Fig 2: Inhibition of PERK decreases DAMPs generation in B16F10 cell treated with P2Et.a SCR or PERK KO clones were treated with P2Et IC50, DOXO IC50, or vehicle for 12 h. Surface exposure of CRT was determined by flow cytometry among viable cells (Aqua negative) and primary antibody for CRT (rabbit anti-mouse) was detected using conjugate goat anti-rabbit secondary antibody Alexa Fluor 488. A representative histogram is shown. b Percentage folding change of MFI from treated cells relative to vehicle treatment expressed as mean ± SEM of three independent experiments is shown. c SCR or PERK KO clones were treated with P2Et IC50, DOXO IC50 or with vehicle for 48 h. Primary antibody for HMGB1 was detected using Alexa Fluor 488 conjugated goat anti-rabbit secondary antibody (green) and DAPI (blue) for the nuclei. Images were acquired with confocal microscope Olympus FV1000 with an oil 60× PlanAPO objective. Representative images of three independent experiments are shown. White bar corresponds to 10 µm d. Scatter plot of Pearson coefficients was assessed in 50 cells for each treatment in 3 independent experiments. Results from one representative experiment expressed as means ± S.E.M are shown. e SCR or PERK KO clones were treated with P2Et IC50 or 0.5 µg/ml MTX for 48 h. After treatment cells were stained with quinacrine (1 µM) and PI. Quinacrine low cells were determined among viable cell (PI-negative cells). A representative dot plot for each treatment is shown f Percentage of quinacrine low cells are expressed as mean ± SEM of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001, ****P < 0.0001

Fig 3: Differential expression of genes related to RNA processing in tumours expressing HMGB1 at high or low levels. (A) Selection of samples expressing mRNA HMGB1 at levels > median + 1.5 SD (HMGB1-high) and samples expressing mRNA HMGB1 at levels < median - 1.5 SD (HMGB1-low). (B) Volcano plots of genes expressing at higher levels in each group; log ratio in HMGB1-high/HMGB1-low. Ov, samples obtained from the PanCancer ovarian cystadenocarcinoma ATCG study in cBioportal; Pr, samples obtained from the PanCancer prostate adenocarcinoma ATCG study in cBioportal. (C) GO term enrichment analysis of genes expressed at higher levels in HMGB1-high samples in ovary or prostate cancer.

Fig 4: DA-MeHA hydrogel-encapsulated spheroids composed of ADSCs downregulated inflammatory responses. A DA-MeHA hydrogel-loaded spheroids composed of ADSCs downregulated the expression of TNF-a. B DA-MeHA hydrogel-loaded spheroids composed of ADSCs downregulated the expression of HMGB1. n = 4 samples/group (*p < 0.05 and **p < 0.01).

Fig 5: Interaction networks of HMGB1-associated proteins in SKOV-3 and PC-3 cell lysates showing principal clusters. Colour code is as follows: red, mRNA splicing, via spliceosome (GO:0000398); dark blue, rRNA processing (GO:0006364); green, ribonucleo-protein complex biogenesis (GO:0022613); yellow, nuclear transport (GO:0051169); pink, chromatin organisation (GO:0006325); purple cell division (GO:0051301); brown, DNA repair (GO:0006281); dark green, NuRD complex (GO:0016581).