Journal: Materials Today Bio

Article Title: Neutrophil membrane-mimicking nanodecoys with intrinsic anti-inflammatory properties alleviate sepsis-induced acute liver injury and lethality in a mouse endotoxemia model

doi: 10.1016/j.mtbio.2022.100244

Figure Lengend Snippet: Construction and characterization of neutrophil membrane-mimicking nanodecoys (NM) and red cell membrane-mimicking nanovesicles (RM). (A) and (B) The Cryo-electron micrograph of NM and RM (scale bar, 100 ​nm). (C) Protein profiles of the neutrophil membrane and NMs assessed by sodium dodecyl sulfate poly-acrylamide gel electrophoresis (SDS-PAGE). (D) Western blotting analysis of lymphocyte antigen 6 complex locus G6D (Ly6G) and tumor necrosis factor receptor −1 (TNFR1) in RM and NM. (E) and (F) Average diameter size and zeta potential of NM and RM. (G) The biodistribution of NM in major organs (n ​= ​3, statistical differences between groups were performed by a one-way ANOVA) and fluorescence intensity of liver at 24 ​h was set as 100%. ∗ p ​< ​0.05 compared with the fluorescence intensity of liver at 10 ​min. (H) Blood circulation time of NM (n ​= ​3). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The following antibodies were used: anti-TNFR1 (Sino Biological, Beijing, China), Ly6G (Thermo Fisher Scientific, NY, USA), COX-2 (Proteintech, IL, USA), and β-actin antibody (Sigma-Aldrich, MO, USA).

Techniques: SDS Page, Western Blot, Fluorescence

Journal: Nature Communications

Article Title: The cytoplasmic nuclear receptor RARγ controls RIP1 initiated cell death when cIAP activity is inhibited

doi: 10.1038/s41467-017-00496-6

Figure Lengend Snippet: RARγ has a role in the formation of death complexes. a Western blot analysis of HT-29 cont-shRNA, and RARγ-shRNA-A treated with DMSO, TS or TSZ for 24 h and cell lysates were immunoblotted with the indicated antibodies. b , c Immunoprecipitation of HT-29 cont-shRNA or RARγ-shRNA-A cells treated with TSZ for the indicated time. Cell lysates were collected and immunoprecipitated with anti-Caspase-8 antibody b or anti-TNFR1 antibody c . The immunoprecipitated complexes were immunoblotted with the indicated antibodies. d Sequential immunoprecipitation of HT-29 cont-shRNA or RARγ-shRNA-A cells treated TSZ for 2 h. First IP : TNFR1 complex I was immunoprecipitated using anti-TNFR1 antibody. Second IP : the remaining lysates were immunoprecipitated again with anti-TNFR1 antibody. Third IP : the remaining lysates were then immunoprecipitated with anti-TRADD antibody. The immunoprecipitated complexes were analyzed by with the indicated antibodies. (M: marker). All blots above are representative of one of three experiments

Article Snippet: Anti-RARγ (C-15) (sc-550) for human, anti-RARα (C-20) (sc-551), anti-caspase-8 (C-20) (sc-6136), anti-cIAP2 (sc7944) and anti-Fas (C-20) (sc-715) from Santa Cruz; anti-RIP1 (610459) and anti-FADD (610400) from BD Biosciences; anti-RARγ1 (ab5905) for mouse, anti-RIP3 (ab72106), anti-MLKL (ab184718) for human, anti-MLKL (ab172868) for mouse and anti-cIAP1 (ab2399) from Abcam; anti-RIP3 (2283) for mouse from ProSci, anti-TRADD (05-473) from Upstate; anti-TRAF2 (MAB3277) and anti-TNFR1 (AF-425-PB) from R&D; anti-Actin (A3853) (dilution 1:10,000), anti-FLAG (F9291) (dilution 1:5,000) and anti-GFP (G6539) (dilution 1:5 000) from Sigma; anti-V5 (R960-25) (dilution 1:5,000) from Invitrogen; anti-PARP1 (BML-SA250-0050) from Enzo Life Science; anti-GAPDH (NB300-22) (dilution 1:5,000) from Novus Biologicals; anti-cleaved caspase-8 (9496), anti-CYLD (4495), anti-RIP1 (137451) and anti-p-RIP1(65746) from Cell Signaling Technology; anti-DsRed (632392) (dilution 1:5,000) from Clontech.

Techniques: Western Blot, shRNA, Immunoprecipitation, Marker

Journal: Nature Communications

Article Title: The cytoplasmic nuclear receptor RARγ controls RIP1 initiated cell death when cIAP activity is inhibited

doi: 10.1038/s41467-017-00496-6

Figure Lengend Snippet: RARγ initiates the formation of death complexes by dissociating RIP1 from TNFR1. a Immunoprecipitation of HT-29 cont-shRNA, TRADD-shRNA or RIP1-shRNA treated with TSZ for the indicated times. Cell lysates were immunoprecipitated using anti-TNFR1 antibody and immunoblotted with the indicated antibodies. b Sequential immunoprecipitation of HEK293T cells co-transfected with FLAG-TNFR1, DsRed-RIP1 and increasing amounts of V5-RARγ-NLSmut plasmids as indicated. First IP : cell lysates were immunoprecipitated with anti-FLAG (TNFR1) antibody. Second IP : the remaining lysates were then immunoprecipitated with anti-V5 (RARγ) antibody. The immunoprecipitated complexes were analyzed by with the indicated antibodies. c Sequential immunoprecipitation of HEK293T cells co-transfected with V5-RARγ-NLSmut, RIP1-Myc, and increasing amounts of DsRed-RIP3 plasmids as indicated. First IP : cell lysates were immunoprecipitated with anti-V5 (RARγ) antibody. Second IP : the remaining lysates were then immunoprecipitated with anti-DsRed (RIP3) antibody. The immunoprecipitated complexes were analyzed with the indicated antibodies. d WT and RIP3−/− MEFs treated with TSZ for the indicated times. Cell lysates were immunoprecipitated using anti-Caspase-8 antibody and immunoblotted with the indicated antibodies. All blots above are representative of one of three experiments

Article Snippet: Anti-RARγ (C-15) (sc-550) for human, anti-RARα (C-20) (sc-551), anti-caspase-8 (C-20) (sc-6136), anti-cIAP2 (sc7944) and anti-Fas (C-20) (sc-715) from Santa Cruz; anti-RIP1 (610459) and anti-FADD (610400) from BD Biosciences; anti-RARγ1 (ab5905) for mouse, anti-RIP3 (ab72106), anti-MLKL (ab184718) for human, anti-MLKL (ab172868) for mouse and anti-cIAP1 (ab2399) from Abcam; anti-RIP3 (2283) for mouse from ProSci, anti-TRADD (05-473) from Upstate; anti-TRAF2 (MAB3277) and anti-TNFR1 (AF-425-PB) from R&D; anti-Actin (A3853) (dilution 1:10,000), anti-FLAG (F9291) (dilution 1:5,000) and anti-GFP (G6539) (dilution 1:5 000) from Sigma; anti-V5 (R960-25) (dilution 1:5,000) from Invitrogen; anti-PARP1 (BML-SA250-0050) from Enzo Life Science; anti-GAPDH (NB300-22) (dilution 1:5,000) from Novus Biologicals; anti-cleaved caspase-8 (9496), anti-CYLD (4495), anti-RIP1 (137451) and anti-p-RIP1(65746) from Cell Signaling Technology; anti-DsRed (632392) (dilution 1:5,000) from Clontech.

Techniques: Immunoprecipitation, shRNA, Transfection

Journal: Nature Communications

Article Title: The cytoplasmic nuclear receptor RARγ controls RIP1 initiated cell death when cIAP activity is inhibited

doi: 10.1038/s41467-017-00496-6

Figure Lengend Snippet: RARγ modulates TNF-induced RIP-initiated cell death. a TRADD is responsible for initiated TC or TCZ-induced cell death. b RARγ is essential for the transition from inflammatory signaling to death pathways in TNF-induced RIP1-initiated cell death. When RARγ is released from the nucleus in response to TNF under the conditions of cIAP inhibition, it initiates the formation of death complex IIa by dissociating RIP1 from TNFR1. When RIP1 recruits RIP3 to necrosome/complex IIb, RARγ is replaced from the complex

Article Snippet: Anti-RARγ (C-15) (sc-550) for human, anti-RARα (C-20) (sc-551), anti-caspase-8 (C-20) (sc-6136), anti-cIAP2 (sc7944) and anti-Fas (C-20) (sc-715) from Santa Cruz; anti-RIP1 (610459) and anti-FADD (610400) from BD Biosciences; anti-RARγ1 (ab5905) for mouse, anti-RIP3 (ab72106), anti-MLKL (ab184718) for human, anti-MLKL (ab172868) for mouse and anti-cIAP1 (ab2399) from Abcam; anti-RIP3 (2283) for mouse from ProSci, anti-TRADD (05-473) from Upstate; anti-TRAF2 (MAB3277) and anti-TNFR1 (AF-425-PB) from R&D; anti-Actin (A3853) (dilution 1:10,000), anti-FLAG (F9291) (dilution 1:5,000) and anti-GFP (G6539) (dilution 1:5 000) from Sigma; anti-V5 (R960-25) (dilution 1:5,000) from Invitrogen; anti-PARP1 (BML-SA250-0050) from Enzo Life Science; anti-GAPDH (NB300-22) (dilution 1:5,000) from Novus Biologicals; anti-cleaved caspase-8 (9496), anti-CYLD (4495), anti-RIP1 (137451) and anti-p-RIP1(65746) from Cell Signaling Technology; anti-DsRed (632392) (dilution 1:5,000) from Clontech.

Techniques: Inhibition

Journal: Frontiers in Neurology

Article Title: Cerebral Immunohistochemical Characterization of the H 2 S and the Oxytocin Systems in a Porcine Model of Acute Subdural Hematoma

doi: 10.3389/fneur.2020.00649

Figure Lengend Snippet: Protein BLAST search for primary antibodies (anti-human) to sus scrofa.

Article Snippet: anti-TNFR1 (Protein Tech, 21574-1-AP, RRID:AB_10734433 ) , Rabbit Polyclonal , 100 , 69.30 , TNFR1 fusion protein Ag16112 , 1:100.

Techniques: Sequencing, Concentration Assay

Journal: Frontiers in Neurology

Article Title: Cerebral Immunohistochemical Characterization of the H 2 S and the Oxytocin Systems in a Porcine Model of Acute Subdural Hematoma

doi: 10.3389/fneur.2020.00649

Figure Lengend Snippet: TNFR1 ( A , n = 1) and TNFR2 ( B , n = 1) in the brainstem (A/B 1) and in the cerebellum (A/B 2). In the brainstem TNFR1 (A1) and TNFR2 (B1) were expressed in neurons and microvasculature, and TNFR2 was also expressed in the parenchyma (B) . In the cerebellum purkinje cells expressed TNFR1 (A2) and TNFR2 was expressed in purkinje, granular cells and parenchyma (B2) . TNFR1, anti-tumor necrosis factor receptor 1; TNFR2, tumor necrosis factor receptor 2.

Article Snippet: anti-TNFR1 (Protein Tech, 21574-1-AP, RRID:AB_10734433 ) , Rabbit Polyclonal , 100 , 69.30 , TNFR1 fusion protein Ag16112 , 1:100.

Techniques:

Journal: Neoplasia (New York, N.Y.)

Article Title: The STAT3 Target Gene TNFRSF1A Modulates the NF-κB Pathway in Breast Cancer Cells.

doi: 10.1016/j.neo.2018.03.004

Figure Lengend Snippet: Figure 4. TNFRSF1A expression modulates NF-κB activity. MDA-MB-468 and BT549 cells were transfected with TNFR1 plasmid (encoding TNFRSF1A) or an empty vector. They were then stimulated with TNFα and analyzed by (A) immunoblot for the expression of TNFR1 (TNFRSF1A) in whole cell lysates (with tubulin serving as a loading control), (B) luciferase reporter assay for NF-κB-dependent transcriptional activity (n = 3), and (C) qRT-PCR for expression of endogenous NF-κB target genes (normalized to GAPDH; n = 3).

Article Snippet: Cells (5 × 105 cells per well in a 6-well plate) were seeded, and the following day were transfected using Lipofectamine 2000 (Invitrogen) with 1 μg of TNFR1 (pBMNZ-neo-Flag-TNFR1 L380A (from Martin Kluger), Addgene plasmid # 43949), or an empty vector as a control.

Techniques: Expressing, Activity Assay, Transfection, Plasmid Preparation, Western Blot, Control, Luciferase, Reporter Assay, Quantitative RT-PCR