MicroRNA-375 sensitizes tumour necrosis factor-alpha (TNF-α)-induced apoptosis in head and neck squamous cell carcinoma in vitro

We investigated the effect of microRNA-375 (miR-375) on tumour necrosis factor-alpha (TNF-α)-induced cell death in head and neck squamous cell carcinoma, and further explored the potential molecular mechanism underlying this phenomenon. Cal27 cells were transfected with miR-375 mimic and subsequently treated with or without TNF-α (10 ng/ml). An additional group of cells were treated with TNF-α alone. The resulting morphological changes were observed, and the percentage of sub-G1 cells was measured. The protein expression and cleavage of caspase 3, caspase 8, and poly(ADP ribose) polymerase (PARP) were determined through Western blotting. The results showed a significant increase in cell death in the combination group, but not in the groups treated with miR-375 mimic, TNF-α alone, or control. The data obtained from sub-G1 cells supported the notion that miR-375 increases the accumulation of sub-G1. In the combination group, the degradation of caspase 3, caspase 8, and PARP was observed and the cleavage of these enzymes was detected. The pan-caspase inhibitor, Z-VAD, inhibited the apoptosis of Cal27 cells treated with a combination of miR-375 mimic and TNF-α. In addition, the apoptosis inhibitory proteins, cFLIP-L and cIAP1, were down-regulated in a time-dependent manner. Taken together, these data suggest that miR-375 sensitizes TNF-α-induced apoptosis, and the reduction in the expression of the apoptosis inhibitory proteins cFLIP-L and cIAP2 plays an important role in this sensitization.