Journal: Reproductive Immunology: Open Access

Article Title: NF-kB Regulation in T-cells in Pregnancy is Mediated via Fas/FasL Interactions: The Signal for which is Derived from Exosomes Present in Maternal Plasma

doi: 10.21767/2476-1974.100008

Figure Lengend Snippet: Figure 2: Factors present in maternal plasma can induce apoptosis in Jurkat T-cells. A) Jurkat T-cells were cultured in the presence of Fas activating antibody CH11 (0.625 µg/mL), or recombinant TRAIL (3 ng/mL) and the level of early (DilC1(5)-PI-) and late (DilC1(5)-PI+) apoptosis determined by flow cytometry. Flow charts are representative of 3 separate experiments with similar results. B) Jurkat T-cells were cultured in the presence of 20% (v/v) non-pregnant (NP) or pregnant (P) plasma from individual patients for 72 hrs and the apoptosis determined. Pooled data from 5 individual patients (NP and P15-19) is indicated in the bar graph. C) Jurkat T-cells were pre-incubated for 1 hr in the presence of ZB4 (500 ng/mL) or anti-TRAIL (0.8 ng/mL) and subsequently cultured for 72 hrs in the presence of 20% (v/v) NP or P plasma. Apoptosis was determined by Flow cytometry. The level of early (DilC1(5)-PI-) apoptosis is shown on the graph where n=4 for both NP and P samples. *p<0.05 relative to NP plasma.

Article Snippet: To assess the role of FasL and TRAIL activation in regulating p65 expression and apoptosis cells were incubated in the presence of the Fas activating antibody CH11 (Millipore) or recombinant human TRAIL (ProSci) at concentrations stated in the figure legends.

Techniques: Clinical Proteomics, Cell Culture, Recombinant, Flow Cytometry, Incubation

Journal: Reproductive Immunology: Open Access

Article Title: NF-kB Regulation in T-cells in Pregnancy is Mediated via Fas/FasL Interactions: The Signal for which is Derived from Exosomes Present in Maternal Plasma

doi: 10.21767/2476-1974.100008

Figure Lengend Snippet: Figure 3: Factors present in maternal plasma can induce suppression of p65 in T-cells; A) Jurkat T-cells were cultured in the presence of increasing concentrations of Fas activating antibody CH11 or recombinant TRAIL and the level of p65 determined. Blot is representative of 3 separate experiments with similar results; B) Jurkat T-cells were cultured in the presence of CH11 (0.625 µg/mL) ±500 ng/mL Fas inactivating antibody ZB4 or mIgG control or IgM control (0.625 µg/mL) or recombinant TRAIL (3 ng/mL) ±80 ng/mL anti-TRAIL or mIgG control or BSA (3 ng/mL) and the level of p65 determined. Blot is representative of 3 separate experiments with similar results; C) Jurkat T-cells were cultured in the presence of CH11 (0.625 µg/mL) ±500 ng/mL Fas inactivating antibody ZB4 or mIgG control or IgM control (0.625 µg/mL) and the level of p65 and CD3ζ determined in cell lysates. Blot is representative of 3seperate experiments each showing similar results.

Article Snippet: To assess the role of FasL and TRAIL activation in regulating p65 expression and apoptosis cells were incubated in the presence of the Fas activating antibody CH11 (Millipore) or recombinant human TRAIL (ProSci) at concentrations stated in the figure legends.

Techniques: Clinical Proteomics, Cell Culture, Recombinant, Control

Journal: Journal of Molecular Medicine (Berlin, Germany)

Article Title: Silencing of Mcl-1 overcomes resistance of melanoma cells against TRAIL-armed oncolytic adenovirus by enhancement of apoptosis

doi: 10.1007/s00109-021-02081-3

Figure Lengend Snippet: Resistance of certain melanoma cell lines to AdV-TRAIL treatment. A Schematic overview of the regulation elements of AdV-TRAIL. E1A lacking the retinoblastoma-binding domain (E1A ΔpRb ) is controlled by a tyrosinase promoter which ensures a selective expression (a) and subsequent viral replication (b) in melanoma cells. E1A ΔpRb also slightly transactivates the bi-directional doxycycline (Dox)-inducible Tight promoter (c) leading to minimal expression of the reverse tetracycline transactivator (rtTA) (d). In the presence of Dox, rtTA is able to bind to the Tight promoter (e) and induces strong rtTA expression (f), resulting in an activation loop. In parallel, activation of the Tight promoter in the presence of Dox leads to TRAIL expression (g), which subsequently induces apoptosis (h). In cells without tyrosinase promoter activity, E1A ΔpRb and TRAIL are not expressed. Adenoviral inverted terminal repeats are labeled as 5′-ITR and 3′-ITR. B Cell viability of four melanoma cell lines after treatment with TRAIL and AdV-TRAIL. Cells were either treated with soluble TRAIL (50 ng/ml) or transduced with AdV-TRAIL (5 MOI). In the latter case, TRAIL expression was induced by Dox as indicated. Cell viability was detected by calcein-AM staining 48 h after transduction with AdV-TRAIL or treatment with soluble TRAIL, respectively. The mean percentages of calcein-AM-positive cells (viable) ± SEM of three independent experiments are shown. Statistical significance: * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001

Article Snippet: Soluble TRAIL (Adipogen, San Diego, CA, USA) was added to the cell culture in a concentration of 50 ng/ml.

Techniques: Binding Assay, Expressing, Activation Assay, Activity Assay, Labeling, Transduction, Staining

Journal: Cell Death & Disease

Article Title: Constitutive localization of DR4 in lipid rafts is mandatory for TRAIL-induced apoptosis in B-cell hematologic malignancies

doi: 10.1038/cddis.2013.389

Figure Lengend Snippet: ( a ) Surface expression of TRAIL receptors in transformed and non-transformed cells. Flow cytometry analysis of surface expression level of DR4 and DR5 in four Burkitt lymphoma cell lines. Histograms obtained in a representative experiment are shown. Numbers represent the median fluorescence intensity. Results obtained in a representative experiment are shown. ( b ) Graphs showing the surface expression level of DR4 and DR5 in four Burkitt lymphoma cell lines (left panel) and in non-transformed cells PBL, fibroblasts and HUVEC (right panel). Mean±S.D. of the median fluorescence intensity obtained in four different experiments is reported. ( c ) Soluble TRAIL-induced apoptosis in transformed and non-transformed cells. Biparametric flow cytometry analysis of sTRAIL-induced apoptosis in four Burkitt lymphoma cell lines after double staining with annexin V-FITC/Trypan blue. Numbers represent the percentage of dead cells either annexin V/Trypan blue double-positive or annexin V single-positive. Results obtained in a representative experiment are shown. ( d ) Bar graphs showing the amount of apoptosis in four Burkitt lymphoma cell lines (left panel) and in non-transformed PBL, fibroblasts and HUVEC (right panel) after sTRAIL administration. Mean±S.D. of the percentages of annexin V-positive cells obtained in four different experiments is reported. Significant differences ( P <0.01) were detected between control and treated cancer cells

Article Snippet: Apoptosis was induced by incubating cells with recombinant soluble TRAIL (TRAIL 100 ng/ml for 48 h, Adipogen International, Inc. San Diego, CA, USA) or with agonist antibodies anti-DR4 and anti-DR5 (2 μ g/ml, both R&D Systems, Minneapolis, MN, USA).

Techniques: Expressing, Transformation Assay, Flow Cytometry, Fluorescence, Double Staining, Control

Journal: Cell Death & Disease

Article Title: Constitutive localization of DR4 in lipid rafts is mandatory for TRAIL-induced apoptosis in B-cell hematologic malignancies

doi: 10.1038/cddis.2013.389

Figure Lengend Snippet: Lipid rafts and TRAIL-induced apoptosis. Left panels. Bar graphs showing the amount of sTRAIL-induced apoptosis in cells pretreated with MBC or perifosine. Mean±S.D. of the percentages of annexin V-positive cells obtained by flow cytometry in four different experiments is reported. Right panels. IVM analysis after triple cell staining with anti-DR4/anti-GM3/Hoechst in control, MBC- or perifosine-treated cells. The yellow fluorescence areas indicate the co-localization. ( a ) Ramos lymphoma cell line, ( b ) Namalwa lymphoma cell line and ( c ) freshly isolated human lymphocytes. * indicates P <0.01 ° indicates P <0.05 versus sTRAIL samples. ( d ) Quantitative evaluation of GM3/DR4 and GM3/DR5 association by FRET technique, as revealed by flow cytometry analysis. Numbers represent the FRET efficiency (calculated by using Riemann algorithm). Note different scales

Article Snippet: Apoptosis was induced by incubating cells with recombinant soluble TRAIL (TRAIL 100 ng/ml for 48 h, Adipogen International, Inc. San Diego, CA, USA) or with agonist antibodies anti-DR4 and anti-DR5 (2 μ g/ml, both R&D Systems, Minneapolis, MN, USA).

Techniques: Flow Cytometry, Staining, Control, Fluorescence, Isolation

Journal: Cell Death & Disease

Article Title: Constitutive localization of DR4 in lipid rafts is mandatory for TRAIL-induced apoptosis in B-cell hematologic malignancies

doi: 10.1038/cddis.2013.389

Figure Lengend Snippet: ( a ) Ex vivo analyses of lymphocytes isolated from patients with CLL. Flow cytometry analysis of surface expression level of CD19 in lymphocytes freshly isolated from a representative HD among six or from two pathological subjects among six (Pt1 and Pt2) affected by B leukemia. Numbers represent the percentage of CD19-positive cells. Results obtained in a representative experiment are shown. ( b ) Surface expression of TRAIL receptors. Flow cytometry analysis of surface expression level of DR4 and DR5 in lymphocytes freshly isolated from a healthy donor or from two pathological subjects affected by B leukemia. Numbers represent the median fluorescence intensity. Results obtained in a representative experiment are shown. ( c ) Apoptosis induction by sTRAIL, mTRAIL and agonist antibodies to DR4 and DR5. Biparametric flow cytometry analysis of apoptosis induced by sTRAIL or by mTRAIL, that is, coculturing CD34 + -armed cells or CD34 + -mock with target cells (1 : 1 ratio). Numbers represent the percentage of apoptotic cells either annexin V/Trypan blue double-positive or annexin V single-positive. Results obtained in a representative experiment are shown. Analyses shown in b and c were restricted to CD19-positive cells

Article Snippet: Apoptosis was induced by incubating cells with recombinant soluble TRAIL (TRAIL 100 ng/ml for 48 h, Adipogen International, Inc. San Diego, CA, USA) or with agonist antibodies anti-DR4 and anti-DR5 (2 μ g/ml, both R&D Systems, Minneapolis, MN, USA).

Techniques: Ex Vivo, Isolation, Flow Cytometry, Expressing, Fluorescence

Journal: International Journal of Ophthalmology

Article Title: Combination of oridonin and TRAIL induces apoptosis in uveal melanoma cells by upregulating DR5

doi: 10.18240/ijo.2021.12.05

Figure Lengend Snippet: A: Blank control group; B: 5 µmol/L paclitaxel positive control group; C: 3.12 µmol/L oridonin; D: 6.25 µmol/L oridonin; E: 12.5 µmol/L oridonin; F: 31.2 ng/mL TRAIL; G: 62.5 ng/mL TRAIL; H: 125 ng/mL TRAIL; I: 3.12 µmol/L oridonin+31.2 ng/mL TRAIL; J: 6.25 µmol/L oridonin+62.5 ng/mL TRAIL; K: 12.5 µmol/L oridonin+125 ng/mL TRAIL.

Article Snippet: Reagents Soluble TRAIL protein (Novoprotein, Shanghai); oridonin (Chengdu Institute of Biology, Chinese Academy of Sciences); MTT (AMRESCO); RPMI1640 (Thermo Fisher); foetal bovine serum (FBS, GIBCO); DMSO (Life science); paclitaxel (Shanghai Yuanye Bio-Technology); penicillin (Life Technology); Streptomycin (GIBCO); DR5 antibody (Abcam); a-caspase-3 antibody (Abcam); x-linked inhibitor of apoptosis protein (XIAP) antibody (Abcam); β-actin antibody (Abcam).

Techniques: Control, Positive Control

Journal: International Journal of Ophthalmology

Article Title: Combination of oridonin and TRAIL induces apoptosis in uveal melanoma cells by upregulating DR5

doi: 10.18240/ijo.2021.12.05

Figure Lengend Snippet: A-G: Flow cytometry analysis of cell apoptosis: A: Blank control group; B: 3.12 µmol/L oridonin; C: 31.2 ng/mL TRAIL; D: 3.12 µmol/L oridonin+31.2 ng/mL TRAIL; E: 12.5 µmol/L oridonin; F: 125 ng/mL TRAIL; G: 12.5 µmol/L oridonin+ 125 ng/mL TRAIL; H: Histogram present the proportion of apoptotic cells in each group, aP<0.05, bP<0.01, compared with control group.

Article Snippet: Reagents Soluble TRAIL protein (Novoprotein, Shanghai); oridonin (Chengdu Institute of Biology, Chinese Academy of Sciences); MTT (AMRESCO); RPMI1640 (Thermo Fisher); foetal bovine serum (FBS, GIBCO); DMSO (Life science); paclitaxel (Shanghai Yuanye Bio-Technology); penicillin (Life Technology); Streptomycin (GIBCO); DR5 antibody (Abcam); a-caspase-3 antibody (Abcam); x-linked inhibitor of apoptosis protein (XIAP) antibody (Abcam); β-actin antibody (Abcam).

Techniques: Flow Cytometry, Control

Journal: International Journal of Ophthalmology

Article Title: Combination of oridonin and TRAIL induces apoptosis in uveal melanoma cells by upregulating DR5

doi: 10.18240/ijo.2021.12.05

Figure Lengend Snippet: A: The relative expression level of DR5 gene after transfection with DR5 siRNA was detected by qPCR; B: The effect of TRAIL and oridonin on the activity of transfected si-DR5 cells was detected by MTT; C: Western blot was used to detect the expression levels of apoptosis-related proteins in each group; D: A histogram obtained by quantizing the gray value of the protein band in C. aStatistically significant difference compared with the control group, P<0.05.

Article Snippet: Reagents Soluble TRAIL protein (Novoprotein, Shanghai); oridonin (Chengdu Institute of Biology, Chinese Academy of Sciences); MTT (AMRESCO); RPMI1640 (Thermo Fisher); foetal bovine serum (FBS, GIBCO); DMSO (Life science); paclitaxel (Shanghai Yuanye Bio-Technology); penicillin (Life Technology); Streptomycin (GIBCO); DR5 antibody (Abcam); a-caspase-3 antibody (Abcam); x-linked inhibitor of apoptosis protein (XIAP) antibody (Abcam); β-actin antibody (Abcam).

Techniques: Expressing, Transfection, Activity Assay, Western Blot, Control

Journal: British Journal of Cancer

Article Title: In vitro sensitivity testing of minimally passaged and uncultured gliomas with TRAIL and/or chemotherapy drugs

doi: 10.1038/sj.bjc.6604459

Figure Lengend Snippet: Chemotherapy drugs and doses used in this study

Article Snippet: We used soluble TRAIL (Peprotech, Rocky Hill, CT, USA) at concentrations commonly employed in vitro (1 μ g ml −1 and 100 ng ml −1 ).

Techniques: Clinical Proteomics, Concentration Assay

Journal: British Journal of Cancer

Article Title: In vitro sensitivity testing of minimally passaged and uncultured gliomas with TRAIL and/or chemotherapy drugs

doi: 10.1038/sj.bjc.6604459

Figure Lengend Snippet: Propidium iodide uptake assay of glioma cell sensitivity to combination treatments. Cells from the indicated early passage glioma cell lines or ex vivo gliomas were incubated in vitro with soluble TRAIL at 100 ng ml −1 (+) or 1000 ng ml −1 (++) alone or together with temozolomide (13.7 μ g ml −1 ) or cisplatin (54 μ g ml −1 ) for 48 h. Flow cytometry measurement of propidium iodide exclusion was used to quantitate the proportion of surviving cells. The areas of the circles denote survival following each treatment. Small circles indicate efficient killing, large circles reflect survival.

Article Snippet: We used soluble TRAIL (Peprotech, Rocky Hill, CT, USA) at concentrations commonly employed in vitro (1 μ g ml −1 and 100 ng ml −1 ).

Techniques: Ex Vivo, Incubation, In Vitro, Flow Cytometry