Figure 6. Inhibition of the CD39/CD73-adenosine pathway ameliorates hypoxia-driven immunosuppression in HCC development.

(A) WT and miR-223KO mice were treated with DEN+CCl₄ as described in Figure 1. Levels of extracellular adenosine ectonucleotidases Cd39 and Cd73 mRNA expression, and adenosine levels in HCC tissues from WT and miR-223KO mice were determined. (B) The relative adenosine levels in cell medium were analyzed in Hepa1-6 cells after miR-223/NC mimics transfection under normoxia and hypoxia. (C) Adenosine and/or A2A adenosine receptor inhibitor CPI-444 were applied in primary macrophages or primary T cells, Cd274 and Pdcd1 mRNA expression were determined by RT-qPCR. (D, E) C57BL/6J mice were subjected to CCl₄+Hepa1-6-derived HCC model and treated with anti-CD73 (n=10) or isotype antibodies (n=8). Gross images of livers are shown, and tumor volume (mm³) was measured (panel D). The percentages of PD-1⁺CD8⁺ T cells and PD-L1⁺ macrophages were analyzed by flow cytometry (panel E). Values represent means ± SEM. *P< 0.05, **P< 0.01, ***P< 0.001.