L Gao1, X Zhang2, S Dou1, X Yue1, J Yang1. 1. School of Medicine, Xijing University, Xi'an 710000, China. 2. Department of Otolaryngology, Affiliated Hospital of Shaanxi University of Chinese Medicine, Xianyang 712000, China.
Abstract
OBJECTIVE: To investigate the effects of RNA interference of long noncoding RNA FOXCUT on epithelial mesenchymal transformation and mitochondrial function in nasopharyngeal carcinoma (NPC) cells. METHODS: FOXCUT expression levels were detected by RT-PCR in tumor tissues and adjacent tissues from 50 patients with NPC and in NP69, CNE1, CNE2, SUNE2, HER2 and 5-8F cell lines. CNE1 cells were transfected with a short hairpin RNA (shRNA) targeting FOXCUT or a negative control RNA construct (shRNA-NC), and the changes in cell proliferation and morphology were assessed with CCK8 assay, clone formation assay and microscopic observation. An immunofluorescence assay was used to examine the vimentin-positive cells, and the levels of SOD, MDA and LDH in the cells were detected. The changes of mitochondrial membrane potential were detected with flow cytometry, and the expression levels of E-cad, N-cad, vimentin, Bax, Bcl-2, caspase-3 and c-Myc in the cells were detected with Western blotting. RESULTS: The expression level of FOXCUT was significantly increased in NPC tissues as compared with the adjacent tissues (P < 0.001). Compared with NP69 cells, CNE1, CNE2, SUNE2, HER2 and 5-8F cells all exhibited significantly increased expressions of FOXCUT (P < 0.001). In CNE1 cells, transfection with FOXCUT shRNA significantly inhibited cell proliferation and clone formation (P < 0.001), and caused obvious changes in cell morphology. FOXCUT knockdown significantly decreased the expressions of N-cad and vimentin, increased E- cad expression and the contents of MDA and LDH (P < 0.05), reduced vimentin- positive cells and the activity of SOD, and caused a shift of red fluorescent cells to green fluorescent cells and an increased percentage of green fluorescent cells. FOXCUT knockdown also resulted in significantly increased expressions of Bax/Bcl2 and cleaved Cas3/Cas3 and a lowered expression of c-Myc. CONCLUSIONS: Interference of FOXCUT can inhibit the proliferation and epithelial-mesenchymal transformation, enhance oxidative stress, induce mitochondrial function injury, and promote apoptosis in NPC cells, suggesting the potential of FOXCUT interference for targeted treatment of NPC.
OBJECTIVE: To investigate the effects of RNA interference of long noncoding RNA FOXCUT on epithelial mesenchymal transformation and mitochondrial function in nasopharyngeal carcinoma (NPC) cells. METHODS: FOXCUT expression levels were detected by RT-PCR in tumor tissues and adjacent tissues from 50 patients with NPC and in NP69, CNE1, CNE2, SUNE2, HER2 and 5-8F cell lines. CNE1 cells were transfected with a short hairpin RNA (shRNA) targeting FOXCUT or a negative control RNA construct (shRNA-NC), and the changes in cell proliferation and morphology were assessed with CCK8 assay, clone formation assay and microscopic observation. An immunofluorescence assay was used to examine the vimentin-positive cells, and the levels of SOD, MDA and LDH in the cells were detected. The changes of mitochondrial membrane potential were detected with flow cytometry, and the expression levels of E-cad, N-cad, vimentin, Bax, Bcl-2, caspase-3 and c-Myc in the cells were detected with Western blotting. RESULTS: The expression level of FOXCUT was significantly increased in NPC tissues as compared with the adjacent tissues (P < 0.001). Compared with NP69 cells, CNE1, CNE2, SUNE2, HER2 and 5-8F cells all exhibited significantly increased expressions of FOXCUT (P < 0.001). In CNE1 cells, transfection with FOXCUT shRNA significantly inhibited cell proliferation and clone formation (P < 0.001), and caused obvious changes in cell morphology. FOXCUT knockdown significantly decreased the expressions of N-cad and vimentin, increased E- cad expression and the contents of MDA and LDH (P < 0.05), reduced vimentin- positive cells and the activity of SOD, and caused a shift of red fluorescent cells to green fluorescent cells and an increased percentage of green fluorescent cells. FOXCUT knockdown also resulted in significantly increased expressions of Bax/Bcl2 and cleaved Cas3/Cas3 and a lowered expression of c-Myc. CONCLUSIONS: Interference of FOXCUT can inhibit the proliferation and epithelial-mesenchymal transformation, enhance oxidative stress, induce mitochondrial function injury, and promote apoptosis in NPC cells, suggesting the potential of FOXCUT interference for targeted treatment of NPC.
Authors: Artur Peter Günther Dingeldein; Šárka Pokorná; Martin Lidman; Tobias Sparrman; Radek Šachl; Martin Hof; Gerhard Gröbner Journal: Biophys J Date: 2017-05-23 Impact factor: 4.033