Journal: Cell Death & Disease

Article Title: YIPF2 promotes chemotherapeutic agent-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in non-small cell lung cancer cells

doi: 10.1038/s41419-020-2436-x

Figure Lengend Snippet: a , b A549 ( a ) and H1792 ( b ) cells were treated with PEM at 5.0 μM for the indicated times (0, 6, 12, 24, 36 and 48 h). Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. c A549 cells were treated with doxorubicin (DOX) at various concentrations (0–2.0 μM) for 18 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. d Overexpression of YIPF2 in H1792 and H1299 cells in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. e Knockdown of YIPF2 expression by YIPF2–1 siRNA in A549 and H1792 cells in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. f Overexpression of YIPF2 in H1299 cells (left) or Knockdown of YIPF2 expression by YIPF2–1 siRNA in H1792 cells (right) in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10A and ACTB. g Three A549 cell lines (Ctrl, YIPF2, YIPF2 + siTNFRSF10B) in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B, CASP8, CASP3 and ACTB. h Three H1299 cell lines (Ctrl, siYIPF2–1, siYIPF2–1 + TNFRSF10B (short isoform)) in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B, CASP8, CASP3 and ACTB.

Article Snippet: The primary antibodies used in western blot assays and Immunoprecipitation were as follows: anti-YIPF2 (Cat. no. HPA019902; Sigma-Aldrich), ACTB (Cat. no. A1978; Sigma-Aldrich), CASP8 (Cat. no. 9746S; CST), CASP3 (Cat. no. NB100-56708; Novus Biologicals), PARP1 (Cat. no. 9542S; CST), TNFRSF10B (Cat. no. 2019; ProSci Incorporated), RAB8 (Cat. no. 6975S; CST), Flag (Cat. no. F7425, F1804; Sigma-Aldrich) and GFP (Cat. no. G1544, Sigma-Aldrich; Cat. no. sc-9996, Santa Cruz).

Techniques: Western Blot, Over Expression, Expressing

Journal: Cell Death & Disease

Article Title: YIPF2 promotes chemotherapeutic agent-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in non-small cell lung cancer cells

doi: 10.1038/s41419-020-2436-x

Figure Lengend Snippet: a Overexpression of YIPF2 in A549 and H1792 cells. The surface expression of TNFRSF10B was confirmed by flow cytometry analyses. b Knockdown of YIPF2 expression by YIPF2–1 and YIPF2–2 siRNA in A549 cells. The surface expression of TNFRSF10B was confirmed by flow cytometry analyses. c Relative RT-qPCR analyses of YIPF2 and TNFRSF10B mRNA levels after YIPF2 overexpression in H1792 (left) and H1299 (right) cells ( n = 3). d Relative RT-qPCR analyses of YIPF2 and TNFRSF10B mRNA levels after YIPF2 knocking down in H1792 (left) and H1299 (right) cells ( n = 3). e Left: Overexpression of YIPF2 in H1299 cells in the presence or absence of cycloheximide (CHX) at 10 μg/ml for the indicated times (0, 4, 8 and 12 h). Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. Right: The band intensity of TNFRSF10B was quantified by ImageJ software and plotted. This experiment was repeated three times independently with similar results. f Left: Knockdown of YIPF2 expression by YIPF2–1 siRNA in A549 cells in the presence or absence of cycloheximide (CHX) at 10 μg/ml for the indicated times (0, 4, 8 and 12 h). Cell lysates were analyzed by Western blotting with antibodies against YIPF2, TNFRSF10B and ACTB. Right: The band intensity of TNFRSF10B was quantified by ImageJ software and plotted. This experiment was repeated three times independently with similar results. (mean ± SEM, n = 3 independent experiments; NS, not significant; * P < 0.05, ** P < 0.01 and *** P < 0.001; P -values in c , e and f were obtained using two-tailed Student’s t -tests, P -values in d were obtained using one-way ANOVA followed by Bonferroni’s post-hoc test).

Article Snippet: The primary antibodies used in western blot assays and Immunoprecipitation were as follows: anti-YIPF2 (Cat. no. HPA019902; Sigma-Aldrich), ACTB (Cat. no. A1978; Sigma-Aldrich), CASP8 (Cat. no. 9746S; CST), CASP3 (Cat. no. NB100-56708; Novus Biologicals), PARP1 (Cat. no. 9542S; CST), TNFRSF10B (Cat. no. 2019; ProSci Incorporated), RAB8 (Cat. no. 6975S; CST), Flag (Cat. no. F7425, F1804; Sigma-Aldrich) and GFP (Cat. no. G1544, Sigma-Aldrich; Cat. no. sc-9996, Santa Cruz).

Techniques: Over Expression, Expressing, Flow Cytometry, Quantitative RT-PCR, Western Blot, Software, Two Tailed Test

Journal: Cell Death & Disease

Article Title: YIPF2 promotes chemotherapeutic agent-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in non-small cell lung cancer cells

doi: 10.1038/s41419-020-2436-x

Figure Lengend Snippet: a , b Knockdown of RAB8 expression by RAB8 siRNA in H1792 ( a ) and A549 ( b ) cells in the presence or absence of PEM at 5.0 μM for 36 h. The surface expression of TNFRSF10B was confirmed by flow cytometry analyses. c Overexpression of RAB8 in H1299 cells in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against RAB8, TNFRSF10B, CASP8, CASP3, PARP1 and ACTB. d Knockdown of RAB8 expression by RAB8 siRNA in H1792 cells in the presence or absence of PEM at 5.0 μM for 36 h. Cell lysates were analyzed by Western blotting with antibodies against RAB8, TNFRSF10B, CASP8, CASP3, PARP1 and ACTB.

Article Snippet: The primary antibodies used in western blot assays and Immunoprecipitation were as follows: anti-YIPF2 (Cat. no. HPA019902; Sigma-Aldrich), ACTB (Cat. no. A1978; Sigma-Aldrich), CASP8 (Cat. no. 9746S; CST), CASP3 (Cat. no. NB100-56708; Novus Biologicals), PARP1 (Cat. no. 9542S; CST), TNFRSF10B (Cat. no. 2019; ProSci Incorporated), RAB8 (Cat. no. 6975S; CST), Flag (Cat. no. F7425, F1804; Sigma-Aldrich) and GFP (Cat. no. G1544, Sigma-Aldrich; Cat. no. sc-9996, Santa Cruz).

Techniques: Expressing, Flow Cytometry, Over Expression, Western Blot

Journal: Cell Death & Disease

Article Title: YIPF2 promotes chemotherapeutic agent-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in non-small cell lung cancer cells

doi: 10.1038/s41419-020-2436-x

Figure Lengend Snippet: a H1299 cells were transfected with pcDNA3.1 or pcDNA3.1-Flag-TNFRSF10B (short isoform) plasmids. Then the co-IP assays were carried out with Flag antibody and the co-eluted proteins were detected by western blot assays with Flag and YIPF2 antibodies. b , c H1792 cells were transfected with pEGFP-N1 or pEGFP-N1-YIPF2 plasmids. Then the co-IP assays were carried out with GFP antibody and the co-eluted proteins were detected by western blot assays with GFP, TNFRSF10B ( b ) and RAB8 ( c ) antibodies. d H1792 cells were transfected with pEGFP-N1 or pEGFP-N1-RAB8 plasmids. Then the co-IP assays were carried out with GFP antibody and the co-eluted proteins were detected by western blot assays with GFP, and TNFRSF10B antibodies. e Cell lysates were prepared from H1792 cells transiently co-expressing pEGFP-N1-RAB8 and pcDNA3.1-YIPF2. The co-IP assays were carried out with GFP antibody and the co-eluted proteins were detected by western blot assays with YIPF2, GFP and TNFRSF10B antibodies. f Schematic illustration showing that YIPF2 promotes PEM-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in NSCLC cells.

Article Snippet: The primary antibodies used in western blot assays and Immunoprecipitation were as follows: anti-YIPF2 (Cat. no. HPA019902; Sigma-Aldrich), ACTB (Cat. no. A1978; Sigma-Aldrich), CASP8 (Cat. no. 9746S; CST), CASP3 (Cat. no. NB100-56708; Novus Biologicals), PARP1 (Cat. no. 9542S; CST), TNFRSF10B (Cat. no. 2019; ProSci Incorporated), RAB8 (Cat. no. 6975S; CST), Flag (Cat. no. F7425, F1804; Sigma-Aldrich) and GFP (Cat. no. G1544, Sigma-Aldrich; Cat. no. sc-9996, Santa Cruz).

Techniques: Transfection, Co-Immunoprecipitation Assay, Western Blot, Expressing

Journal: Cell Death & Disease

Article Title: YIPF2 promotes chemotherapeutic agent-mediated apoptosis via enhancing TNFRSF10B recycling to plasma membrane in non-small cell lung cancer cells

doi: 10.1038/s41419-020-2436-x

Figure Lengend Snippet: a Box plots of YIPF2 mRNA levels determined from two Oncomine datasets, namely TCGA Lung 2 and Weiss Lung (** P < 0.01 and *** P < 0.001; P- values were obtained using two-tailed Student’s t -tests). b Box plots of TNFRSF10B mRNA levels determined from two Oncomine datasets, namely TCGA Lung 2 and Bhattacharjee Lung. c Kaplan-Meier plots of the first-progression survival (FPS, upper) and post-progression survival (PPS, lower) of chemotherapy-treated patients stratified by YIPF2 expression. The data were acquired from the Kaplan-Meier plotter database ( P -values were obtained using the log-rank test). d Kaplan-Meier plots of the first-progression survival (FPS, upper) and post-progression survival (PPS, lower) of chemotherapy-treated patients stratified by TNFRSF10B expression. The data were acquired from the Kaplan-Meier plotter database ( P -values were obtained using the log-rank test). e Scatter plots showing the correlation of YIPF2 expression with TNFRSF10B expression in lung cancer cells in two GEO datasets (upper: GDS1688 which contains 29 lung cancer cell lines; lower: GDS3627 which contains 58 NSCLC cell lines). The r value was calculated via Spearman’s rank correlation coefficient analysis.

Article Snippet: The primary antibodies used in western blot assays and Immunoprecipitation were as follows: anti-YIPF2 (Cat. no. HPA019902; Sigma-Aldrich), ACTB (Cat. no. A1978; Sigma-Aldrich), CASP8 (Cat. no. 9746S; CST), CASP3 (Cat. no. NB100-56708; Novus Biologicals), PARP1 (Cat. no. 9542S; CST), TNFRSF10B (Cat. no. 2019; ProSci Incorporated), RAB8 (Cat. no. 6975S; CST), Flag (Cat. no. F7425, F1804; Sigma-Aldrich) and GFP (Cat. no. G1544, Sigma-Aldrich; Cat. no. sc-9996, Santa Cruz).

Techniques: Two Tailed Test, Expressing

Journal: Cell Reports Medicine

Article Title: Combining gemcitabine and MSC delivering soluble TRAIL to target pancreatic adenocarcinoma and its stroma

doi: 10.1016/j.xcrm.2024.101685

Figure Lengend Snippet:

Article Snippet: Rabbit anti-human TRAIL , Peprotech , 500-P135; RRID:AB_147781.

Techniques: Control, Virus, Variant Assay, Luciferase, Recombinant, Saline, Infection, Blocking Assay, Proliferation Assay, cDNA Synthesis, SYBR Green Assay, Enzyme-linked Immunosorbent Assay, Gene Expression, Software

Journal:

Article Title: Costimulating aberrant T cell responses by B7-H1 autoantibodies in rheumatoid arthritis

doi: 10.1172/JCI200316015

Figure Lengend Snippet: B7-H1 mAb promotes programmed cell death of activated CD4+ T cells. (a) Human CD4+ T cells were cultured with 10 μg/ml of immobilized control (ctl) Ab, B7-H1 mAb, PD-1Ig in the presence of precoated optimal dose (500 ng/ml) of CD3 mAb. The cells were analyzed by FACS to determine apoptotic cells (positive in AV and negative in PI staining). (b) Blocking of B7-H1 mAb-induced apoptosis by soluble B7-H1Ig. Control Ig or B7-H1Ig at 10 μg/ml was preincubated with immobilized control Ab or B7-H1 mAb for 30 minutes before the addition of T cells. Percentages of apoptotic CD4+ T cells were shown at 72 hours of culture. (c) Expression of apoptotic genes by B7-H1 mAb costimulation. Purified human CD4+ T cells (5 × 106) were cultured with immobilized B7-H1 mAb or control mAb at 10 μg/ml in the presence of optimal dose of CD3 mAb. The mRNA levels of each gene from B7-H1 mAb-stimulated T cells were presented as the fold induction, relevant to that from control mAb-treated T cells. (d) FACS analysis of expression of TRAIL protein on CD4+ T cells 3 days after anti-CD3/B7-H1 mAb costimulation. (e) Purified human CD4+ T cells were stimulated in the presence of indicated mAb or fusion protein in immobilized form as indicated in the legend of a and the activated form of caspase-3 at the indicated time point was stained by FAM-DEVD-FMK and subjected to FACS analysis. (f) Blocking of B7-H1 mAb-induced apoptosis of activated CD4+ T cells by anti–IL-10 neutralizing mAb. Purified human CD4+ T cells were stimulated with immobilized CD3 mAb and B7-H1 mAb for 72 hours, and the apoptotic cells were analyzed by AV and PI staining. Control Ab and mAbs to Fas ligand, IL-2, or IL-10 at 10 μg/ml was included at the beginning of the culture.

Article Snippet: The mAb specific for CD4 (RPA-T4), CD8 (RPA-T8), and CD45RO (UCHL1) were purchased from BD-PharMingen (San Diego, California, USA), and rabbit anti-human TNF-related apoptosis–inducing ligand (TRAIL) polyclonal Ab’s were purchased from Alexis Biochemical Corp. (San Diego, California, USA).

Techniques: Cell Culture, Staining, Blocking Assay, Expressing, Purification