Fig. 3.

Negative regulation of Gal-1 by miR-22 in HCC. a. qPCR analysis revealed upregulation of miR-22 and reduced level of Gal-1 in cells transfected with miR-22 mimic. b. Reduction of Gal-1 was also detected by western blotting. c. Reduction of miR-22 expression by miR-22 inhibitor and the subsequent Gal-1 increased expression by qPCR and western blot (d). e. Verification of direct correlation of Gal-1 and miR-22 through the putative binding site of miR-22 and Gal-1 3’UTR mRNA. Dual luciferase assay revealed the reduction in luciferase activity when miR-22 mimic was co-treated in these cells, compared to negative control (NC). f. A negative correlation was identified in HCC clinical samples when comparing Gal-1 and miR-22 expression in non-tumorous (NT) and tumorous (T) tissues of HCC patients. g. miR-22 expression was analyzed in 81 paired HCC cases. Comparing tumor and non-tumor samples, miR-22 was found to be underexpressed in 49.4% of the cases (40/81). miR-22 expression was normalized with RNU6, a housekeeping gene for data normalization. h. The overall expression of miR-22 was reduced in tumorous tissues when compared to non-tumorous tissues. P < 0.05 is considered statistically significant