| Background: Nasopharyngeal carcinoma(NPC)is the most common head and neck cancer in China.Recent epidemiological studies have shown that the 5-year survival rate of NPC in China is about80%.At present,the diagnosis and treatment mode of NPC is relatively simple,and there is no chemotherapy drug that can specifically block tumor progression according to the disease process with great difference in pathological type and tumor differentiation degree or different clinical symptoms and patients' personal conditions.This requires that more new anti-tumor drugs be further explored in the future Research of NPC diagnosis and treatment,so as to better adapt to different patient characteristics and provide more effective antitumor drugs Multiple individualized treatment plan.Quinone oxidoreductase(NQO1)gene is a gene sequence located in the q12-q22 region of human chromosome 16.Previous studies have shown that NQO1 expression level is closely correlated with the progression and prognosis of patients with head and neck squamous cell carcinoma(HNSCC).NQO1 is highly expressed in HNSCC,suggesting that NQO1 may be a new target for HNSCC treatment.Chemotherapy drugs represented by ?-lapaquinone are effective inhibitors of NQO1.?-lapachone is a kind of naphthoquinone compounds obtained from natural plant structure in central and South America.In recent years,researchers have also found that ?-lapachone can show significant antitumor activity.More in-depth studies have shown that tumor cells in breast,digestive tract,respiratory tract and urinary system show significant growth inhibition under the action of ?-lapachone.At present,the role of ?-lapachone in the treatment of nasopharyngeal carcinoma and its related mechanisms are less,so it may be of great value to explore the efficacy of ?-lapachone in the treatment of NPC.Methods: Firstly,the expression levels of NQO1 protein in NPC and normal tissues were detected and compared by immunohistochemistry;CCK-8 experiment was used to explore and compare the effects of different doses of ?-lapachone on the survival rate of NPC HNE1 cells and normal nasopharyngeal epithelial cells np460;the tumor volume was monitored and recorded after the intervention of ?-lapachone And quality changes.Secondly,we evaluated the function of ?-lapachone in inhibiting the invasion and metastasis of HNE1 cells by Transwell test and invasion test.Thirdly,we detected the expression level of reactive oxygen species(ROS)in HNE1 cells after intervention by using h2dcf-da immunofluorescence probe technology,and detected mitochondrial membrane potential(MMP)by rhodamine123 fluorescence staining,so as to further explore the molecular mechanism of ?-lapachone inhibiting HNE1 cells.Finally,The expression of p62 protein and membrane associated protein lc3II/ I in HNE1 and subcutaneous tumor cells of nude mice were detected by Western blot.The function and value of autophagy in anti NPC process of ?-lapachone were discussed at cell and animal levels.we also detected mammalian target of rapamycin(m TOR),phosphatidylinositol-3 kinase(PI3K),protein kinase B(AKT)and other cytokines closely related to the occurrence and development of NPC in HNE1 and HNE1 subcutaneous tumor cells inoculated with HNE1 after different doses of ?-lapachone.Results: The results of immunohistochemistry showed that the positive rate and strong positive rate of NQO1 in NPC group were significantly higher than those in normal group;the strong positive rate of NPC group in stage III + IV was significantly higher than that in stage I + II group;the strong positive rate in NPC group with lymph node metastasis was significantly higher than that in group without lymph node metastasis.Secondly,CCK-8 test showed that the survival rate of HNE1 and np460 cells treated with different concentrations of ?-lapachone for 24 hours decreased in a dose-dependent manner.The semi inhibitory concentration(IC50)of?-lapachone on HNE1 cells and np460 cells were 30 ? mol / L and 100 ? mol / L,respectively.In addition,the results of subcutaneous tumorigenesis model of HNE1 cells in nude mice showed that with the increase of dose,?-lapachone could significantly inhibit the growth of NPC cells.Transwell assay showed that the migration function of HNE1 cells decreased in a dose-dependent manner with the increase of ?-lapachone concentration in the culture medium;further invasion experiment showed that the invasion function of HNE1 cells decreased significantly with the increase of ?-lapachone concentration in culture medium.The results of H2DCF-DA immunofluorescence probe and rhodamine 123 fluorescence staining showed that with the increase of ?-lapachone concentration,ROS level in HNE1 cells increased in a dose-dependent manner,while MMP concentration decreased significantly.Finally,The number of autophagic lysosomes in HNE1 and subcutaneous tumor cells of nude mice increased with the increase of ?-lapachone concentration.The results of Western blotting showed that the expression of p62 protein in HNE1 and subcutaneous tumor cells decreased significantly with the increase of ?-lapachone concentration.The expression level of lc3 ii protein increased significantly in a dosedependent manner,while the expression level of lc3 i decreased significantly in a dose-dependent manner.Western blotting showed that the expression levels of phosphorylated PI3 K,phosphorylated AKT and phosphorylated m TOR proteins in HNE1 and subcutaneous tumor cells of nude mice decreased significantly with the increase of ?-lapachone concentration,while the expression levels of PI3 K,AKT and m TOR did not change significantly with the increase of ?-lapachone concentration.Conclusion: The expression level of NQO1 in NPC cells was significantly higher than that in normal nasopharyngeal epithelial cells,and the expression level of NQO1 in advanced tumor cells was significantly higher.?-lapachone can significantly inhibit the survival,migration and distant invasion of NPC cells,and its cytotoxicity increases in a dose-dependent manner.The results suggest that its antitumor activity may be closely related to the expression levels of ROS and MMP,autophagyand the phosphorylation of proteins in PI3K-AKT-m TOR signaling pathway. |
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