Fig 1: CXCLs activate the ERK1/2 signaling pathway to up-regulate LIF expression in CAAs. (A and B) SB225002 or Reparixin was added to the co-culture system. (A) The total RNA of adipocytes was extracted for q-PCR analysis, or (B) the CAA culture medium was collected for ELISA analysis. (C) The protein expression level of adipocytes was detected by western blot. (D) Adipocytes were treated with rhCXCL3 or rhCXCL8 in combination with SB225002 and Reparixin. The LIF mRNA levels in adipocytes of each group were detected by q-PCR. (E) Adipocytes were treated with rhCXCL3 and rhCXCL8. The expression and localization of p-ERK1/2, NF-?B p65 and Stat3 in CAAs were observed using a fluorescent confocal microscope, scale bar: 20 µm. (F and G) a-CXCL3 (5 µg/mL) was added in the co-cultured system. The adipocyte RNA was extracted for q-PCR analysis (F), or the CAA culture medium was collected for ELISA analysis (G). (H) The protein expression level of adipocytes was analyzed by western blot. (I) The mRNA level of CXCR2 and LIF in adipocytes next to the normal breast tissue and breast cancer tissue were detected by q-PCR. (J) The mRNA level of CXCL1-3 and CXCL8 in normal breast tissue and breast cancer tissue was detected by q-PCR. Typical microscopic fields and blots are shown and quantitative data are presented as the mean ± SD from at least three independent experiments. #p < 0.05, ##p < 0.01, ###p < 0.001, Co-culture or rhCXCL3/8 VS. Control; *p < 0.05, **p < 0.01, ***p < 0.001, Co-culture + inhibitors/antibody or rhCXCL3/8 + inhibitors VS. Co-culture or rhCXCL3/8.