Journal: Molecular Cancer Research

Article Title: 5-Hydroxy-2-Methyl-1,4-Naphthoquinone, a Vitamin K3 Analogue, Suppresses STAT3 Activation Pathway through Induction of Protein Tyrosine Phosphatase, SHP-1: Potential Role in Chemosensitization

doi: 10.1158/1541-7786.mcr-09-0257

Figure Lengend Snippet: FIGURE 6. A. Overexpression of constitutive STAT3 rescues A293 cells from plumbagin-induced cytotoxicity. First, A293 cells were transfected with constitutive STAT3 plasmid. After 24 h of transfection, the cells were treated with 2.5 μmol/L plumbagin for 24 h, the cytotoxicity was determined by Live/Dead assay, and 20 random fields were counted. B. Plumbagin potentiates the apoptotic effect of thalidomide and bortezomib. U266 cells (1 × 106) were treated with 2.5 μmol/L plumbagin, and 10 μg/mL thalidomide or 20 nmol/L bortezomib alone or in combination for 24 h at 37°C. Cells were stained with a Live/Dead assay reagent for 30 min and then analyzed under a fluorescence microscope. The results shown are representative of two independent experiments.

Article Snippet: Bortezomib (Velcade, PS-341) was obtained from Millennium, and thalidomide was from Tocris Cookson.

Techniques: Over Expression, Transfection, Plasmid Preparation, Live Dead Assay, Staining, Fluorescence, Microscopy

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide impairs DC cross-priming. Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to ovalbumin (50 μ g/ml) or PBS pulse. (a) Mixed lymphocyte reaction was performed by incubating these cells with the same H-2 background splenocytes at the ratio of 1 : 10. The ability of T cell proliferation was accessed by BrdU cell proliferation assay. IL-12 concentration in supernatants of mixed lymphocyte reaction was determined by ELISA. (b) C57BL/6 mouse was intraperitoneally transferred with these 1 × 10 4 cells and Ag-specific IFN- γ ELISPOT assays, and (c) granzyme B expression determination was performed 5 d after adoptive transfer. The ELISPOT data were presented as spot-forming units per million cells. For qPCR, β -actin was used as an internal control. Data were presented as the mean ± SEM, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. Tha: thalidomide; PYR: PYR-41.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, BrdU Cell Proliferation Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Enzyme-linked Immunospot, Expressing, Adoptive Transfer Assay, Control

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: PYR-41 and thalidomide do not increase CD80/CD86 expression in long-term- and short-term-treated conditions. Murine bone marrow-derived DC (cultured for 4 d) conferred PYR-41 (5 μ M), thalidomide (30 μ M), LPS (10 ng/ml), or DMSO stimulation for 12 (a, b) or 2 (c–f) hrs. The expressions of CD80 and CD86 were determined by flow cytometry. Data were presented as the mean ± SEM, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Expressing, Derivative Assay, Cell Culture, Flow Cytometry

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide has not increased IL-12 secretion. Murine bone marrow-derived DC (cultured for 4 d) conferred PYR-41 (5 μ M), thalidomide (30 μ M), LPS (10 ng/ml), or DMSO stimulation for 12 hrs, and IL-12 expression was determined by flow cytometry with intracellular IL-12 staining. The positive percentages (a) and the mean fluorescence intensity (b) of analyzed population were shown. Data were presented as the mean ± SEM and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Expressing, Flow Cytometry, Staining, Fluorescence

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide inhibits DC cross-presentation. Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to ovalbumin (50 μ g/ml) or PBS pulse. The effects of PYR-41 and thalidomide on cross-presentation were determined by flow cytometric analyses (a) and confocal microscope (b), respectively. For immunofluorescence observations, cross-presented OVA was stained with red (25-D1.16), EEA1 and Rab7 are all stained with green, and nuclei were counterstained with blue (DAPI). Original magnification, ×600. Data were presented as the mean ± SEM, ∗ p < 0.05, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Microscopy, Immunofluorescence, Staining

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide abolishes the endosomal recruitment of p97. Murine bone marrow-derived DC (cultured for 4 d) firstly conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to ovalbumin (50 μ g/ml) or PBS pulse. The relocation of p97 from endoplasmic reticulum to endosomes (a) was determined by confocal microscope by Rab5, calnexin, and p97 antibody staining. The colocalized plots of p97 with Rab5 and calnexin (b) were counted and analyzed. The expressions of Rab5/p97 and calnexin/p97 in the cells which are corresponding to the (a) colocalized cells were shown (c). Nuclei were counterstained with DAPI (blue). Original magnification, ×600. Data were presented as the mean ± SEM, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown. Rab5: early endosome marker; calnexin: endoplasmic reticulum marker.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Microscopy, Staining, Marker

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide abrogates the endosomal recruitment of Sec61 α . Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to ovalbumin (50 μ g/ml) or PBS pulse. The relocation of Sec61 α from endoplasmic reticulum to endosomes was determined by confocal microscope by Rab5, calnexin, and Sec61 α antibody staining. The colocalized plots of Sec61 α (b) with Rab5/calnexin were counted and analyzed. The expressions of Rab5/Sec61 α and calnexin/Sec61 α in the cells which are corresponding to the (a) colocalized cells were shown (c). Nuclei were counterstained with DAPI (blue). Original magnification, ×600. Data were presented as the mean ± SEM, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown. Rab5: early endosome marker; calnexin: endoplasmic reticulum marker.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Microscopy, Staining, Marker

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide abrogates the endosomal recruitment of Sec61 β . Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to ovalbumin (50 μ g/ml) or PBS pulse. The relocation of Sec61 β (a) from endoplasmic reticulum to endosomes was determined by confocal microscope by Rab5, calnexin, and Sec61 β antibody staining. The colocalized plots of Sec61 β (b) with Rab5/calnexin were counted and analyzed. The expressions of Rab5/Sec61 β and calnexin/Sec61 β in the cells which are corresponding to the (a) colocalized cells were shown (c). Nuclei were counterstained with DAPI (blue). Original magnification, ×600. Data were presented as the mean ± SEM, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown. Rab5: early endosome marker; calnexin: endoplasmic reticulum marker.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Microscopy, Staining, Marker

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide inhibits LPS-induced NF- κ B activation. Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to LPS (50 ng/ml) stimulation. (a–c) The effects of LPS, PYR-41, and thalidomide on NF- κ B activation were determined by Western blot by revealing I κ B α degradation and p65 phosphorylation. GAPDH was used as internal control. (d) The effects of LPS, PYR-41, and thalidomide on NF- κ B activation were determined by confocal microscope by revealing the nuclei relocation of phosphorylated p65. Nuclei were counterstained with DAPI (blue). Original magnification, ×600. Data were presented as the mean ± SEM, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test. One representative from 3 independent experiments was shown.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Activation Assay, Derivative Assay, Cell Culture, Western Blot, Phospho-proteomics, Control, Microscopy

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Treatment with PYR-41 or thalidomide inhibits LPS-induced myddosome formation. Murine bone marrow-derived DC (cultured for 4 d) conferred thalidomide (30 μ M), PYR-41 (5 μ M), or DMSO treatment prior to LPS (50 ng/ml) stimulation. The relocation of TLR4 (a) and MyD88 (b) was determined by confocal microscope by EEA1, TLR4, and MyD88 antibody staining. Nuclei were counterstained with DAPI (blue). Original magnification, ×600. The colocalized plots of TLR4 (a) or MyD88 (b) with EEA1 were counted and analyzed. (c–d) The effects of thalidomide and PYR-41 on TLR4 expression on DC were determined via flow cytometry. Numbers in histogram indicate MFI of analyzed population. The data were presented as the mean ± SEM, ∗ p < 0.05, ∗∗∗ p < 0.001, and one-way ANOVA with Newman–Keuls post test.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Derivative Assay, Cell Culture, Microscopy, Staining, Expressing, Flow Cytometry

Journal: Journal of Immunology Research

Article Title: PYR-41 and Thalidomide Impair Dendritic Cell Cross-Presentation by Inhibiting Myddosome Formation and Attenuating the Endosomal Recruitments of p97 and Sec61 via NF- κ B Inactivation

doi: 10.1155/2018/5070573

Figure Lengend Snippet: Mechanism of PYR-41- and thalidomide-impaired DC cross-presentation. PYR-41 and thalidomide inhibit myddosome formation via reducing NF- κ B activation. Decreased myddosome formation recruited less p97 and Sec61 from the ER toward the endosomes, leading to less internalized antigens exported to the cytosol to be degraded, resulting in a reduced cross-presentation and subsequent cross-priming.

Article Snippet: PYR-41 and thalidomide were bought from Selleck Chemicals (Houston, TX, USA).

Techniques: Activation Assay

Journal: Reproductive toxicology (Elmsford, N.Y.)

Article Title: Exposure-based assessment of chemical teratogenicity using morphogenetic aggregates of human embryonic stem cells

doi: 10.1016/j.reprotox.2019.10.004

Figure Lengend Snippet: Compounds tested in the present study

Article Snippet: Thalidomide , 50–35–1 , Santa Cruz (sc-201445) , 10 mM (DMSO) , Contraindicated (1.9–5.8 μM).

Techniques: Concentration Assay, Solvent, Control, Proliferation Assay

Journal: Reproductive toxicology (Elmsford, N.Y.)

Article Title: Exposure-based assessment of chemical teratogenicity using morphogenetic aggregates of human embryonic stem cells

doi: 10.1016/j.reprotox.2019.10.004

Figure Lengend Snippet: Effects of teratogenic exposures on morphology and gene expression in HESCA-CSR

Article Snippet: Thalidomide , 50–35–1 , Santa Cruz (sc-201445) , 10 mM (DMSO) , Contraindicated (1.9–5.8 μM).

Techniques: Gene Expression, Concentration Assay