Phospho-NCF1/p47-phox (Ser359) Antibody - #AF3167

Fig. 1: ASC-interacting protein NCF4 is associated with colorectal cancer development. a Immunoblot analysis of ASC from the immunoprecipitated products generated with immunoprecipitation with an ASC antibody from the lysates of WT and Asc–/– BMDMs infected with F. novicida (100 MOI) for 12 h. b Mass spectrometry analysis of the IP product in (a). The peptides of ASC, NCF4, NCF1, and NCF2 were detected in the IP product with ASC antibody from WT and Asc–/– BMDMs in two repeated experiments. c Immunoblot analysis of Myc-ASC co-IP with FLAG-NCF4, FLAG-NCF4-PX, FLAG-NCF4-SH3 and FLAG-NCF4-PB1 from lysates of HEK293T cells transfected with the indicated plasmids. d Immunoblot analysis of FLAG-NCF4 co-IP with V5-ASC, V5-ASC-PYD, and V5-ASC-CARD from lysates of HEK293T cells transfected with the indicated plasmids. e Co-IP analysis of endogenous ASC interacting with total and phosphorylated NCF4, NCF1, and NCF2 in WT and Asc–/– BMDMs treated with the NLRP3 activator LPS plus ATP (LPS, 500 ng/mL for 4 h and ATP, 5 mM for 15 min). f Gene expression analysis of NCF1, NCF2, and NCF4 in total (left) and paired (right, n = 41) colorectal tumors (n = 480) and control tissues (n = 41) from 480 colorectal cancer (CRC) patients of pooled colon and rectal adenocarcinoma datasets in the TCGA database. g Correlation analysis between the gene expression of NCF4 and survival rate of CRC patients. (High, n = 466; Low, n = 131) Data are from 2 (a, b) or representative of 3 independent experiments with similar results (c–e). Wilcoxon signed rank test for (f), Log-rank (Mantel–Cox) test for (g), p-value is indicated in the graph. Source data are provided as a Source Data file.

Figure S6 Oxidized rGO increased lipid peroxidation and NOX2 activation in PC12 cells. Oxidized rGO (c-rGO) was collected after pristine rGO (p-rGO) was cultured with PC12 cells for 3 h. (A-B) PC12 cells were pretreated with NAC for 30 min before being treated with two kinds of rGO (20 µg/mL) for 1 h and stained with BODIPY 581/591 C11 reagent. The lipid peroxidation degrees were quantified via FITC fluorescence intensity, with n = 10 fields per experimental condition from 3 independent tests. Scale bar: 100 μm. (C-D) After PC12 cells were treated as described in (A), plasma membrane proteins were isolated to detect the expression levels of NOX2, p-p47phox, p-p67phox and Rac2. The protein levels were quantified after normalization to ATPase. The results represent the mean ± SD of three independent experiments. *p<0.05, **p<0.01, ***p<0.001 compared with the control group and with the p-rGO group.

Figure 5 TZG significantly regulated the expression of NETs formation-related indicators, and the corresponding proteins in PI3K-RAC2 signaling (A–E). Serum levels of dsDNA, IL-8, TNF-α, CitH3, and MPO detected by ELISA analyses, respectively. Samples in (A)∼(E) were obtained from the blood supernatant of rats in different groups. (F–I) The ratios of p-PI3K/PI3K and p-P47/P47, and the expression levels of RAC2 and PAD4 proteins detected by western blot analyses, respectively. Samples in (F)∼(I) were obtained from MTrPs tissues of rats in different groups. The number of rats in each group was 8. Data are expressed as the mean ± S.D. “∗,” “∗∗,” and “∗∗∗,” p < 0.05, p < 0.01, and p < 0.001, respectively, comparison with the normal control group. “#,” “##,” and “###,” p < 0.05, p < 0.01, and p < 0.001, respectively, comparison with the MPS model group.