human il 8 sandwich elisa kit  (Proteintech)

Journal: The FASEB Journal

Article Title: CXCL8 induces M2 macrophage polarization and inhibits CD8⁺ T cell infiltration to generate an immunosuppressive microenvironment in colorectal cancer

doi: 10.1096/fj.202201982rrr

Figure Lengend Snippet: FIGURE 1 CXCL8 is the key regulator in CRC malignant progression. (A and B) Authentication of 37 common upregulated DEGs (A) and 94 common downregulated DEGs (B) in the four datasets through Venn diagrams software. (C) The PPI network contains 85 nodes and 152 edges. The nodes meant DEGs, the edges meant the interaction of proteins. The yellow area represents the module analysis via MCODE of Cytoscape software that contains 8 nodes and 25 edges. (D) The mRNA expression of CXCL8 between tumors and normal tissues was analyzed in TCGA. (E and F) IHC staining of CXCL8 in 16 paired CRC tissue and adjacent tissue. Representative IHC images of CXCL8 were shown in E and quantitative results were in F. Scale bar: 200 μm (4×), 50 μm (20×), 20 μm (40×). (G) Western blot analysis of CXCL8 expression in normal colon cell line and CRC cell lines. β-Actin was used as a loading control. (H) qRT-PCR was used to determine the mRNA level of CXCL8 in normal and CRC cell lines. Data represent the average of three independent experiments (Student's t-test, *p < .05, **p < .01, ***p < .001).

Article Snippet: For the quantification of CXCL8 levels in the cell culture supernatant, the supernatant from CRC cell lines with or without IL- 1β treatment was collected, and the CXCL8 levels were measured by human CXCL8 sandwich ELISA kits (VAL103, NOVUS, USA) according to the manufacturer's instructions.

Techniques: Software, Expressing, Immunohistochemistry, Western Blot, Control, Quantitative RT-PCR

Journal: The FASEB Journal

Article Title: CXCL8 induces M2 macrophage polarization and inhibits CD8⁺ T cell infiltration to generate an immunosuppressive microenvironment in colorectal cancer

doi: 10.1096/fj.202201982rrr

Figure Lengend Snippet: FIGURE 2 CXCL8 may act as a regulator by increasing the infiltration of M2 macrophage in CRC immune microenvironment. (A and B) A total of 1229 DEGs were enriched by using the DAVID database for KEGG pathway (A) and GO functional enrichment analysis (B). The y-axis showed the terms and x-axis showed the -log10 (Q value) of each term. (C) Violin diagram showed the relative abundance of 14 TILs of CXCL8 in TCGA-COAD dataset (left) and TCGA-READ dataset (right). Red and blue colors represented the CXCL8 high- and low-expression samples, respectively. (D) Representative IHC images of CXCL8, CD68, and CD163 were analyzed in CXCL8 low-expression group (upper) and CXCL8 high-expression group (lower) from clinical paraffin tissue chip. Scale bar: 200 μm (4×), 50 μm (20×). (E and F) The coefficient of correlation between CXCL8, CD163 (E), and CD68 (F) were shown, respectively. (G) Representative IHC images of CXCL8, CD68, and CD163 in CXCL8/BALB/c and control/BALB/c group. Scale bar: 200 μm (4×), 50 μm (20×), 10 μm (40×). (H) Evaluation of IHC staining in G was shown in H. Data represent the average of three independent experiments (Student's t-test, ***p < .001).

Article Snippet: For the quantification of CXCL8 levels in the cell culture supernatant, the supernatant from CRC cell lines with or without IL- 1β treatment was collected, and the CXCL8 levels were measured by human CXCL8 sandwich ELISA kits (VAL103, NOVUS, USA) according to the manufacturer's instructions.

Techniques: Functional Assay, Expressing, Control, Immunohistochemistry

Journal: The FASEB Journal

Article Title: CXCL8 induces M2 macrophage polarization and inhibits CD8⁺ T cell infiltration to generate an immunosuppressive microenvironment in colorectal cancer

doi: 10.1096/fj.202201982rrr

Figure Lengend Snippet: FIGURE 4 IL-1β facilitates CXCL8 secretion and enhances the chemotaxis of M2 macrophage. (A) Heatmap of top 10 upregulated cytokines genes in CXCL8 high-expression group. (B) Correlations analysis between the expression of top 10 genes and CXCL8. The number represents the correlation coefficient, and the cross represents no statistical difference. (C-H) Western blot analysis of CXCL8 expression in SW480 (C) or HCT116 (F), qRT-PCR analysis of CXCL8 mRNA in SW480 (D) and HCT116 (G), and ELISA analysis of CXCL8 concentrations of SW480 (E) or HCT116 (H) after addition with recombinant human IL-1β. (I and J) Migration assay was used for detection of M2 macrophage chemotaxis ability in SW480 (I) and HCT116 (J). IL-1β was added at 50 ng/mL, and Reparixin (Rep) was added at 500 nmol/L to the lower-champer supernatant. Scale bar: 200 μm. Data represent the average of three independent experiments (Student's t-test, *p < .05, **p < .01).

Article Snippet: For the quantification of CXCL8 levels in the cell culture supernatant, the supernatant from CRC cell lines with or without IL- 1β treatment was collected, and the CXCL8 levels were measured by human CXCL8 sandwich ELISA kits (VAL103, NOVUS, USA) according to the manufacturer's instructions.

Techniques: Chemotaxis Assay, Expressing, Western Blot, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Recombinant, Migration

Journal: The FASEB Journal

Article Title: CXCL8 induces M2 macrophage polarization and inhibits CD8⁺ T cell infiltration to generate an immunosuppressive microenvironment in colorectal cancer

doi: 10.1096/fj.202201982rrr

Figure Lengend Snippet: FIGURE 5 CXCL8 promotes M2 polarization of macrophages by activating STAT3. (A) Representative image, CD163 and Ym-1 expression of THP-1-M0 macrophage after CXCL8 treatment for 24 h. Scale bar: 200 μm. (B) GSEA analysis showed JAK–STAT signaling pathways were enriched in the CXCL8 high-expression group. (C) Correlations analysis between CXCL8 expression and the expression of related genes in JAK–STAT signaling pathway. (D and E) Correlations assessed by Spearman's correlation between CXCL8 and JAK2 (D) or STAT3 (E) at mRNA level (TCGA database). (F and G) After CXCL8 treatment, western blot was used to detect STAT3 and p-STAT3 expression in M2 macrophage (control panel). M2 macrophages were pretreated with Stattic (STAT3 inhibitor) followed by CXCL8 addition (Stattic panel). Expression of CD163 was tested by qRT-PCR (G). Data represent the average of three independent experiments (Student's t-test, *p < .05, **p < .01, ***p < .001).

Article Snippet: For the quantification of CXCL8 levels in the cell culture supernatant, the supernatant from CRC cell lines with or without IL- 1β treatment was collected, and the CXCL8 levels were measured by human CXCL8 sandwich ELISA kits (VAL103, NOVUS, USA) according to the manufacturer's instructions.

Techniques: Expressing, Protein-Protein interactions, Western Blot, Control, Quantitative RT-PCR

Journal: The FASEB Journal

Article Title: CXCL8 induces M2 macrophage polarization and inhibits CD8⁺ T cell infiltration to generate an immunosuppressive microenvironment in colorectal cancer

doi: 10.1096/fj.202201982rrr

Figure Lengend Snippet: FIGURE 6 CXCL8 inhibits PD-1+CD8+ T cell infiltration. (A) Representative IHC images of CXCL8, CD3, and CD8 were analyzed in CXCL8 low-expression group (upper) and CXCL8 high-expression group (lower) from clinical paraffin tissue chip. (B) The correlation coefficient of CXCL8 and CD8 from A. (C) Representative IHC images of CXCL8, CD8, and PD-1 in CXCL8/BALB/c and control/BALB/c groups. (D and E) The quantification of the immunohistochemical staining of C, the percentage of CD8+ cells shown in D, and the CD8+PD-1+ cells were analyzed in E. (F) Representative IHC images of CXCL8, CD68, CD163, and PD-L1 in CXCL8/BALB/c and Control/ BALB/c groups. (G and H) The quantification of the immunohistochemical staining of F, and the percentages of CD163+ cells (G) and CD163+PD-L1+ cells (H) were analyzed. (I) WB was used to detect p-STAT3, STAT3, and PD-L1 expression in THP-1-M0 treated with CXCL8. (J) mIHC was performed to analyze colocalization of CD8 (brownish red), PD-1 (green) in murine CRC slice, and the nucleus was stained with DAPI (blue). The percentages of CD8+ cells or CD8+PD-1+ cells were analyzed with inForm Advanced Image Analysis software. Data represent the average of three independent experiments (Student's t-test, *p < .05, ***p < .001).

Article Snippet: For the quantification of CXCL8 levels in the cell culture supernatant, the supernatant from CRC cell lines with or without IL- 1β treatment was collected, and the CXCL8 levels were measured by human CXCL8 sandwich ELISA kits (VAL103, NOVUS, USA) according to the manufacturer's instructions.

Techniques: Expressing, Control, Immunohistochemical staining, Staining, Software

Journal: Nutrients

Article Title: Ellagitannins from Castanea sativa Mill. Leaf Extracts Impair H. pylori Viability and Infection-Induced Inflammation in Human Gastric Epithelial Cells

doi: 10.3390/nu15061504

Figure Lengend Snippet: Effect of Castanea sativa Mill. leaf extracts on IL-8 release. GES-1 cells were treated for 6 h with TNFα (10 ng/mL) or H. pylori (ratio 50:1, bacteria/cell), in addition to leaf extracts before ( A , B ) and after in vitro simulated digestion ( C , D ). IL-8 was measured by ELISA assay. Results are expressed as the mean ± SEM ( n = 3) of the relative percentage in comparison to stimulus (black bar), to which the value of 100% was arbitrarily assigned. * p < 0.05, ** p < 0.01, and *** p < 0.001 vs. stimulus. VR, Castanea sativa Mill. var. verdesa leaf extract; VN, Castanea sativa Mill. var. venegon leaf extract; dig., in vitro simulated digestion.

Article Snippet: Cells were seeded at the density of 3 × 10 4 cells/well (24-well plate) for 48 h. The chemokine IL-8 was quantified in cell medium in at least three independent experiments, after 6 h treatments with TNFα or H. pylori and the extracts, using a sandwich enzyme-linked immunosorbent assay (human IL-8 ABTS ELISA Development Kit, Peprotech).

Techniques: Bacteria, In Vitro, Enzyme-linked Immunosorbent Assay, Comparison

Journal: Nutrients

Article Title: Ellagitannins from Castanea sativa Mill. Leaf Extracts Impair H. pylori Viability and Infection-Induced Inflammation in Human Gastric Epithelial Cells

doi: 10.3390/nu15061504

Figure Lengend Snippet: Effect of ellagitannins from Castanea sativa Mill. on IL-8 release. GES-1 cells were treated for 6 h with TNFα (10 ng/mL) ( A ) or H. pylori (ratio 50:1, bacteria:cell) ( B ), in the presence of castalagin or vescalagin. IL-8 was measured by ELISA assay. Results are expressed as the mean ± SEM ( n = 3) of the relative percentage in comparison to stimulus (black bar), to which the value of 100% was arbitrarily assigned. ** p < 0.01, and *** p < 0.001 vs. stimulus.

Article Snippet: Cells were seeded at the density of 3 × 10 4 cells/well (24-well plate) for 48 h. The chemokine IL-8 was quantified in cell medium in at least three independent experiments, after 6 h treatments with TNFα or H. pylori and the extracts, using a sandwich enzyme-linked immunosorbent assay (human IL-8 ABTS ELISA Development Kit, Peprotech).

Techniques: Bacteria, Enzyme-linked Immunosorbent Assay, Comparison