Abstract
Objective The mechanism of enhanced radiosensitivity induced by mitochondrial uncoupling protein UCP2 was investigated in HeLa cells to provide a theoretical basis as a novel target for cervical cancer treatment. Methods HeLa cells were irradiated with 4 Gy X-radiation at 1.0 Gy/min. The expression of UCP2 mRNA and protein was assayed by real-time quantitative polymerase chain reaction and western blotting. UCP2 siRNA and negative control siRNA fragments were constructed and transfected into HeLa cells 24 h after irradiation. The effect of UCP2 silencing and irradiation on HeLa cells was determined by colony formation, CCK-8 cell viability, γH2AX immunofluorescence assay of DNA damage, Annexin V-FITC/PI apoptosis assay, and propidium iodide cell cycle assay. The effects on mitochondrial structure and function were investigated with fluorescent probes including dichlorodihydrofluorescein diacetate (DCFH-DA) assay of reactive oxygen species (ROS), rhodamine 123, and MitoTracker Green assay of mitochondrial structure and function. Results Irradiation upregulated UCP2 expression, and UCP2 knockdown decreased the survival of irradiated HeLa cells. UCP2 silencing sensitized HeLa cells to irradiation-induced DNA damage and led to increased apoptosis, cell cycle arrest in G2/M, and increased mitochondrial ROS. Increased radiosensitivity was associated with an activation of P53, decreased Bcl-2, Bcl-xl, cyclin B, CDC2, Ku70, and Rad51 expression, and increased Apaf-1, cytochrome c, caspase-3, and caspase-9 expression. Conclusions UCP2 inhibition augmented the radiosensitivity of cervical cancer cells, and it may be a potential target of radiotherapy of advanced cervical cancer.
Highlights
Cervical cancer is a significant threat to women’s health
This study found a significant elevation of UCP2 expression and reactive oxygen species (ROS) production in irradiated HeLa cervical cancer cells that inhibited cell proliferation and promoted apoptosis
UCP2 expression in HeLa cells was assayed at different times after irradiation, and the qPCR and western blot results indicate that both UCP2 mRNA and protein levels increased in response to irradiation (Figure 1)
Summary
Cervical cancer is a significant threat to women’s health. Vaccination or screening for early diagnosis has reduced the incidence of cervical cancer in developed countries, but it is the fourth highest cause of cancer-related death in the developing world [1]. Inhibition of UCP activity increases the formation of ROS, mitochondrial transition pore permeability, and damage to DNA, proteins, and lipids that causes changes in cell growth [7, 8]. Previous studies have found significant associations between mitochondrial UCP2 and tumor grade in primary breast cancer and reduced sensitivity of breast cancer cells to therapeutic agents by increasing oxidative stress, and some studies show that UCP2 has a critical role in chemotherapeutic drug resistance in pancreatic ductal adenocarcincoma [21,22,23]. This study found a significant elevation of UCP2 expression and ROS production in irradiated HeLa cervical cancer cells that inhibited cell proliferation and promoted apoptosis. UCP2 may be a target molecule for the development of novel cervical cancer-specific radiosensitizers
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