Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: Cytotoxicity of ultravist in P815 cells. P815 cells were incubated with different doses of ultravist for 24 h and cell viability was measured by the CCK-8 test. * P<0.05 vs. control.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques: Incubation, CCK-8 Assay, Control

Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: Mast cells could be activated by ultravist stimulation. P815 cells were stimulated with ultravist for 24 h; (A) histamine and (B) β-hexosaminidase were measured by ELISA. * P<0.05, ** P<0.01.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: miRNA expression profile of ultravist-stimulated mast cells. P815 cells were treated with 50 mg/ml ultravist for 24 h. miRNA from the control and ultravist treatment groups was extracted and analyzed by miRNA array. miRNA/miR, microRNA.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques: Expressing, Control

Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: KEGG Pathway Analysis of ultravist-treated mast cells. P815 cells were stimulated with 50 mg/ml ultravist for 24 h. Subsequent KEGG pathway analysis of the mTOR signaling pathway revealed upregulation of genes related to (A) miR-19A-3p and (B) miR-362-3p, which are highlighted in red. KEGG, Kyoto Encyclopedia of Genes and Genomes; miRNA/miR, microRNA.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques:

Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: miR-19a-3p and miR-362-3p could be upregulated by ultravist treatment. P815 cells were treated with different doses of ultravist and (A) miR-19a-3p and (B) miR-362-3p were detected by reverse transcription-quantitative PCR. * P<0.05, ** P<0.01. miRNA/miR, microRNA.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques: Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Biomedical Reports

Article Title: Radiocontrast medium induces histamine release in association with upregulation of miR‑19a‑3p and miR‑362‑3p expression

doi: 10.3892/br.2024.1780

Figure Lengend Snippet: Network analysis of miRNA-target interactions in mast cells. P815 cells were stimulated with 50 mg/ml ultravist for 24 h, the predicted interactions between miR-19a-3p and miR-362-3p were analyzed and the key target genes-Aox1, Akap2, and Uba2l were highlighted in yellow. miRNA/miR, microRNA.

Article Snippet: The mouse mast cell cell line, P815 (Elabscience Biotechnology, Inc.), was cultured in RPMI1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (Cytiva) and 1% penicillin and streptomycin (Gibco; Thermo Fisher Scientific, Inc.) at 37 ̊C incubator with 5% CO 2 .

Techniques:

Journal: Nature communications

Article Title: Distinctive phenotypes and functions of innate lymphoid cells in human decidua during early pregnancy.

doi: 10.1038/s41467-019-14123-z

Figure Lengend Snippet: Fig. 6 dNK responses triggered by activating KIR. a Representative XCL1 and CD107a staining by CyTOF in lineage negative (Lin−) CD56+ decidual cells co-expressing 1–3 Killer-cell immunoglobulin-like receptors (KIR) following a 6 h co-culture with K562 (n = 10). b Representative XCL1 and CD107a staining by mass cytometry in Lin-CD56+ cells co-expressing 1–3 KIR following 4 h stimulation by PMA plus ionomycin (n = 8). c Representative XCL1 staining by flow cytometry in Lin-CD56+KIR2DS4+ decidual cells co-expressing 1–3 additional KIRs following activation via P815 cells coated with anti-KIR2DS4 (n = 6). d Correlation of frequency of XCL1+Lin-CD56+ decidual cells and mean side scatter for the same subset (n = 20). Two-tailed p-value calculated for Pearson correlation coefficients. Two-tailed one-way ANOVA of matched data points with Tukey correction and 95% confidence level was used. *p < 0.05, **p < 0.01, ***p < 0.001. Source data are provided as a Source Data file.

Article Snippet: The murine cell line P815 and human cell line K562 were purchased from DSMZ in Germany.

Techniques: Staining, Expressing, Co-Culture Assay, Mass Cytometry, Cytometry, Activation Assay, Two Tailed Test

Journal: Oncotarget

Article Title: Crenolanib is a type I tyrosine kinase inhibitor that inhibits mutant KIT D816 isoforms prevalent in systemic mastocytosis and core binding factor leukemia

doi: 10.18632/oncotarget.19970

Figure Lengend Snippet: Dilution series of crenolanib to treat HMC1.2 or p815 mastocytosis cell lines were set up and inhibition of cellular proliferation (A, B) or induction of apoptosis (C, D) was assessed. Samples were compared using one-way ANOVA and Tukey’s honestly significant difference (HSD) test. Values in black indicate significance compared to the untreated controls.

Article Snippet: Notably, sensitivity profiles for crenolanib obtained in the Ba/F3 isogenic cell model corresponds to phosphoprotein sensitivities in the native mastocytosis cell lines HMC1.2 (harboring KIT D816V, Figure ) and p815 (harboring murine Kit D814Y, which is homologous to human KIT D816Y, Figure ).

Techniques: Inhibition

Journal: Oncotarget

Article Title: Crenolanib is a type I tyrosine kinase inhibitor that inhibits mutant KIT D816 isoforms prevalent in systemic mastocytosis and core binding factor leukemia

doi: 10.18632/oncotarget.19970

Figure Lengend Snippet: Isogenic Ba/F3 cells harboring a mutant-KIT D816V isoform ( A , left and mid panels), the corresponding mastocytosis cell line HMC1.2 (D816V+, A , right panel), Ba/F3 cells harboring a mutant-KIT D816Y isoform ( B , left panel) and the corresponding mastocytosis cell line p815 (murine D814Y, B, right panel) were treated with crenolanib or dasatinib (A) in dose-dilution assays for 90‘. Western immunoblots are shown for inhibition of autophosphorylation of KIT and consecutive downstream signaling via ERK1/2, STAT5 and AKT. Two different phospho-KIT antibodies (monoclonal Tyr703, 145 KDa, polyclonal Tyr719, 120/145 KDa) provide similar results via dephosphorylation of KIT (not degradation as indicated by stable KIT blots) upon crenolanib as well as dasatinib exposure. Tubulin serves as loading control.

Article Snippet: Notably, sensitivity profiles for crenolanib obtained in the Ba/F3 isogenic cell model corresponds to phosphoprotein sensitivities in the native mastocytosis cell lines HMC1.2 (harboring KIT D816V, Figure ) and p815 (harboring murine Kit D814Y, which is homologous to human KIT D816Y, Figure ).

Techniques: Mutagenesis, Western Blot, Inhibition, De-Phosphorylation Assay, Control

Journal: Oncotarget

Article Title: Crenolanib is a type I tyrosine kinase inhibitor that inhibits mutant KIT D816 isoforms prevalent in systemic mastocytosis and core binding factor leukemia

doi: 10.18632/oncotarget.19970

Figure Lengend Snippet: HMC1.2 (A/B) or p815 (C/D) were sequentially treated with one or both agents and induction of apoptosis was assessed using an annexin V-based flow cytometry assay. (E) Validation in native mastocytosis patient samples (n=11) treated with 2-CDA followed by crenolanib (Cb). Reduction of the CD25+ viable cohort was assessed flow cytometrically. A normalized (untreated set as 1) dose-effect plot is shown. Significance of the combination compared to the monoagents is provided using Tukey Kramer test and Student’s t-test.

Article Snippet: Notably, sensitivity profiles for crenolanib obtained in the Ba/F3 isogenic cell model corresponds to phosphoprotein sensitivities in the native mastocytosis cell lines HMC1.2 (harboring KIT D816V, Figure ) and p815 (harboring murine Kit D814Y, which is homologous to human KIT D816Y, Figure ).

Techniques: Flow Cytometry, Biomarker Discovery

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Role of regulatory T cells and checkpoint inhibition in hepatocellular carcinoma

doi: 10.1007/s00262-019-02427-4

Figure Lengend Snippet: Expression of PD-1/PD-L1 on HCC tumour tissues and tumour cell lines. a Representative images of immunohistochemistry sections in the tumour tissue of HCC patient #20. Positive cells are stained brown by immunoperoxidase. Arrows mark PD-1-positive mononuclear cells (upper left), CD8+ T cells (upper right) and perforin-positive cells (lower right), respectively. PD-L1 staining shown in the lower-left panel indicates a greater number of positive mononuclear cells (macrophages and CD8+ T cells). Of note, HCC tumour cells did not stain positive for PD-1 and PD-L1. Tu tumour. b The flow cytometric analysis of PD-L1/L2 expression on P815 mouse mastocytoma cells in contrast to various hepatoma cell lines (HepG2, HepT1, Huh4, Hep3B, Huh7). Unlike hepatoma cells, which up-regulated PD-L1 and PD-L2, P815 cells remained PD-L1 and PD-L2 negative in cell culture. Thus, we used P815 cells as target cells in lectin-dependent cellular cytotoxicity (LDCC) assays to study the functional role of Tregs on T cell functions in a model of PD-L1/L2-negative tumour cells

Article Snippet: Cell line authentication of murine P815 cells was performed by the Swiss DNA company Microsynth (Supplementary Table 3).

Techniques: Expressing, Immunohistochemistry, Staining, Cell Culture, Functional Assay

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Role of regulatory T cells and checkpoint inhibition in hepatocellular carcinoma

doi: 10.1007/s00262-019-02427-4

Figure Lengend Snippet: IFN-gamma production and degranulation by CD8+ T cells in lectin-dependent cellular cytotoxicity (LDCC) assays and effects of checkpoint inhibition. a The gating strategy to analyse IFN-gamma production and T cell degranulation in CD8+ T effector cells by flow cytometry in co-cultures with Con A-loaded P815 cells and Tregs (Treg to Teff ratio: 1:2). b IFN-gamma production (left) and CD107a degranulation (right) of CD8+ T effector cells in LDCC assays before (BL), with exposure to Con A-loaded P815 target cells (stim = peak secretion) and upon adding Tregs (stim + Tregs). Adding autologous Tregs reduced P815-induced peak IFN-gamma secretion and degranulation of CD8+ T cells in all study groups. p values refer to significances obtained by paired Student t test marked by bars. c Provides the summary statistics concerning the differences of IFN-gamma-production (left) and CD107a degranulation (right) by CD8+ T cells after sequential addition of Con A-loaded P815 target cells and autologous Tregs (Treg to Teff ratio: 1:2), as well as in the presence of added neutralizing anti-PD-1, anti-PD-L1, and anti-CTLA-4, and anti-GITR (10 µg/ml each). Columns represent the mean ± SD of 6–18 different donors. Anti-PD-1 and anti-PD-L1—but not anti-CTLA-4 nor anti-GITR—partially reversed Treg-associated inhibition of IFN-gamma secretion in CD8+ T cells from patients with HCC. Treg-mediated inhibition of T cell degranulation was not reversed by any of the antibodies. p values refer to significances obtained by paired Student t test marked by bars

Article Snippet: Cell line authentication of murine P815 cells was performed by the Swiss DNA company Microsynth (Supplementary Table 3).

Techniques: Inhibition, Flow Cytometry

Journal: Frontiers in Immunology

Article Title: NLRP3 Inflammasome Activation of Mast Cells by Estrogen via the Nuclear-Initiated Signaling Pathway Contributes to the Development of Endometriosis

doi: 10.3389/fimmu.2021.749979

Figure Lengend Snippet: NLRP3 expression was increased by estrogen stimulation via ER-α in MCs. (A) Bar plot of differentially expressed genes (DEGs) from the KEGG pathway enrichment analysis; a comparison between β-estradiol treated P815 cells and the control. (B) A volcano plot was used for visualization of the differentially expressed genes. The gray point in the plot represents genes with no statistical differences (-1< log 2 (FC) < 1, P > 0.05), the red point represents upregulated genes (log 2 (FC) > 1, P < 0.05), and the blue point represents downregulated genes (log 2 (FC) < -1, P < 0.05) with statistical significance. (C) Quantitative PCR analysis of ESR1 in P815 cells after ESR1-specific shRNA transfection. (D) Western blot analysis of ER-α in P815 cell nuclei after stable transfection of ESR1-specific shRNA. (E) Relative quantification of NLRP3 mRNA level in P815 was measured by FPKM reads using RNA-seq. (F) Quantitative PCR analysis NLRP3 mRNA levels in P815 cells treated with β-estradiol after knockdown of ESR1 compared with the negative control. (G) Representative images of immunofluorescence staining for NLRP3 (red) in mast cell line P815 incubated with or without 100 pmol/mL β-estradiol for 24 h Data in (C, D, F) represent three independent experiments. Data are expressed as mean ± SEM. Statistics: Unpaired Student’s t-test. * P < 0.05, ** P < 0.01, *** P < 0.001; ns, statistically not significant.

Article Snippet: Mouse mast cell line P815 from ScienCell Research Laboratories (CA, USA) was cultured with RPMI-1640 (Gibco, USA) supplemented with 10% fetal bovine serum (FBS) and 1% Penicillin-Streptomycin (PS) (Thermo Fisher).

Techniques: Expressing, Real-time Polymerase Chain Reaction, shRNA, Transfection, Western Blot, Stable Transfection, RNA Sequencing Assay, Negative Control, Immunofluorescence, Staining, Incubation

Journal: Frontiers in Immunology

Article Title: NLRP3 Inflammasome Activation of Mast Cells by Estrogen via the Nuclear-Initiated Signaling Pathway Contributes to the Development of Endometriosis

doi: 10.3389/fimmu.2021.749979

Figure Lengend Snippet: ERE sites in the NLRP3 promoter mediated estrogen-induced NLRP3 transcription. (A) ChIP analysis was performed using anti-ER-α or anti-Histone H3 antibody to ascertain the existence of the ERE in the promoter of the NLRP3 gene. The PCR results showed that a 159-bp fragment containing the presumed ERE could be precipitated after P815 cells were treated with β-estradiol for 24 h. (B) The pulled-down band was excised from the gel and sequenced. (C) Schematic diagram of luciferase reporter constructs. Basic-Luc: pGL4-basic plasmid; NLRP3-Luc: pGL4-basic plasmid with the NLRP3 promoter fragment including presumed ERE-like sequence; delNLRP3-Luc: pGL4-basic plasmid with the mutant NLRP3 promoter fragment, with the presumed ERE-like sequence deleted. (D) Luciferase activities of three report systems with or without ESR1 plasmid co-transfection in 293T cells were compared with each other. Renilla luciferase plasmid was used to normalize transfection efficiencies. The experiments were repeated three times and data are presented as means ± SEM. Statistics: Unpaired Student’s t-test. * P < 0.05, ** P < 0.01.

Article Snippet: Mouse mast cell line P815 from ScienCell Research Laboratories (CA, USA) was cultured with RPMI-1640 (Gibco, USA) supplemented with 10% fetal bovine serum (FBS) and 1% Penicillin-Streptomycin (PS) (Thermo Fisher).

Techniques: Luciferase, Construct, Plasmid Preparation, Sequencing, Mutagenesis, Cotransfection, Transfection

Journal: Frontiers in Immunology

Article Title: NLRP3 Inflammasome Activation of Mast Cells by Estrogen via the Nuclear-Initiated Signaling Pathway Contributes to the Development of Endometriosis

doi: 10.3389/fimmu.2021.749979

Figure Lengend Snippet: The NLRP3 inflammasome pathway was activated by estrogen via ER-α. (A) Western blot analysis of NLRP3, cleaved caspase-1, caspase-1 precursor, cleaved IL-1β, IL-1β precursor, and ASC in P815 cells treated with 100 pmol/mL β-estradiol for 1 h, 3 h, 6 h, 12 h, and 24 h. (B) Quantified results of western blot assays. (C) Intracellular concentration of K + was determined by the ratio of the fluorescence intensities obtained by exciting PBFI at 340/380 nm wavelengths while monitoring emission at 500 nm, and presented as percent of untreated control. (D) The concentration of IL-1β in the P815 cell culture supernatant was tested using ELISA. Data in (B–D) represent three independent experiments. Data are expressed as mean ± SEM. Statistics: ANOVA followed by Dunnett t-test. * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: Mouse mast cell line P815 from ScienCell Research Laboratories (CA, USA) was cultured with RPMI-1640 (Gibco, USA) supplemented with 10% fetal bovine serum (FBS) and 1% Penicillin-Streptomycin (PS) (Thermo Fisher).

Techniques: Western Blot, Concentration Assay, Fluorescence, Cell Culture, Enzyme-linked Immunosorbent Assay