Fig 1: Screening IL8 as the therapeutic target after chemotherapy resistance.A The volcano plot of RNA transcription sequencing of the first-line chemotherapy group and the sequential chemotherapy group. A total of 256 differentially expressed genes (DEGs) with fold-change greater than 2. B KEGG pathways enriched by DEGs in the transcriptome affected by sequential chemotherapy compared with first-line chemotherapy. C qRT-PCR assays were performed to detect the changes of the DEGs in the three enriched signaling pathways most associated with cancer in tumor tissues treated with sequential and first-line chemotherapy. D, E IL8 expression at different periods of chemotherapy was detected by qRT-PCR assays and western blot analysis. Data are mean ± standard deviation. *P < 0.05, **P < 0.01. F Upregulation of IL8 in GC samples was obtained in the GEPIA database. G Kaplan–Meier OS curves in GC patients according to the expression of IL8. H Kaplan–Meier OS curves in GC patients after 5-Fu therapy according to the expression of IL8. I Western blot analysis of IL8 expression in BGC823 cells transfected with siRNAs against IL8. J CCK8 assays was carried out to detect the viability of BGC823 cells transfected with si-IL8#2 and si-IL8#3. **P < 0.01, ***P < 0.001. K, L Representative images (K) and quantification (L) of BGC823 cells transfected with si-IL8#2 and si-IL8#3 by EdU staining assays. Data are mean ± standard deviation. **P < 0.01. M–O Representative images (M) and quantification (N, O) of BGC823 cells transfected with si-IL8#2 and si-IL8#3 by colony-forming experiments and transwell assays. Data are mean ± standard deviation. **P < 0.01, ***P < 0.001.
Fig 2: ROS inhibition suppresses the IL8 and MCP1 transcripts expression in monocytic cells co-stimulated with TNF-a and H2O2. THP-1 cells were stimulated, in triplicate, with TNF-a (10 ng/mL) and/or H2O2 (10 mM) while control cells were treated with vehicle only. In ROS inhibition assays, cells were preincubated for 30 min with either NAC (1 mM) or curcumin (10 µM) before stimulation with TNF-a and/or H2O2. Total RNA was extracted and IL8/MCP1 gene expression was assessed by qRT-PCR as described in methods. Two-way ANOVA was used to calculate group differences and p-values less than 0.05 were considered as significant. The representative data (mean ± SEM) obtained from three independent experiments with similar results show effective suppression of (A) IL8 mRNA by NAC (p ? 0.0001) and (B) MCP1 mRNA by both NAC and curcumin (p = 0.01).
Fig 3: Upregulated adipose tissue chemokine CXC motif ligand-8 (CXCL8) expression in obesity. Adipose CXCL8 gene expression was determined in five lean, 18 overweight, and 27 obese non-diabetic individuals using real-time RT-PCR, while CXCL8 protein expression was determined in six lean, 14 overweight, and 10 obese non-diabetic individuals using immunohistochemistry (IHC) as described in Materials and Methods. The data (mean ± SEM) show that (A) adipose CXCL8 gene expression was upregulated in obese (P = 0.05) compared with lean individuals. (B) CXCL8 protein expression (IHC intensity in arbitrary units) was also higher in obese (P = 0.0001) compared with lean individuals. (C) The representative IHC images obtained from three independent determinations with similar results show upregulated adipose CXCL8 expression (purple arrows, 100× magnification) in overweight and obese as compared with lean individuals. Scale bar = 50 µM. (D) IRF5 and CXCL8 gene expression was associated with each other in the adipose tissue (r = 0.40, P = 0.0004). The asterisks * and *** represent significance levels of P = 0.05 and P < 0.001, respectively.
Fig 4: IL8 and MCP1 gene expression in human monocytic THP-1 cells stimulated with TNF-a and/or oxidative stress. THP-1 cells were stimulated, in triplicate, with TNF-a (10 ng/mL) and/or H2O2 (10 mM) for 24 h while control cells were treated with vehicle only. Likewise, THP-1 cells were also stimulated, in triplicate, with TNF-a under oxidative stress by 1% hypoxia. Total RNA was collected from cells for measuring gene expression of IL8 and MCP1 using qRT-PCR as detailed in methods. One-way ANOVA (Tukey’s multiple comparisons test) was used to calculate group differences and p-values less than 0.05 were considered as significant. The representative data (mean ± SEM) obtained from three independent experiments with similar results show the upregulated transcripts of (A) IL8 and (B) MCP1 in cells co-stimulated with TNF-a and H2O2, compared to TNF-a stimulation alone (p = 0.0001). Similarly, the representative data (mean ± SEM) from three independent experiments with similar results show increased transcripts expression of (C) IL8 and (D) MCP1 in the cells stimulated with TNF-a under 1% hypoxia, compared to TNF-a stimulation under normoxia (20% O2) (p < 0.0001).
Supplier Page from Abcam for Anti-IL-8 antibody