Fig. 6: Effect of CAF-derived CXCL12 on CXCR4/XRCC5 axis and stemness of HCC cells. | Nature Communications

Fig. 6: Effect of CAF-derived CXCL12 on CXCR4/XRCC5 axis and stemness of HCC cells.

From: Multi-omic analysis reveals a CAF-stemness-governed classification in HCC liver transplant recipients beyond the Milan criteria

Fig. 6

A CXCL12 concentration in CAF conditioned medium (CAF-CM) and LX-2 conditioned medium (LX-2-CM) was detected using ELISA. B, C XRCC5 expression was detected by western blotting in HCC cells treated with 0 nM, 10 nM, 100 nM, and 1 μM recombinant human CXCL12 protein for 48 h. Band intensities were semi-quantified using IMAGE LAB software and normalized with β-actin. Values were presented as the means under the bands. D Effect of siCXCR4 was detected by western blotting. E, F XRCC5 and CXCR4 expression was detected by western blotting in HCC cells treated with recombinant human CXCL12 protein (1 μM) and siCXCR4 or not. The effect of CXCL12 stimulation (1 μM) with CXCR4 knockdown or not, as well as CXCR4 inhibitor administration, AMD3100 (1 μM), or not on the stemness of HCC cells were shown by mRNA expression levels of stemness-related genes (G, H), cell migration (I), in vitro self-renewal capacity (J). K Schematic of the transwell co-culture model of HCC cells and CAFs. Created with BioRender.com. L, M The effect of co-culture with CXCR4 knockdown or not as well as AMD3100 (1 μM) administration or not on stemness of HCC cells were shown by mRNA expression levels of stemness-related genes. N IHC staining of CXCR4 and CXCL12 in patients with HCC undergoing liver transplantation. O Positive correlation between XRCC5 with CXCR4 and CXCL12 in 79 HCC samples. P Graphical summary of the mechanisms based on the combination of integrated multi-omics and experimental evidence. The data of ELISA (four individual experiments), cell functional assays and RT-qPCR analysis (three individual experiments) were presented as mean ± SD. Two-tailed Student’s t-test was used for comparisons, P values are shown on the graphs. ELISA, enzyme linked immunosorbent assay; HCC, hepatocellular carcinoma; IHC immunohistochemistry, TFs transcriptional factors, NC negative control, CAF cancer-associated fibroblast, ATAC-seq assay for transposase-accessible chromatin sequencing, CSC cancer stem cell. Source data are provided in the Source Data file.

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