Journal: Discover Oncology

Article Title: CXCL10/CXCR3 axis facilitates the M2 polarization of macrophages in colorectal cancer via activating RAF

doi: 10.1007/s12672-025-04081-y

Figure Lengend Snippet: CXCL10 released by CRC cells facilitated the M2 polarization of macrophages. (A). The silence of CXCL10 was identified by Western blot. (B). The overexpression of CXCL10 was identified by Western blot. (C). Transfected HCT116 cells were co-cultured with macrophages in the Transwell. Percentages of M2 macrophages (CD68 + CD206 + cells) were checked by flow cytometry. (D). Percentages of M1 macrophages (CD68 + CD86 + cells) were checked by flow cytometry. (E). Productions of IL-6, IL-10, and TNF-α in macrophages were determined by ELISA. (F). Expressions of iNOS and Arg1 in macrophages were determined by Western blot (* p < 0.05 vs. si-NC, ** p < 0.01 vs. si-NC, ## p < 0.01 vs. OE-NC) (All experiments were performed in triplicate ( n = 3 biological replicates)

Article Snippet: The PVDF membrane was blocked with 5% skimmed milk while shaking at room temperature for 1 h and then incubated with the primary antibody against CXCL10 (1:1000, DF6417, affinity, USA), inducible nitric oxide synthase (iNOS) (1:1000, ab3523, Abcam, USA), arginase 1 (Arg1) (1:1000, DF6657, affinity, USA), RAF1 (1:1000, ab181115, Abcam, USA), ERK1/2 (1:1000, 4695s, CST, USA), p-ERK1/2 (1:1000, 4370T, CST, USA), and GAPDH (1:10000, 10494-1-AP, Proteintech, USA) overnight at 4°C.

Techniques: Western Blot, Over Expression, Transfection, Cell Culture, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Discover Oncology

Article Title: CXCL10/CXCR3 axis facilitates the M2 polarization of macrophages in colorectal cancer via activating RAF

doi: 10.1007/s12672-025-04081-y

Figure Lengend Snippet: Influences of CXCL10 released by CRC cells on the M2 polarization of macrophages were abolished by the inhibitor of CXCR3 or RAF. CXCL10-overexpressed HCT116 cells were cultured with an inhibitor of CXCR3 (NBI-74330, 3 nM) or RAF (MCP110, 20 µM) for 24 h. Then, HCT116 cells were co-cultured with macrophages in the Transwell. (A). Percentages of M2 macrophages (CD68 + CD206 + cells) were checked by flow cytometry. (B). Percentages of M1 macrophages (CD68 + CD86 + cells) were checked by flow cytometry. (C). Productions of IL-6, IL-10, and TNF-α in macrophages were determined by ELISA. (D). Expressions of iNOS and Arg1 in macrophages were determined by Western blot (** p < 0.01 vs. OE-NC, # p < 0.05 vs. CXCL10 OE, ## p < 0.01 vs. CXCL10 OE) (All experiments were performed in triplicate ( n = 3 biological replicates)

Article Snippet: The PVDF membrane was blocked with 5% skimmed milk while shaking at room temperature for 1 h and then incubated with the primary antibody against CXCL10 (1:1000, DF6417, affinity, USA), inducible nitric oxide synthase (iNOS) (1:1000, ab3523, Abcam, USA), arginase 1 (Arg1) (1:1000, DF6657, affinity, USA), RAF1 (1:1000, ab181115, Abcam, USA), ERK1/2 (1:1000, 4695s, CST, USA), p-ERK1/2 (1:1000, 4370T, CST, USA), and GAPDH (1:10000, 10494-1-AP, Proteintech, USA) overnight at 4°C.

Techniques: Cell Culture, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Western Blot