Fig 2: ADAR1 promotes the production of pro-Inflammatory cytokines via ROS in astrocytes. Astrocytes were primary cultured from wild type (WT) and ADAR ±mice. A, Representative DHE fluorescent images showing the ROS levels. B, ROS levels in A were quantified relative to the WT control (Ctrl) group. C-D, Western blot analyses of IL-1, IL-6 and TNF-α production in primary astrocytes cultured with or without Oxygen-Glucose-Deprivation (OGD) and H2O2 (0.1 mM) for 24 h (C). The protein levels were quantified by normalizing to α-Tubulin (D). *P < 0.05 vs. WT astrocytes with OGD (WT, OGD); **P < 0.05 vs. ADAR1 ± astrocytes with OGD (ADAR1+/−, OGD); n = 6.

Fig 3: Exhausted CD4+ T cells revealed the increased expression of genes related to cell cycle, angiogenesis, and cytokines. During the process of exhaustion, restimulated CD4+ T cells exhibited altered expression of (A) cell cycle inhibitor p21, which was strongly elevated in CD4+ T cells cocultured with cancer cells, (B) upregulation of angiogenesis-related genes, and (C) overexpression of interleukins IL-6 and IL-1B only in CD4+ T cells co-cultured with cancer cells. (D) The highly elevated intracellular levels of IL-6 and IL-1B were found only in T cells co-cultured with cancer cells (T15M). (A) The data from 3 independent biological replicates (donors) are presented. (B,C) The data are from 1 representative biological replicate, the other biological replicates are presented in Figure S7. The statistical significance was calculated using Student’s t-test p < 0.05 and statistically significant values are indicated by asterisks. The uncropped blots and molecular weight markers are shown in Supplementary Materials.

Fig 4: Recipient MoDCs take up exos, altering inflammatory cytokine profiles. (A) Dil (red)-labeled MoDC exos (arrows) inside recipient MoDCs counterstained with DAPI nuclear stain (blue), and phalloidin (green) for the cell membrane and visualized using confocal microscopy (upper panel images captured with 20× lens and lower panel with 63× lens). (B) Colocalization of TGFB1 in MoDCs +/− regMoDCexo or iMoDCexo treatment using immunogold TEM. (C) Q-rt-PCR through TaqMan gene expression using delta-delta CT and plotted as relative fold change of inflammatory cytokines IL-6, IL-10, IL-1B, TNFa, and IL-23.

Fig 5: Neutralizing antibody cocktail against IL-1, 6 and TNF-α blocks neuron cell apoptosis caused by activated astrocytes. Primary neurons were treated with astrocyte-conditioned medium with or without (Ctrl) oxygen-glucose-deprivation (OGD). The medium was supplemented with PBS or neutralizing antibodies against IL-1β, −6 and/or TNF-α, as indicated. (A-B) TUNEL staining of neuron cells treated with astrocyte-conditioned medium (A), and TUNEL + cells were quantified relative to the total neuron cells (B). &P < 0.01 vs. group without OGD (Ctrl, PBS); @P < 0.01 vs. group with OGD & PBS (OGD, PBS); n = 6. (C-D) Cleaved-Caspase 3 and Cleaved-PARP levels were detected by Western blot (C) and quantified by normalizing to GAPDH (D). &P < 0.01 vs. group without OGD (Ctrl, PBS); @P < 0.01 vs. group with OGD & PBS (OGD, PBS) for the corresponding proteins, respectively; n = 6.