siRNA targeting HIF-1alpha induces apoptosis of pancreatic cancer cells through NF-kappaB-independent and -dependent pathways under hypoxic conditions.

OBJECTIVE: The primary objective was to explore the molecular mechanism of small interference RNA (siRNA) targeting hypoxic inducible factor-1alpha (HIF-1alpha) for inducing apoptosis of pancreatic cancer cells through a nuclear factor-kappaB (NF-kappaB)-independent or -dependent pathway under hypoxic conditions.

MATERIALS AND METHODS: A cassette encoding siRNA targeting HIF-1alpha mediated by recombinant adeno-associated virus (rAAV) was constructed, giving rAAV-siHIF. rAAV-siHIF or rAAV-hrGFP were transfected into exponentially growing MiaPaCa2 cells under hypoxic conditions. The expression of HIF-1alpha mRNA and protein and the activity of NF-kappaB were observed by real-time PCR, Western blot and electromobility shift assay (EMSA). The proliferation and apoptosis of MiaPaCa2 cells with or without PDTC, as an inhibitor of NF-kappaB, were investigated by MTT and TUNEL.

RESULTS: rAAV-siHIF inhibited the expression of HIF-1alpha mRNA and protein and the activity of NF-kappaB in MiaPaCa2 cells under hypoxic conditions. At the same time, rAAV-siHIF decreased MiaPaCa2 cell proliferation and induced apoptosis, but these effects were not abrogated by PDTC. Moreover, PDTC also inhibited MiaPaCa2 cell proliferation and induced apoptosis while rAAV-hrGFP did not have these effects.

CONCLUSION: Under hypoxic conditions, HIF-1alpha plays a key role in the proliferation of MiaPaCa2 cells and inhibition of HIF-1alpha expression may lead to MiaPaCa2 cell apoptosis through NF-kappaB-independent and -dependent pathways.