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. 2021 Apr 6;9:631931. doi: 10.3389/fcell.2021.631931

FIGURE 4.

FIGURE 4

CAFs activated SDF-1/CXCR4/PI3K/AKT pathway and consequently inhibited HCC cell apoptosis. (A) Both RT-PCR and western immunoblotting assays showed that LX2 SULF2 cells expressed significantly more SDF-1 than LX2 Vector cells. Cell fractions: LX2 cells. (B) It was found by ELISA assay that there was more SDF-1 protein in conditioned medium from LX2 SULF2 cells than that from LX2 Vector cells (the protein was extracted from total cells). (C) Microarray profiling of mRNA expression showed that most downstream genes of SDF-1/CXCR4 signaling were remarkably upregulated in Hep3B CAFs cells in contrast to Hep3B Vector cells. (D) Co-culture with CAFs was found by western immunoblotting to increase expression of SDF-1, CXCR4, p-PI3K, and p-AKT in Hep3B cells, while treatment of CXCR4 inhibitor (AMD3100) did not alter the expression of phosphorylation of PI3K, AKT, BAD, caspase 9, and FKHRL 1 in Hep3B cells. Cell fractions: Hep 3B cells (the protein was extracted from total cells). (E) By both DAPI staining and caspase 3/7 activity assay, it was found that co-culture with CAFs suppressed cell apoptosis, whereas AMD3100 treatment did not influence Hep3B cell apoptosis apparently. CAF, carcinoma-associated fibroblast; HCC, hepatocellular carcinoma; SULF2, sulfatase 2.