PC cells were cultured under normoxic and hypoxic conditions. Expression of miR-210 and hypoxia-inducible factor (HIF)-1伪 was detected using quantitative reverse-transcription polymerase chain reaction. Cancer cells were transiently transfected with HIF-1伪 small interfering RNA (siRNA) and miR-210 mimics, and cell proliferation was measured using the CCK-8 assay. Potential targets for miR-210 were then identified using a dual luciferase reporter assay.
Hypoxic conditions induced miR-210 expression in six PC cell lines (AsPC-1, BxPC-3, MIAPaCa-2, PANC-1, Su86.86 and SW1990), but not in Capan-1 or T3M4 cells. Transfection of HIF-1伪 siRNA into PANC-1 cells markedly inhibited HIF-1伪 expression, and subsequently down-regulated miR-210 expression under hypoxic conditions. MiR-210 had no observable impact on the proliferation of PANC-1 or Su86.86 cells and dual luciferase reporter assays showed significantly reduced luciferase activity in the wildtype E2F3, EFNA3, GIT2, MNT, ZNF462 and EGR3 constructs, compared to the corresponding mutants, but not in HOXA3.
These results suggest that miR-210 expression in PC cells is induced by hypoxia through a HIF-1伪-dependent pathway, but does not influence PC cell proliferation. Also, E2F3, EFNA3, GIT2, MNT, ZNF462 and EGR3 may be potential miR-210 targets in PC.
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