Fig. 4: β-catenin signaling activation excludes DC infiltration by suppressing CXCL12 expression in ICC with LNM. | Cell Death Discovery

Fig. 4: β-catenin signaling activation excludes DC infiltration by suppressing CXCL12 expression in ICC with LNM.

From: Mobilization and activation of tumor-infiltrating dendritic cells inhibits lymph node metastasis in intrahepatic cholangiocarcinoma

Fig. 4

A Representative IHC staining images of β-catenin or non-phosphorylated β-catenin in ICC samples from TMA cohort. Red arrows indicate positive nuclear staining of β-catenin. B Association between β-catenin or non-phosphorylated β-catenin staining and LNM status in TMA cohort. Cell counts of CD45+ CD11c+ DCs (C) and CD8+ TILs (D) in ICC samples grouped by β-catenin or non-p-β-catenin staining intensity in TMA cohort. The mRNA (E) and protein expression (F) of CTNNB1 (β-catenin) assessed by qPCR and western blot in three human ICC cell lines 48 h following siRNA transfection. CXCL12, CCL4, and CCL5 mRNA expression (G) quantitated by qRT-PCR and amount of secreted CXCL12, CCL4, and CCL5 (H) in 48h-conditioned siRNA-treated human ICC cell supernatants detected by enzyme-linked immunosorbent assay (ELISA). I Putative β-Catenin/LEF1/TCF complex binding site (5’-TACAAAG-3’) in the promoter region (site 2) of CXCL12 was identified by in silico analysis. Specific primers for this promoter region (−800bp to −400bp) were designed. J ChIP-qPCR assay was performed in ICC cell lines with IgG and β-Catenin-specific antibodies. The results are presented as percentage of the total input chromatin DNA. K Schematic representation for the establishment of mouse ICC cell lines MuCCA1 and mIC-23 cells. MuCCA1 cells were generated from AKT/NICD-driven ICCs after 6 months of ex vivo culture. mIC-23 cells were generated from AKT/YAP-induced ICCs. Anti-CK19 antibody was used for IF staining of ICC cells. L Knockdown efficiency validated by qRT-PCR and western blot in two mouse ICC cell lines 48 h following siRNA transfection. M Secreted CXCL12, CCL4, and CCL5 in 48h-conditioned siRNA-treated mouse ICC cell supernatants detected by ELISA. N Trans-well migration assay of BMDCs attracted by condition medium and siRNA-treated MuCCA1 cell supernatants, with or without IT1t dihydrochloride (20 µM) for 24 h. *p < 0.05, **P < 0.01, ***p < 0.001, ****p < 0.0001.

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