Fig 2: EZH2 promoted OGD-induced apoptosis and inflammation in cardiomyocytes by repressing Socs3.a Cardiomyocytes were assigned to five groups, i.e., no treatment (control), deprivation of oxygen and glucose (OGD), OGD treated with EZH2 inhibitor (DZNep), pre-transfected with scramble shRNA plus OGD and EZH2 inhibitor treatment (OGD + DZNep + shNC), and pre-transfected with SOCS3-specific shRNA plus OGD and EZH2 inhibitor treatment (OGD + DZNep + shSOCS3). The cell viability of each group was detected by MTT assay. b Apoptosis was detected by flow cytometry after annexin V/PI staining. c Statistical analysis of Fig. 6b. d The mRNA levels of IL-1β, IL-6 and TNF-α were assessed by qPCR. e The protein levels of p-p65, p65, H3K27me3, EZH2 and SOCS3 were determined by western blotting. The statistical analysis is presented. Error bars represent the mean ± SD. *P < 0.05 and **P < 0.01.

Fig 3: In vitro, the YAP inhibitor VP reverses the regulatory effect of Gap19 on the Cx43-YAP-SOCS axis. a, b At the beginning of hemin stimulation, astrocytes were treated with the YAP inhibitor VP. WB analysis showed that VP suppressed the increase in SOCS1/SOCS3 levels induced by Gap19 treatment. c, d Representative photos showing the levels of TLR4, p-IKKβ, IKKβ, IKBα, p65, p-p65, and β-tubulin after treatment with the YAP inhibitor VP. e, f Representative photographs showing the levels of JAK2, p-JAK2, STAT3, p-STAT3, and β-tubulin after treatment with the YAP inhibitor VP. g qRT-PCR analysis of the transcriptional activities of IL-1β, IL-6, TNF-α, and MCP1 in astrocytes after VP treatment. The bars represent the SEM of the data from 3 samples per group. *P < 0.05, **P < 0.01 compared to the hemin group. #P < 0.05, ##P < 0.01 compared with the Gap19 + hemin group

Fig 4: MDSC-secreted IL10 enhanced STAT3/SOCS3 signaling in vitro.A Western blot analyses were used to determine the expression patterns of IL10-stimulated p-STAT3 and SOCS3 protein levels in CCD841 and IEC-6 cells. B, C Western-blot analysis was used to determine the expression of p-STAT3, STAT3, SOCS3, IL10, and IL6 in co-cultured IEC-6 cells with 0.5 Gy/MDSCs or 5 Gy/MDSCs directly or indirectly using a transwell system. D The expression of p-STAT3, SOCS3, IL10, and IL6 in IEC-6 cells depleted of SOCS3 using siRNA and co-cultured with splenic 0.5 Gy/MDSCs after IR exposure were determined using western blot analysis. E The expression of p-STAT3, SOCS3, IL10, and IL6 in IEC-6 cells depleted of SOCS3 using siRNA and co-cultured with recombinant IL10 after IR exposure were determined using western blot analysis. F Representative immunofluorescence images of spleen cells stained for phosphorylated STAT3 (green), IL6 (red), and both p-STAT3 and IL6 (yellow). DAPI staining (blue) was used to determine the number of nuclei. Scale bar = 100 µm. IR irradiation.

Fig 5: Paeoniflorin can inhibit the LPS-induced expression of Tlr2 and p38 genes in Raw264.7 cells and inhibit the LPS-induced increase in ASK1, TLR2, and p38 expression and decrease in SOCS3 expression in Raw264.7 cells. (A, B: tlr2 and p38 mRNA expression; C: SCOS3, ASK1, TLR2, and p38 protein expression; D-G: relative expression levels).