human fibroblast cell line hs 5  (ATCC)

Journal: Cell

Article Title: Pan-cancer analyses reveal cancer-type-specific fungal ecologies and bacteriome interactions

doi: 10.1016/j.cell.2022.09.005

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Article Snippet: HS-5 human fibroblast cell line , ATCC , Cat# CRL-11882.

Techniques: Negative Control, Construct, Purification, Staining, Polymer, Clinical Proteomics, Software

Journal: Frontiers in Oncology

Article Title: Stromal Fibroblasts Counteract the Caveolin-1-Dependent Radiation Response of LNCaP Prostate Carcinoma Cells

doi: 10.3389/fonc.2022.802482

Figure Lengend Snippet: Stromal fibroblasts limit the radiosensitizing effect of SRC inhibition by dasatinib in LNCaP spheroid co-cultures. CAV1 expressing stromal fibroblasts (HS5) were co-cultured with LNCaP control (Ctrl) cells expressing low endogenous CAV1 levels and the reporter green fluorescent protein (GFP) in hanging drops for 24 h. After formation of spheroids, cells were plated in growth factor-reduced Matrigel mixed with normal growth medium (1/2, v/v) supplemented with dasatinib or vehicle control (VC) and irradiated with 0 Gy or 10 Gy. Pictures were taken at the time of irradiation (0 h) and 48 h later. (A) Spheroid growth was measured and the respective volumes were calculated for 0 and 48 h after irradiation. Data were derived from 3 to 5 individual experiments, where at least 10 spheroids per condition and per experiment each were measured. (B) Cell death was analyzed afterwards by fluorescence microscopy using propidium iodide. Hoechst 33342 was used for nuclei staining. Representative fluorescent images from the individual experiments are shown (48 h time point). Scale bars represent 100 µm.

Article Snippet: The human PCa cell line LNCaP and the human skin fibroblast cell line HS5 were from ATCC (Manassas, VA) and cultured in RPMI Medium (Gibco, ThermoFisher, Waltham, MA) supplemented with 10% fetal bovine serum and 100 U/ml Penicillin/Streptomycin (Sigma-Aldrich, St. Louis, MO) under standard cell culture conditions (37°C, 5% CO2, 95% humidity).

Techniques: Inhibition, Expressing, Cell Culture, Control, Irradiation, Derivative Assay, Fluorescence, Microscopy, Staining

Journal: Frontiers in Oncology

Article Title: Stromal Fibroblasts Counteract the Caveolin-1-Dependent Radiation Response of LNCaP Prostate Carcinoma Cells

doi: 10.3389/fonc.2022.802482

Figure Lengend Snippet: Dasatinib treatment does not impact on survival or growth of stromal fibroblasts, nor in combination with RT. CAV1 expressing stromal fibroblasts (HS5) were cultured as monolayers (A, B) or spheroids (C, D) with or without radiation treatment (0 and 10 Gy) in the presence of dasatinib or vehicle control (VC), (A) Apoptotic cells (subG1) were analyzed 48 h post treatment by flow cytometry. Graphs consist of data from 3 individual experiments (with SEM). P-values indicate ***p ≤0.005 as estimated by one-way ANOVA with Tukey’s multiple comparison test. (B) Clonogenic survival was evaluated 10 days post treatment by counting respective colonies. Data show the surviving fractions from 3 independent experiments (means ± SD) plated in triplicates each. (C) Spheroid growth was measured and the respective volumes were calculated for 0 and 48 h after irradiation from 3 to 4 individual experiments where at least 10 spheroids per condition and per experiment each were measured. (D) Cell death was analyzed afterwards by fluorescence microscopy using propidium iodide. Hoechst 33342 was used for nuclei staining. Representative fluorescent images from individual experiments are shown (48 h time point). Scale bar represents 100 µm.

Article Snippet: The human PCa cell line LNCaP and the human skin fibroblast cell line HS5 were from ATCC (Manassas, VA) and cultured in RPMI Medium (Gibco, ThermoFisher, Waltham, MA) supplemented with 10% fetal bovine serum and 100 U/ml Penicillin/Streptomycin (Sigma-Aldrich, St. Louis, MO) under standard cell culture conditions (37°C, 5% CO2, 95% humidity).

Techniques: Expressing, Cell Culture, Control, Flow Cytometry, Comparison, Irradiation, Fluorescence, Microscopy, Staining

Journal: Frontiers in Oncology

Article Title: Stromal Fibroblasts Counteract the Caveolin-1-Dependent Radiation Response of LNCaP Prostate Carcinoma Cells

doi: 10.3389/fonc.2022.802482

Figure Lengend Snippet: RT treatment causes an increased metabolic potential of fibroblasts, an effect that is not affected upon SRC inhibition; LNCaP PCa cells are not affected. Extracellular flux analyses of LNCaP and fibroblast cell cultures were performed following RT treatment (0 and 10 Gy) in the presence of dasatinib or vehicle control (VC) 24 h post treatments. OCR (A) and ECAR (B) levels of LNCaP PCa cells are shown. Data were summarized as mean values ± SD (measured in 4–7 replicates each). (C) OCR levels of stromal HS5 fibroblasts are shown. (D) Basal respiration, ATP production and proton leak were depicted in separate bar diagrams. (E) Respective ECAR measurement over time are shown. (F) Basal and stressed (following oligomycin treatment) measurements of ECAR were depicted in separate bar diagrams. Data were summarized as mean values ± SD (measured in 4–7 replicates each). P-values indicate: *p ≤ 0.05, by one-way ANOVA with Tukey’s multiple comparison test and additionally by unpaired (two-tailed) t-tests depicted as # p ≤0.05.

Article Snippet: The human PCa cell line LNCaP and the human skin fibroblast cell line HS5 were from ATCC (Manassas, VA) and cultured in RPMI Medium (Gibco, ThermoFisher, Waltham, MA) supplemented with 10% fetal bovine serum and 100 U/ml Penicillin/Streptomycin (Sigma-Aldrich, St. Louis, MO) under standard cell culture conditions (37°C, 5% CO2, 95% humidity).

Techniques: Inhibition, Control, Comparison, Two Tailed Test

Journal: Leukemia

Article Title: Citrullination of histone H3 drives IL-6 production by bone marrow mesenchymal stem cells in MGUS and multiple myeloma

doi: 10.1038/leu.2016.187

Figure Lengend Snippet: ( a ) Confirmation of knockdown by siPADI2 by two independent siRNA sequences. ( b ) The expression of the BMMSC component of critical chemokine signalling axes in myeloma (IL-6, CXCL12 and cMET) is modulated by PADI2 expression in the immortalised BMMSC cell line, HS5. Expression of all three transcripts are reduced in response to reduction in PADI2 expression (siPADI2), or the addition of the pan-PADI inhibitor, Cl-Amidine (200 μ m , 72 h). However, siPADI4 does not alter expression of any of the transcripts. ** and *** indicate the level of significance assigned by a Dunn's comparison post-test after a significant ANOVA for each dataset ( P <0.0001 [IL-6] P =0.0002 [CXCL12] P =0.0019 [cMET]). ( c ) IL-6 expression, as measured through its secretion into the tissue culture medium by the HS5 cell line, is reduced after treatment with Cl-Amidine and/or siPADI2. Data shown are mean of n =3±s.d. * P =0.039, unpaired student's t -test with Welch's correction. ( d ) Overexpression of PADI2 increases IL-6 expression, whereas the catalytic mutant, C645S, does not increase expression over basal levels. Data shown are mean of n =3±s.d. ** P =0.004, unpaired student's t -test with Welch's correction.

Article Snippet: BMMSCs and the HS5 human bone marrow fibroblast cell line (ATCC CRL11882, LGC Standards, Teddington, UK, purchased December 2013, tested for mycoplasma every 3 months by PCR) were cultured in RPMI1640 medium with 10% fetal bovine serum (Hyclone, GE Healthcare, Little Chalfont, UK) at 37 °C in a humidified incubator with 5% CO 2 .

Techniques: Knockdown, Expressing, Comparison, Over Expression, Mutagenesis