prostate cancer cells, A549 lung cancer cells, MDA-MB-468 breast cancer cells and OVCAR3 ovarian cancer cells). a.u. arbitrary units. c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids (48 h) and subjected to LG (24 h). Upper panel: relative cell viability and relative cell death. Lower panel: immunoblot analysis of total proteins. β-actin: loading control. d Transcription analysis of selected mitochondrial ( ND6 and COX1 ) and nuclear ( BIM ) genes by RT-PCR in HCT116-FoxO3A −/− cells transfected with the indicated plasmids (48 h) and subjected to LG (24 h). e HCT116-FoxO3A −/− cells, transfected with the indicated plasmids (48 h), were subjected to metabolic stress with 2-DG (1 mM, 6 h). The graph reflects the quantification of tetramethylrhodamine ethyl ester (TMRE) fluorescence of active mitochondria in transfected cells. f Clonogenic assay on HCT116-FoxO3A +/+ cells cultured in LG (24 h) and treated with increasing concentrations of trametinib and/or compound C, as indicated, for 24 h. Cell growth percent inhibition at each drug concentration is presented. g Left panel: immunoblot analysis of total and mitochondrial proteins isolated from tumors ( n ≥ 7 for each group) derived from HCT116-xenografted nude mice subjected to 2-DG treatment (100 mg/kg, 6 days). β-actin and HSP60 were used as total and mitochondrial loading control, respectively. Right panel: densitometric analysis of full-length and cleaved FoxO3A normalized against the mitochondrial loading control and the results of the densitometric analysis of the phosphorylated-AMPK and ERK normalized against total AMPK and ERK, respectively, and the loading control. Data are presented as mean ± SEM and significance was calculated with Student’s t test; * p

    Article Snippet: Compound C (5 μM, #171260), AICAR (5 mM, #A9978), PD98059 (0,02 mM, #P215), 2-deoxy-glucose (1 mM, #D8375), iodoacetic acid (0,5 μM, #I4386), metformin (10 μM, #D150959), 5-fluorouracil (2 μM, #F6627), cisplatin (30 μM, #P4394), etoposide (40 μM, #E1383) and irinotecan (30 μM, #347609) were all from Sigma; trametinib (#S2673) was from Selleckchem (Munich, German).

    Techniques: Cycling Probe Technology, Multiple Displacement Amplification, Transfection, Reverse Transcription Polymerase Chain Reaction, Fluorescence, Clonogenic Assay, Cell Culture, Inhibition, Concentration Assay, Isolation, Derivative Assay, Mouse Assay

    mtFoxO3A is involved in cancer cell response to chemotherapeutic agents. a – c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with irinotecan (CPT-11, 30 μM, 24 h). a Upper panel: relative cell viability and relative cell death. Lower panel: immunoblot analysis of total proteins. β-actin: loading control. b Transcription analysis of selected mitochondrial ( ND6 and COX1 ) and nuclear ( BIM ) genes by RT-PCR. c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with irinotecan (CPT-11, 30 μM, 24 h). Relative cell viability and relative cell death were calculated. d Left panel: immunoblot analysis of total and mitochondrial proteins isolated from tumors ( n ≥ 7 for each group) derived from HCT116-xenografted nude mice subjected to cisplatin treatment (CDDP, 2 mg/kg, 6 days). β-actin and HSP60 were used as total lysate and mitochondrial fraction controls, respectively. Right panel: densitometric analysis of full-length and cleaved FoxO3A normalized against the mitochondrial fractionation loading control and the results of the densitometric analysis of the phosphorylated forms of AMPK and ERK normalized against total AMPK and ERK, respectively, and the loading control. e Clonogenic assay on HCT116-FoxO3A +/+ cells treated with increasing concentrations of trametinib (24 h) and/or irinotecan (24 h), as indicated. f Immunoblot analysis of total proteins isolated from HCT116-FoxO3A +/+ cells upon metformin treatment (MET, 10 μM, 72 h). β-actin: loading control. g HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with metformin (MET, 10 μM, 72 h). Relative cell viability and relative cell death were calculated. h Clonogenic assay on HCT116-FoxO3A +/+ cells treated with increasing concentrations of metformin (24 h) and/or irinotecan (24 h), as indicated. e , h Cell growth percent inhibition at each drug concentration is presented. The data presented are the mean of at least three independent experiments. Where applicable, data are presented as mean ± SEM and significance was calculated with Student’s t test; * p

    Journal: Cell Death & Disease

    Article Title: Uncoupling FoxO3A mitochondrial and nuclear functions in cancer cells undergoing metabolic stress and chemotherapy

    doi: 10.1038/s41419-018-0336-0

    Figure Lengend Snippet: mtFoxO3A is involved in cancer cell response to chemotherapeutic agents. a – c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with irinotecan (CPT-11, 30 μM, 24 h). a Upper panel: relative cell viability and relative cell death. Lower panel: immunoblot analysis of total proteins. β-actin: loading control. b Transcription analysis of selected mitochondrial ( ND6 and COX1 ) and nuclear ( BIM ) genes by RT-PCR. c HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with irinotecan (CPT-11, 30 μM, 24 h). Relative cell viability and relative cell death were calculated. d Left panel: immunoblot analysis of total and mitochondrial proteins isolated from tumors ( n ≥ 7 for each group) derived from HCT116-xenografted nude mice subjected to cisplatin treatment (CDDP, 2 mg/kg, 6 days). β-actin and HSP60 were used as total lysate and mitochondrial fraction controls, respectively. Right panel: densitometric analysis of full-length and cleaved FoxO3A normalized against the mitochondrial fractionation loading control and the results of the densitometric analysis of the phosphorylated forms of AMPK and ERK normalized against total AMPK and ERK, respectively, and the loading control. e Clonogenic assay on HCT116-FoxO3A +/+ cells treated with increasing concentrations of trametinib (24 h) and/or irinotecan (24 h), as indicated. f Immunoblot analysis of total proteins isolated from HCT116-FoxO3A +/+ cells upon metformin treatment (MET, 10 μM, 72 h). β-actin: loading control. g HCT116-FoxO3A −/− cells were transfected with the indicated plasmids for 48 h and then treated with metformin (MET, 10 μM, 72 h). Relative cell viability and relative cell death were calculated. h Clonogenic assay on HCT116-FoxO3A +/+ cells treated with increasing concentrations of metformin (24 h) and/or irinotecan (24 h), as indicated. e , h Cell growth percent inhibition at each drug concentration is presented. The data presented are the mean of at least three independent experiments. Where applicable, data are presented as mean ± SEM and significance was calculated with Student’s t test; * p

    Article Snippet: Compound C (5 μM, #171260), AICAR (5 mM, #A9978), PD98059 (0,02 mM, #P215), 2-deoxy-glucose (1 mM, #D8375), iodoacetic acid (0,5 μM, #I4386), metformin (10 μM, #D150959), 5-fluorouracil (2 μM, #F6627), cisplatin (30 μM, #P4394), etoposide (40 μM, #E1383) and irinotecan (30 μM, #347609) were all from Sigma; trametinib (#S2673) was from Selleckchem (Munich, German).

    Techniques: Transfection, Cycling Probe Technology, Reverse Transcription Polymerase Chain Reaction, Isolation, Derivative Assay, Mouse Assay, Fractionation, Clonogenic Assay, Inhibition, Concentration Assay