Fig 2: EBNA1 inhibits p65 phosphorylation in carcinoma cell lines. Western blot analyses of total and phosphorylated (ser 536) p65 in (A) Ad/AH and (B) Hone1 cells stably expressing EBNA1 or a neomycin control vector (neo) under basal conditions or following stimulation with TNFa. Western blotting for EBNA1 and ß-actin serve as EBNA1 expression and protein loading controls, respectively.

Fig 3: TIF-IA degradation in response to multiple stress stimuli of NF-?B (A and C) SW480 or Hela cells were mock or UV-C (20 or 40 J/m2) irradiated. Following the times specified, immunoblots was performed on WCL with the indicated antibodies. (B) Immunomicrograph (×63) showing increased nucleolar area (as depicted by nucleolin staining) in SW480 cells in response to aspirin (3 mM, 16 h) or UV-C (40 J/m2, 2 h). (D) SW480 cells were treated with carrier, 10 µM Camptothecin (CPT) or aspirin (3 mM) for 16 h. Top: Western blot was performed with the indicated antibodies on WCL. Bottom: ?H2AX immunocytochemistry confirmed DNA damage in response to CPT. (E) SW480 cells were treated with DMSO (carrier), CX5461 (500 nM), BMH-21 (4 µM), ActinomycinD (ActD, 50 ng/ml) or aspirin (10 mM) for 4 h, or TNF (10 ng/ml) for 30 min, WCL were examined by western blot using the antibodies indicated. (F) SW480 or Hela cells were treated with carrier (DMSO), ceramide-2 (C2, 10 uM)) or ceramide-6 (C6, 10 uM) for 16 h. Top: WCL were analysed by western blot with the indicated antibodies. Bottom: Representative immunomicrographs (×63) show the localisation of fibrillarin and RelA in SW480 cells. (G) SW480 cells were pre-treated with 100 µM FumonisinB1 (FMB1), prior to aspirin (3 mM,16 h) exposure. Top: Immunoblots indicate cellular levels of TIF-IA. TIF-IA intensity (relative to actin) was determined for each condition using ImageJ analysis. Results are presented as the percentage relative TIF-IA compared to control. Mean (±s.e.m.) is shown for six experiments. Bottom: Representative immunomicrograph (×63) demonstrating the cellular localisation of RelA. Fibrillarin acts as a nucleolar marker. The percentage of cells showing nucleolar RelA was determined manually. At least 200 cells from at least 5 fields of view per were counted per experiment. The results are the mean (±s.e.m.). N = 3. P values were derived using a two tailed Student's t test. Scale bars = 10 µm. Actin or a tubulin act as loading controls.

Fig 4: GEN inhibits NF-?B activation, Müller cell activation, and inflammatory secretion through the Nrf2 antioxidant pathway. Nrf2 siRNA transfection was used to knockdown Nrf2 expression, and scramble siRNA (Scr siRNA) was used as the negative control. GEN was administered to Müller cells 24 h prior to stimulation with HG. (a–c) The protein expression of HO-1 and NQO1 was measured by western blotting. **P < 0.01 vs. the HG group and #P < 0.05 vs. the HG+GEN group. (d) ROS production was measured by an ROS assay kit. (e–h) The protein expression of I?Ba, p-I?Ba, P65, p-P65, and GFAP was measured by western blotting. **P < 0.01 vs. the HG group and #P < 0.05 vs. the HG+GEN group. (i–k) ELISA was used to measure the protein levels of TNF-a, IL-1ß, and IL-6 in Müller cells. **P < 0.01 vs. the HG group and ##P < 0.01 vs. the HG+GEN group. n = 4/group.

Fig 5: Molecular docking analysis. (A) The binding affinity of triptolide for CDKN1A (lowest Vina score = -6.9, cavity size = 18). (B) The binding affinity of triptolide for c-JUN (lowest Vina score = -5.7, cavity size = 24). (C) The binding affinity of triptolide for RELA (lowest Vina score = -7.5, cavity size = 304). (D) The binding affinity of triptolide for TP53 (lowest Vina score = -8.9, cavity size = 1320).