Journal: PLoS ONE

Article Title: Electroacupuncture-Like Stimulation at the Baihui (GV20) and Dazhui (GV14) Acupoints Protects Rats against Subacute-Phase Cerebral Ischemia-Reperfusion Injuries by Reducing S100B-Mediated Neurotoxicity

doi: 10.1371/journal.pone.0091426

Figure Lengend Snippet: Representative western blot images show the expression of (A) mitochondrial cytochrome c and (C) cytosolic cytochrome c, TRADD, FADD, cleaved caspase-8 and cleaved caspase-3 in the ischemic cortical penumbra in the Sham-7 d, Model-7 d, EA-7 d, and Non-acup-7 d groups 7 d after reperfusion. COX IV and actin were used as internal controls for the mitochondrial and cytosolic fractions, respectively. The relative mitochondrial expression of (B) cytochrome c, and the relative cytosolic expression of (D) cytochrome c, (E) TRADD, (F) FADD, (G) cleaved caspase-8, and (H) cleaved caspase-3 were evaluated in the ischemic cortical penumbra in the Sham-7 d, Model-7 d, EA-7 d, and Non-acup-7 d groups (n = 4). Data are presented as mean ± SD. * P <0.05 compared with the Sham-7 d group; # P <0.05 compared with the Model-7 d group.

Article Snippet: They were then incubated with a mouse anti-GFAP (1∶1000 dilution, #3670 Cell Signaling Technology), rabbit anti-phospho-SAPK/JNK (p-JNK (Thr183/Tyr185); 1∶1000 dilution, #9251S Cell Signaling Technology), rabbit anti-phospho-p44/42 mitogen-activated protein kinase (MAPK (p-ERK); 1∶1000 dilution, #9101 Cell Signaling Technology), rabbit anti-phospho-p38 MAP kinase (p-p38 MAP kinase (Thr180/Tyr182); 1∶1000 dilution, #9212 Cell Signaling Technology), rabbit anti-cytochrome c (1∶1000 dilution, #4272 Cell Signaling Technology), rabbit anti-tumor necrosis factor receptor type 1-associated death domain (TRADD) (1∶1000 dilution, #3694 Cell Signaling Technology), rabbit anti-Fas-associated death domain (FADD) (1∶1000 dilution, #341282 Calbiochem), rabbit anti-cleaved caspase-8 (1∶1000 dilution, 3259-100 BioVision), or rabbit anti-cleaved caspase-3 (1∶1000 dilution, #9661S Cell Signaling Technology) antibody overnight at 4°C.

Techniques: Western Blot, Expressing

Journal: FEBS letters

Article Title: Agonist antibody activates death receptor 6 downstream signaling involving TRADD recruitment.

doi: 10.1016/j.febslet.2013.12.010

Figure Lengend Snippet: Fig. 5. TRADD is involved in DR6-induced IL-6 activation. (A and B) HEK293T cells were transiently transfected with IL-6 reporter plasmid, Renilla and increasing amounts of expression vectors for TRADD or TRADD-DN (A) or together with DR6-FL (B). 24 h later, promoter activity was measured by luciferase assay. (C) HEK293T cells were co- transfeced with IL-6-Luc, Renilla, DR6-FL and TRADD as described in (B), and then stimulated with DQM3. 16 h later, luciferase activity was measured. (D) HEK293T cells were co-transfected with TRADD siRNA (si-TRADD) or negative control siRNA (si-NC) and expression plasmid for TRADD. Western blot analysis was performed to verify the efficiency of siRNA. (E) TRADD siRNA-treated cells were transfected with IL-6 reporter plasmid and DR6-FL. 24 h after transfection, cells were subjected to DQM3 stimulation for 16 h, and then luciferase activity was monitored as described above. Results are shown as the mean ± S.E.M. ⁄P < 0.05; ⁄⁄P < 0.01.

Article Snippet: The rabbit polyclonal anti-TRADD antibody was purchased from Abcam.

Techniques: Activation Assay, Transfection, Plasmid Preparation, Expressing, Activity Assay, Luciferase, Negative Control, Western Blot

Journal: FEBS letters

Article Title: Agonist antibody activates death receptor 6 downstream signaling involving TRADD recruitment.

doi: 10.1016/j.febslet.2013.12.010

Figure Lengend Snippet: Fig. 6. DR6 interacts with TRADD in co-immunoprecipitation assays. (A and B) HEK293T cells in 60-mm dishes co-transfected with DR6-FL/EX and Flag-tagged TRADD or TRADD-DN were harvested 48 h after transfection. Total cell lysates were immunoprecipitated with anti-DR6 (DQM3) and immunoblotted with anti-TRADD (A) or anti-Flag (B). Western blot analysis of DR6, TRADD and Flag was performed on whole cell lysates as a control (Input). (C) HEK293T cells were co-transfected with expression vectors for DR6-FL and Flag-tagged TRADD, and then stimulated or not with DQM3 for the indicated times. The interaction of TRADD with DR6 was assessed after DQM3 stimulation by co-immunoprecipitation. The immunoprecipitates and cell extracts were analyzed by immunoblot. Results representative of at least three independent experiments are shown. b-Actin was used as loading control for each fraction. ⁄, Heavy chain (DQM3); IP, immunoprecipitates; IB, immunoblot.

Article Snippet: The rabbit polyclonal anti-TRADD antibody was purchased from Abcam.

Techniques: Immunoprecipitation, Transfection, Western Blot, Control, Expressing