Abstract:

Objective To discuss the effect of microRNA-153-3p （miR-153-3p） on the apoptosis of oral squamous cell carcinoma （OSCC） cells， and to clarify its mechanism. Methods The Cancer tissue and adjacent normal tissue were collected from 84 patients diagnosed with OSCC， and five kinds of OSCC cells and normal human oral mucosa HOK-16A cells were cultured. Real-time fluorescence quantitative PCR （RT-qPCR） method was used to detect the expression levels of miR-153-3p in the OSCC cancer tissue and adjacent normal tissue， as well as in the five kinds of cells. The Tca8113 cells were randomly divided into blank control group， mimics NC group，mimics group， mimics+vector group， and mimics+peroxisome proliferator-activated receptor γ coactivator-1α （PGC-1α） group， and were transfected with mimics NC plasmid，miR-153-3p mimic plasmid， PGC-1α overexpression plasmid， or co-transfected with both plasmids. RT-qPCR method was used to detect the expression levels of miR-153-3p in the cells in various groups after transfection； MTT method was used to detect the proliferation activities of the cells in various groups； flow cytometry was used to detect the apoptotic rates of the cells in various groups； 2'，7'- dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence staining was used to detect the levels of reactive oxygen species （ROS） in the cells in various groups； JC-1 probe staining was used to detect the mitochondrial membrane potential of the cells in various groups； Western blotting method was used to detect the expression levels of cytochrome C （Cyt C） protein in the cytoplasm and mitochondria， as well as the expression levels of Cleaved-Caspase-3， PGC-1α， nuclear respiratory factor 1 （NRF-1）， and mitochondrial transcription factor A （TFAM） proteins in the cells in various groups； dual-luciferase reporter assay was used to verify the targeting relationship between miR-153-3p and PGC-1α. Results Compared with adjacent normal tissue and HOK-16A cells， the expression levels of miR-153-3p in OSCC cancer tissue and OSCC cells were significantly decreased （P<0.05）， while the lowest expression level was observed in the Tca8113 cells. After transfection， compared with blank control group and mimics NC group， the expression level of miR-153-3p in the Tca8113 cells in mimics group was significantly increased （P<0.001）； the proliferation activity of the cells at each time point was significantly decreased （P<0.001）； the apoptotic rate and ROS level of the cells were significantly increased （P<0.001）， while the mitochondrial membrane potential of the cells was significantly decreased （P<0.001）； the expression level of Cleaved-Caspase-3 protein in the cells and the expression level of Cyt C protein in the cytoplasm were significantly increased （P<0.001）， while the expression level of Cyt C protein in the mitochondria and the expression levels of PGC-1α/TFAM pathway-related proteins PGC-1α， NRF-1， and TFAM in the cells were significantly decreased （P<0.05）. The dual-luciferase reporter assay results confirmed that miR-153-3p could specifically target and regulate the expression of the PGC-1α gene. After co-transfection， compared with blank control group， the apoptotic rate and ROS level in the cells in mimics group were significantly increased （P<0.001）， while the mitochondrial membrane potential was significantly decreased （P<0.001）； compared with mimics group， there were no significant differences in apoptotic rate， ROS level， and mitochondrial membrane potential in the cells in mimics+vector group （P>0.05）； compared with mimics+vector group， the apoptotic rate and ROS level in the cells in mimics+PGC-1α group were significantly decreased （P<0.01）， while the mitochondrial membrane potential was significantly increased （P<0.001）. Compared with blank control group， the expression levels of PGC-1α， NRF-1， and TFAM proteins in the cells in mimics group were significantly decreased （P<0.001）； compared with mimics+vector group， the expression levels of PGC-1α， NRF-1， and TFAM proteins in the cells in mimics+PGC-1α group were significantly increased （P<0.001）. Conclusion ?Overexpression of miR-153-3p can induce apoptosis in the Tca8113 cells by targeting and down-regulating the expression of PGC-1α， thereby blocking the PGC-1α/TFAM signaling pathway， inducing mitochondrial dysfunction， and activating the mitochondrial apoptotic pathway.

Key words: Oral squamous cell carcinoma, MicroRNA-153-3p, Peroxisome proliferator-activated receptor γ coactivator-1α, Mitochondria, Apoptosis