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Abstract：
Glycolysis is a key energy-providing process and one of the hallmarks of cancer. Nitric oxide (NO), a free radical molecule, regulates glycolysis in various cancers. NO can alter the cell cycle and apoptosis in head and neck squamous cell carcinoma (HNSCC) cells. However, the effect of NO on glycolysis in HNSCC cells remains unresolved. The present study investigated the effects of NO on cell proliferation, glucose transporter (GLUT) gene expression and glycolytic indicators in HNSCC cell lines. Two pairs of isogenic HNSCC cell lines, HN18/HN17 and HN30/HN31, were treated with a NO donor, diethylamine NONOate (DEA-NONOate), for 24, 48 and 72 h. Cell proliferation was assessed using MTT assay and NO concentration was measured using the Griess Reagent System. GLUT1, GLUT2, GLUT3, and GLUT4 gene expression was analyzed using reverse transcription-quantitative PCR. Furthermore, hexokinase (HK) activity and lactate production were measured in NO-treated cells using colorimetric assay. NO exhibited concentration-dependent pro- and anti-proliferative effects on the HNSCC cell lines. Lower NO concentrations (5-200 µM) had pro-proliferative effects, whereas NO >200 µM had an anti-proliferative effect on HNSCC cells. NO (5 µM) promoted proliferation and glycolysis in HN18 cells by upregulating GLUT1 and GLUT2 gene expression and increasing HK activity and lactate levels. At 5-20 µM, NO-induced HN17 and HN30 cells demonstrated enhanced proliferation and GLUT2, GLUT3 and GLUT4 gene expression, whereas the glycolytic pathway was not affected. In conclusion, the present study demonstrated distinct proliferative effects of NO on HNSCC cells. NO may promote cell proliferation by stimulating glucose consumption and the glycolytic rate in HN18 cells. The effects of NO in other cell lines may be mediated by a non-glycolysis mechanism and require further investigation.
摘要：
糖酵解是一个关键的能量提供过程，也是癌症的标志之一。一氧化氮(NO),自由基分子,调节各种癌症的糖酵解。NO可以改变头颈部鳞状细胞癌（HNSCC）细胞的细胞周期和凋亡。然而,NO对HNSCC细胞糖酵解的影响仍未解决。本研究探讨了NO对细胞增殖的影响,HNSCC细胞系中葡萄糖转运蛋白（GLUT）基因表达和糖酵解指标。两对等基因HNSCC细胞系，HN18/HN17和HN30/HN31，用NO供体治疗，二乙胺NONOate（DEA-NONOate），24、48和72小时。使用MTT测定法评估细胞增殖，并使用Griess试剂系统测量NO浓度。使用逆转录-定量PCR分析GLUT1、GLUT2、GLUT3和GLUT4基因表达。此外，使用比色测定法在NO处理的细胞中测量己糖激酶(HK)活性和乳酸产生。NO对HNSCC细胞系表现出浓度依赖性的促增殖和抗增殖作用。较低的NO浓度（5-200µM）具有促增殖作用，而NO>200μM对HNSCC细胞具有抗增殖作用。NO（5µM）通过上调GLUT1和GLUT2基因表达并增加HK活性和乳酸水平来促进HN18细胞的增殖和糖酵解。在5-20µM，NO诱导的HN17和HN30细胞表现出增强的增殖和GLUT2，GLUT3和GLUT4基因表达，而糖酵解途径不受影响。总之,本研究证明了NO对HNSCC细胞的明显增殖作用。NO可能通过刺激HN18细胞的葡萄糖消耗和糖酵解率来促进细胞增殖。NO在其他细胞系中的作用可能由非糖酵解机制介导，需要进一步研究。
