Regional expression of CXCL12/CXCR4 in liver and hepatocellular carcinoma and cell-cycle variation during in vitro differentiation

The CXCL12/CXCR4 system may be important in carcinoma. Expression of the alpha-chemokine SDF-1alpha (stromal cell derived factor-1alpha)/CXCL12 mRNA is reduced in many carcinomas, yet its tissue protein expression may guide metastasis. Here! we first compare the mRNA and protein expression of CXCL12 and its receptor CXCR4 in human liver, hepatocellular carcinoma, and malignant cell lines, and then assess cell cycle variation in CXCR4 expression. CXCR4 mRNA was present in most normal human tissues and malignant cell lines; it was only marginally reduced in hepatomas, while CXCL12 was markedly reduced, P<0.0001. Immuno-histochemical staining of adjacent non-malignant liver showed regional CXCR4 cytoplasmic and cell-surface staining, limited to those hepatocytes around the central vein, a distribution resembling that of CXCL12. CXCL12 protein was not present in hepatocellular carcinoma cells in vivo, nor was cytoplasmic CXCR4 staining; nuclear CXCR4 protein expression in some malignant hepatocytes and CXCR4 staining of capillary endothelial cells around tumor cells were noted. In some malignant cell lines that had no CXCL12 on northern blots CXCL12 was weakly detectable by RT-PCR or protein staining in the cytoplasm of a few cells. With a view to future manipulation of CXCL12/CXCR4 expression and growth we noted that in HT-29 cells CXCR4 protein expression was less on confluent than on non-confluent cells and varied during the cell cycle. Higher expression was associated most closely with the percentage of cells in the S-phase and inversely with the percentage of cells in the G1-phase. Treatment of HT-29 cells with butyrate reduced CXCR4 cell surface expression and reduced the percentage of cells in S-phase. In summary, CXCL12 protein expression parallels its mRNA, being markedly reduced in malignant cell lines and hepatomas; in liver, the regional distributions of CXCL12 and cytoplasmic CXCR4 are similar; finally, in HT-29, CXCR4 expression correlates with the S-phase of the cell cycle and is reduced during butyrate-induced differentiation.