kat2a primary cell based screening assays cell culture ht1080 human fibrosarcoma cell line ht1080  (ATCC)

Journal: Journal of orthopaedic research : official publication of the Orthopaedic Research Society

Article Title: The Histone Deacetylase Inhibitor Vorinostat Selectively Sensitizes Fibrosarcoma Cells to Chemotherapy

doi: 10.1002/jor.21274

Figure Lengend Snippet: Vorinostat inhibits soft tissue sarcoma and normal fibroblast proliferation. (A) Representative phase-contrast photomicrographs (×200) indicating decreased HT1080, SK-LMS-1, SW872 (STS), and WS-1 normal fibroblast proliferation and altered morphology after vorinostat treatment for 72 h. (B) STS and normal fibroblast proliferation were decreased by vorinostat treatment, as determined by counting cells after 24 h (white bars) or 72 h (black bars). Bars represent the mean and whiskers represent the SEM of at least four biological replicates. # and * denote statistical significance versus cells treated with vehicle after 24 or 72 h, respectively (p < 0.05).

Article Snippet: Cell Culture Human HT1080 fibrosarcoma, SK-LMS-1 leiomyosarcoma, and SW872 liposarcoma cells, and WS-1 normal human fibroblasts were obtained from the ATCC (Manassas, VA) and cultured in minimal essential media (Gibco, Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), nonessential amino acids (Gibco), sodium pyruvate (Gibco), and penicillin–streptomycin (Gibco) at 37°C under 5% CO 2 .

Techniques:

Journal: Journal of orthopaedic research : official publication of the Orthopaedic Research Society

Article Title: The Histone Deacetylase Inhibitor Vorinostat Selectively Sensitizes Fibrosarcoma Cells to Chemotherapy

doi: 10.1002/jor.21274

Figure Lengend Snippet: Vorinostat does not affect basal apoptosis levels in soft tissue sarcoma cells and normal fibroblasts. STS cells and normal fibroblasts were treated with 1.0 μM vorinostat or vehicle for 24 or 72 h and then trypsinized, fixed, stained for cleaved caspase 3 (BD Pharmingen™) and analyzed by flow cytometry. (A) Representative histograms of cleaved caspase 3 staining in HT1080 fibrosarcoma cells treated with vehicle or vorinostat. The negative control sample (first row, first histogram) was treated with vehicle for 72 h and then stained with the secondary Alexa Fluor® 647 antibody (Invitrogen) only. Puromycin (first row, second histogram) was used to induce apoptosis as a positive control for cleaved caspase 3 staining. (B) Vorinostat only moderately affects caspase 3 cleavage and then only at the supratherapeutic 3.0 μM dose in all cell lines tested. Bars represent the mean and whiskers represent the SEM of at least four biological replicates. # and * denote statistical significance versus cells treated with vehicle after 24 or 72 h, respectively (p < 0.05).

Article Snippet: Cell Culture Human HT1080 fibrosarcoma, SK-LMS-1 leiomyosarcoma, and SW872 liposarcoma cells, and WS-1 normal human fibroblasts were obtained from the ATCC (Manassas, VA) and cultured in minimal essential media (Gibco, Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), nonessential amino acids (Gibco), sodium pyruvate (Gibco), and penicillin–streptomycin (Gibco) at 37°C under 5% CO 2 .

Techniques: Staining, Flow Cytometry, Negative Control, Positive Control

Journal: Journal of orthopaedic research : official publication of the Orthopaedic Research Society

Article Title: The Histone Deacetylase Inhibitor Vorinostat Selectively Sensitizes Fibrosarcoma Cells to Chemotherapy

doi: 10.1002/jor.21274

Figure Lengend Snippet: Vorinostat sensitizes fibrosarcoma but not normal fibroblasts to doxorubicin. STS cells and normal fibroblasts were treated with 1.0 μM vorinostat or vehicle for 72 h. Doxorubicin was added at the beginning of the third day. (A) Representative (×200) phase-contrast and fluorescent photomicrographs of HT1080 cells plated in chamber slides, treated with vorinostat, doxorubicin, or both and then fixed and stained with anti-cleaved caspase 3 (BD Pharmingen™) and anti-rabbit Alexa Fluor® 488 (Invitrogen) antibodies. I: Phase contrast; II: cleaved caspase 3; III: doxorubicin; IV: DAPI. (B) Vorinostat and doxorubicin synergistically induce apoptosis in HT1080 but not SK-LMS-1 or SW872 STS cells, or WS-1 normal fibroblasts. After a 72-h treatment with vorinostat, doxorubicin, or both, cells were trypsinized, fixed, stained for cleaved caspase 3 and analyzed by flow cytometry. Bars represent the mean and whiskers represent the SEM of at least four biological replicates. Statistical significance is denoted by: * versus vehicle, # versus vorinostat alone, † versus doxorubicin alone.

Article Snippet: Cell Culture Human HT1080 fibrosarcoma, SK-LMS-1 leiomyosarcoma, and SW872 liposarcoma cells, and WS-1 normal human fibroblasts were obtained from the ATCC (Manassas, VA) and cultured in minimal essential media (Gibco, Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), nonessential amino acids (Gibco), sodium pyruvate (Gibco), and penicillin–streptomycin (Gibco) at 37°C under 5% CO 2 .

Techniques: Staining, Flow Cytometry

Journal: Journal of orthopaedic research : official publication of the Orthopaedic Research Society

Article Title: The Histone Deacetylase Inhibitor Vorinostat Selectively Sensitizes Fibrosarcoma Cells to Chemotherapy

doi: 10.1002/jor.21274

Figure Lengend Snippet: Vorinostat inhibits growth and induces chemosensitivity of fibrosarcoma xenografts. Crl:NU-Foxn1nu nude mice were injected subcutaneously with HT1080 cells. One week after inoculation, mice began receiving daily injections of vorinostat or vehicle and weekly injections of doxorubicin or vehicle. (A,B) HT1080 xenograft growth is inhibited by vorinostat alone and further inhibited by the combination of vorinostat and doxorubicin, but not by doxorubicin alone. Data are represented in (A) as the median and interquartile range and in (B) as the minima and maxima (whiskers) and the median, 25th and 75th percentiles (box) at day 21. N = 16 (Veh, VOR) or N = 9 (Veh + Dox, VOR + Dox). (C) Vorinostat treatment induces histone H3 accumulation in xenografts. Statistical significance is denoted by: *p < 0.05 versus vehicle, #p < 0.01 versus vehicle, †p < 0.01 versus doxorubicin alone.

Article Snippet: Cell Culture Human HT1080 fibrosarcoma, SK-LMS-1 leiomyosarcoma, and SW872 liposarcoma cells, and WS-1 normal human fibroblasts were obtained from the ATCC (Manassas, VA) and cultured in minimal essential media (Gibco, Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), nonessential amino acids (Gibco), sodium pyruvate (Gibco), and penicillin–streptomycin (Gibco) at 37°C under 5% CO 2 .

Techniques: Injection

Journal: Journal of orthopaedic research : official publication of the Orthopaedic Research Society

Article Title: The Histone Deacetylase Inhibitor Vorinostat Selectively Sensitizes Fibrosarcoma Cells to Chemotherapy

doi: 10.1002/jor.21274

Figure Lengend Snippet: Combined vorinostat and doxorubicin treatment induces cell death in fibrosarcoma xenografts. (A) Cleaved caspase 3 immunohistochemistry revealed that cell death was greatest in HT1080 xenografts from vorinostat/doxorubicin-treated mice (upper panel). The high magnification inset indicates cleaved caspase 3-positive apoptotic cells in the xenograft from vorinostat/doxorubicin-treated mice. TUNEL staining (middle panel) confirmed that vorinostat/doxorubicin induced more apoptosis in HT1080 compared to vehicle, doxorubicin alone, or vorinostat alone. Nuclei were counterstained using DAPI (lower panel). (B) Quantitation of TUNEL staining in HT1080 xenografts. Statistical significance is denoted by *p < 0.05 versus vehicle, #p < 0.05 versus doxorubicin alone, †p < 0.01 versus vorinostat alone. Scale bar = 50 μm.

Article Snippet: Cell Culture Human HT1080 fibrosarcoma, SK-LMS-1 leiomyosarcoma, and SW872 liposarcoma cells, and WS-1 normal human fibroblasts were obtained from the ATCC (Manassas, VA) and cultured in minimal essential media (Gibco, Life Technologies, Grand Island, NY) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), nonessential amino acids (Gibco), sodium pyruvate (Gibco), and penicillin–streptomycin (Gibco) at 37°C under 5% CO 2 .

Techniques: Immunohistochemistry, TUNEL Assay, Staining, Quantitation Assay

Journal: Journal of cell science

Article Title: Role of membrane-tension gated Ca 2+ flux in cell mechanosensation.

doi: 10.1242/jcs.208470

Figure Lengend Snippet: Fig. 1. Response of HT1080 cells to mechanical compression. (A) Experimental setup for the compression device. Fabrication of the device follows the method in Si et al. (2015). Cells in the device are compressed by the application of air pressure above the PDMS layer. The compression depth is limited by the support pillars with height 4–6 µm. (B) Epifluorescence image of cell (YFP channel) before and after compression, and after the release of compression. The cross-section area of the cell increases under compression, and decreases after the release of compression. Scale bars: 20 µm. (C) Scaled FRET-to-CFP ratio (FRET/CFP) and cell spreading area over time. Compression takes place at t=0 min (indicated by the black arrow). Cells are then released at t=38 min (indicated by the red arrow). Results are mean±s.e.m. (n=40). (D) Summary of mean±s.e.m. FRET/CFP when cell is uncompressed, compressed and released from compression. All results are scaled to the mean value of uncompressed cells and represent three biological replicates (n=40 cells). For each biological replicate, there is one technical replicate. ***P<0.001; n.s., not significant (Student’s t-test).

Article Snippet: Cell culture Human fibrosarcoma HT1080 (ATCC) cells were cultured in high glucose (4.5 g/l) Dulbecco’s modified Eagle’s medium (Mediatech), supplemented with 10% fetal bovine serum (Hyclone) and 1% penicillin-streptomycin (Sigma).

Techniques:

Journal: Journal of cell science

Article Title: Role of membrane-tension gated Ca 2+ flux in cell mechanosensation.

doi: 10.1242/jcs.208470

Figure Lengend Snippet: Fig. 4. Intracellular Ca2+ dye signal after cells are treated with TRPV4 inhibitor or incubated in Ca2+-free medium for 30 min. (A) Image of a cell in the DIC channel and the fluorescence channel showing Ca2+ dye in the cytoplasm. Scale bar: 20 µm. (B) Total intracellular Ca2+ dye signal over time for HT1080 cells with no treatment (control), TRPV4 inhibitor treatment and Ca2+-free medium treatment. The Ca2+ dye signal is generally stable for 15 min after imaging. The slight decrease in signal is due to dye leakage out of the cell, and occurs for all conditions. (C) Population and time average of total intracellular Ca2+ dye signal for control, TRPV4 inhibitor treatment and Ca2+-free medium treatment. Results are mean±s.e.m.; the number of cells in each set of experiments is labeled in C; there were two biological repeats and two technical repeats for each biological repeat in all conditions. ***P<10−9 (Student’s t-test).

Article Snippet: Cell culture Human fibrosarcoma HT1080 (ATCC) cells were cultured in high glucose (4.5 g/l) Dulbecco’s modified Eagle’s medium (Mediatech), supplemented with 10% fetal bovine serum (Hyclone) and 1% penicillin-streptomycin (Sigma).

Techniques: Incubation, Fluorescence, Control, Imaging, Labeling

Journal: Oncology Letters

Article Title: Regulation of radiosensitivity by 4-methylumbelliferone via the suppression of interleukin-1 in fibrosarcoma cells

doi: 10.3892/ol.2019.9990

Figure Lengend Snippet: Radiosensitization by 4-MU. (A) Clonogenic potency of HT1080 cells, (B) representative dot plots of FITC-Annexin V and PI staining, and (C) quantitative analysis of apoptotic cell fractions. Values are presented as the mean ± standard deviation. *P<0.05 and **P<0.01 vs. control. 4-MU, 4-methylumbelliferone; FITC, fluorescein isothiocyanate; PI, propidium iodide.

Article Snippet: Cell culture HT1080 human fibrosarcoma cells from American Type Culture Collection (Manassas, VA, USA) were cultured in Roswell Park Memorial Institute 1640 medium (Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Japan Bio Serum, Fukuyama, Japan) and 1% penicillin/streptomycin (Life Technologies) at 37°C in a humidified atmosphere of 5% CO 2 .

Techniques: Staining, Standard Deviation, Control

Journal: Oncology Letters

Article Title: Regulation of radiosensitivity by 4-methylumbelliferone via the suppression of interleukin-1 in fibrosarcoma cells

doi: 10.3892/ol.2019.9990

Figure Lengend Snippet: Analysis of signal transduction pathways and upstream genes, altered by X-ray irradiation with or without 4-MU treatment. HT1080 cells treated with 500 µM 4-MU and/or 2 Gy X-ray irradiation were cultured for 24 h. (A) Heatmaps of canonical pathways and (B) upstream genes were generated from z-scores calculated with Ingenuity Pathway Analysis software based on the results of global gene expression profiling. Red, green and white squares represent z-score values of ‘2>’ and ‘2<’ and ‘no data’, respectively. 4-MU, 4-methylumbelliferone.

Article Snippet: Cell culture HT1080 human fibrosarcoma cells from American Type Culture Collection (Manassas, VA, USA) were cultured in Roswell Park Memorial Institute 1640 medium (Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Japan Bio Serum, Fukuyama, Japan) and 1% penicillin/streptomycin (Life Technologies) at 37°C in a humidified atmosphere of 5% CO 2 .

Techniques: Transduction, Irradiation, Cell Culture, Generated, Software, Gene Expression

Journal: Oncology Letters

Article Title: Regulation of radiosensitivity by 4-methylumbelliferone via the suppression of interleukin-1 in fibrosarcoma cells

doi: 10.3892/ol.2019.9990

Figure Lengend Snippet: Validation of microarray results. (A) HT1080 cells treated with 500 µM 4-MU and/or 2 Gy X-ray irradiation were cultured for 24 h. mRNA levels of IL-1α, −1β, and −6 were evaluated by reverse transcription-quantitative polymerase chain reaction. (B) Supernatants of HT1080 cells cultured in the absence of serum with 500 µM 4-MU and/or 2 Gy X-ray irradiation for 24 h were collected and assayed by ELISA to determine IL-1α, −1β and −6 concentrations. Values represent the mean ± standard deviation of the mean. *P<0.05 and **P<0.01, as indicated. 4-MU, 4-methylumbelliferone; IL, interleukin.

Article Snippet: Cell culture HT1080 human fibrosarcoma cells from American Type Culture Collection (Manassas, VA, USA) were cultured in Roswell Park Memorial Institute 1640 medium (Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Japan Bio Serum, Fukuyama, Japan) and 1% penicillin/streptomycin (Life Technologies) at 37°C in a humidified atmosphere of 5% CO 2 .

Techniques: Biomarker Discovery, Microarray, Irradiation, Cell Culture, Reverse Transcription, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Oncology Letters

Article Title: Regulation of radiosensitivity by 4-methylumbelliferone via the suppression of interleukin-1 in fibrosarcoma cells

doi: 10.3892/ol.2019.9990

Figure Lengend Snippet: Flow cytometry analysis of inflammatory cytokine receptor expression. HT1080 cells treated with 500 µM 4-MU and/or 2 Gy X-ray irradiation were cultured for 24 h. Cells were labeled with fluorophore-conjugated antibodies and analyzed by flow cytometry. Histograms of (A) PE-CD121a, (B) PE-CD126 and (C) APC-CD130, and (D) the quantitative analysis of mean fluorescence intensity. Values represent the mean ± standard deviation of the mean. *P<0.05 and **P<0.01 vs. control/as indicated. 4-MU, 4-methylumbelliferone; PE, phycoerythrin; CD, cluster of differentiation; APC, allophycocyanin.

Article Snippet: Cell culture HT1080 human fibrosarcoma cells from American Type Culture Collection (Manassas, VA, USA) were cultured in Roswell Park Memorial Institute 1640 medium (Thermo Fisher Scientific, Inc., Waltham, MA, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Japan Bio Serum, Fukuyama, Japan) and 1% penicillin/streptomycin (Life Technologies) at 37°C in a humidified atmosphere of 5% CO 2 .

Techniques: Flow Cytometry, Expressing, Irradiation, Cell Culture, Labeling, Fluorescence, Standard Deviation, Control

Journal: American Journal of Cancer Research

Article Title: Vascular-endothelial response to IDH1 mutant fibrosarcoma secretome and metabolite: implications on cancer microenvironment

doi:

Figure Lengend Snippet: Representative images and quantification of vascular-endothelial tube formation in basal medium and the human fibrosarcoma secretome. A: Vascular-endothelial cells treated with basal medium showing sparse tube formation. B: The fibrosarcoma secretome stimulated abundant vascular-endothelial tube formation. Scale bars: 400 μm. C: Quantification of vascular-endothelial tube lengths in basal medium and the fibrosarcoma secretome. Rel.: relative; s.e.: standard error; neg.: negative; ctrl.: control; n=3 independent biological samples per group; ***: P < 0.001.

Article Snippet: Cell culture Human fibrosarcoma cells (HT-1080) were acquired from ATCC (Cat. #CCL-121; Manassas, VA, USA).

Techniques: Control

Journal: American Journal of Cancer Research

Article Title: Vascular-endothelial response to IDH1 mutant fibrosarcoma secretome and metabolite: implications on cancer microenvironment

doi:

Figure Lengend Snippet: Representative images and quantification of vascular-endothelial tube formation in the cancer secretome and a small molecule (SM)-treated cancer secretome. A: Abundant vascular-endothelial tubes induced by the fibrosarcoma secretome. B: Substantial reduction in vascular-endothelial tubes induced by the GSK864-treated fibrosarcoma secretome. SM: small molecule, GSK864; Scale bars: 400 μm. C: Quantification of vascular-endothelial tube lengths in the fibrosarcoma secretome and a small molecule (SM)-treated fibrosarcoma secretome. SM: GSK864. Rel.: relative; s.e.: standard error; pos.: positive; ctrl.: control; n=3 independent biological samples per group; ***: P < 0.001.

Article Snippet: Cell culture Human fibrosarcoma cells (HT-1080) were acquired from ATCC (Cat. #CCL-121; Manassas, VA, USA).

Techniques: Control

Journal: American Journal of Cancer Research

Article Title: Vascular-endothelial response to IDH1 mutant fibrosarcoma secretome and metabolite: implications on cancer microenvironment

doi:

Figure Lengend Snippet: Representative images and quantification of vascular-endothelial tube formation in the fibrosarcoma secretome, a small molecule (SM)-treated fibrosarcoma secretome, and 2HG supplemented and SM-treated fibrosarcoma secretome. A: The fibrosarcoma secretome induced abundant vascular-endothelial tube formation. B: The GSK864-treated fibrosarcoma secretome reduced tube formation. C: 2HG supplemented in the SM-treated fibrosarcoma secretome enhanced tube formation. Scale bars: 400 μm. D: Quantification of vascular-endothelial tube lengths in the fibrosarcoma secretome, a small molecule (SM)-treated fibrosarcoma secretome (by GSK864) and the 2HG supplemented, GSK864-treated fibrosarcoma secretome. Rel.: relative; s.e.: standard error; pos.: positive; ctrl.: control; n=6 independent biological samples per group; ***: P < 0.001.

Article Snippet: Cell culture Human fibrosarcoma cells (HT-1080) were acquired from ATCC (Cat. #CCL-121; Manassas, VA, USA).

Techniques: Control

Journal: American Journal of Cancer Research

Article Title: Vascular-endothelial response to IDH1 mutant fibrosarcoma secretome and metabolite: implications on cancer microenvironment

doi:

Figure Lengend Snippet: Representative images and quantification of vascular-endothelial tube formation in the fibrosarcoma secretome and 2HG in endothelial basal medium. A: Abundant vascular-endothelial tube formation induced by the fibrosarcoma secretome. B: 2HG in endothelial basal medium induced similar vascular-endothelial tube formation. Scale bars: 400 μm. C: Quantification of vascular-endothelial tube lengths in the fibrosarcoma secretome and 2HG in endothelial basal medium. Rel.: relative; s.e.: standard error; pos.: positive; ctrl.: control; n=3 independent biological samples per group; n.s.: no (statistical) significance.

Article Snippet: Cell culture Human fibrosarcoma cells (HT-1080) were acquired from ATCC (Cat. #CCL-121; Manassas, VA, USA).

Techniques: Control

Journal: American Journal of Cancer Research

Article Title: Vascular-endothelial response to IDH1 mutant fibrosarcoma secretome and metabolite: implications on cancer microenvironment

doi:

Figure Lengend Snippet: Cell proliferation. A: 2-hydroxyglutarate (2HG) stimulated human fibrosarcoma cell (HT-1080) proliferation beyond its baseline in a dose-dependent manner. s.d.: standard deviation. n=5 independent biological samples. B: GSK864 inhibited the proliferation of both human fibroblasts and human fibrosarcoma cells (HT-1080) in a dose-dependent manner. s.d.: standard deviation. Proliferation of human fibrosarcoma cells (HT-1080) and fibroblasts at 0.8, 1.6 and 2 μmol GSK864 concentrations were significantly lower than control (ctrl) and 0.2 and 0.4 μmol (P < 0.001; n=5 independent biological samples per group).

Article Snippet: Cell culture Human fibrosarcoma cells (HT-1080) were acquired from ATCC (Cat. #CCL-121; Manassas, VA, USA).

Techniques: Standard Deviation, Control