VEGF與Cyclin D1基因多型性與台灣口腔鱗狀上皮細胞癌之相關性探討

背景：細胞週期失去調控和血管新生與腫瘤的發生有關，血管內皮生長因子 (Vascular endothelial growth factor；VEGF) 已經被認為是血管生成的重要因子，它參與了許多腫瘤中血液的提供。Cyclin D1參與細胞有絲分裂中G1-S phase變化的檢查點上。但關於VEGF和Cyclin D1基因多型性與口腔組織病變的相關性，目前仍不清楚。研究目的：以PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) 的方法分析VEGF基因在核酸序列在 -460、+405和+936位置的基因多型性和Cyclin D1基因在核酸序列在 +870位置的基因多型性與口腔癌前病變以及口腔鱗狀上皮細胞癌的關係，並觀察VEGF基因在核酸序列在 -460、+405、+936位置的haplotypes與口腔組織病變的相關性；再進一步探討這兩個基因與菸、檳榔、酒間是否有交互作用，進而影響口腔組織病變的發生。研究方法：我們收集OSCC患者324位、口腔黏膜纖維化病患64位和口腔白斑症病患83位。疾病診斷由高雄醫學大學口腔顎面外科病理切片報告確認。331位對照組來自高雄縣某鄉鎮居民及高雄市某工廠員工，經健康篩檢和口腔檢查後排除掉有口腔病變、口腔有明顯發炎症狀、癌症病史、慢性疾病或免疫系統疾病的對象。經簽署同意書後抽取10ml的周邊血液來萃取DNA，以PCR-RFLP的方法獲得VEGF及Cyclin D1的基因多型性，並回答一份標準化問卷來收集研究對象的相關資料。將所有的資料以Excel與Access軟體建檔，在轉入JMP5.01統計軟體進行分析。結果：我們發現經調整年齡、性別、種族、教育程度以及菸、檳榔、酒的使用後，Cyclin D1 +870GG基因型會降低罹患口腔鱗狀上皮細胞癌 (OR = 0.48，95% CI = 0.23 - 0.98)，尤其是在沒有咀嚼檳榔者 (ORadjusted = 0.30，95% CI = 0.07 - 0.98) 和沒有喝酒習慣者 (ORadjusted = 0.14，95% CI = 0.03 - 0.49) 中，而VEGF 基因在-460C>T、+405C>G、+936C>T等基因多型性與口腔病變間無明顯相關。但在抽菸者中觀察到攜帶VEGF -460(CT+CC) 基因型可能為口腔鱗狀上皮細胞癌發生的危險因子 (OR adjusted = 2.11，95% CI = 1.05 - 4.43)。在喝酒者中攜帶Cyclin D1 +870GG基因型可能是促使口腔白斑症惡化成口腔鱗狀上皮細胞癌的危險因子 (OR adjusted = 2.20，95% CI = 1.03 - 4.98)；在沒有喝酒者中攜帶Cyclin D1 +870GG基因型會降低促使口腔白斑症惡化成口腔鱗狀上皮細胞癌的危險性 (OR adjusted = 0.15，95% CI = 0.02 - 0.87)。進一步依研究對象的不同基因型進行分層分析，發現在不同的VEGF基因型的分組下，可以觀察到菸、檳榔、酒的使用習慣對口腔鱗狀上皮細胞癌的危險性是有不同的。結論：我們建議Cyclin D1基因在核酸序列 +870位置基因多型性會與檳榔、酒的使用習慣間有交互作用影響罹患口腔鱗狀上皮細胞癌的相關性，且喝酒與否會改變Cyclin D1基因在核酸序列 +870位置的基因多型性與口腔白斑症癌化成口腔鱗狀上皮細胞癌的相關性。VEGF基因在核酸序列-460、+405、+936位置基因多型性可能會改變菸、檳榔、酒與口腔黏膜下纖維化症轉變成口腔鱗狀上皮細胞癌、罹患口腔鱗狀上皮細胞癌、口腔黏膜下纖維化症和口腔白斑症的相關性。

關鍵字

口腔鱗狀上皮細胞癌；口腔黏膜下纖維化症；口腔白斑症； VEGF ； Cyclin D1 ；基因多型性

並列摘要

Background: Vascular endothelial growth factor (VEGF) has been considered to be a critical angiogenic cytokine involved in the development of several tumors. The dysfunction in the control of cell cycle accelerates the tumor development. The cyclin D1 gene was involved in the G1-S checkpoint of cell cycle. However, the association between polymorphisms of VEGF and cyclin D1 genes and oral squamous cell carcinoma remain unclear. Study objective: Our aims were to investigate the risks of VEGF -460C>T, +405C>G, and +936C>T polymorphisms and Cyclin D1 +870A>G polymorphisms to OSCC. Methods: A total of 324 oral squamous cell carcinoma (OSCC), 64 oral submucosal fibrosis (OSF) and 83 oral leukoplakia (OL) patients were recruited from the department of Oral and Maxillofacial Surgery, Kaohsiung Medical University Hospital. A total of 331 participants were recruited from a community health survey and a company health examination in the southern Taiwan. After participants signed the informed consent, we collected 10ml of peripheral blood for DNA isolation. A standardized questionnaire was applied to collect the information about substance usage and demographic data. The polymorphisms of VEGF -460C>T, +405C>G, and +936 C>T, and Cyclin D1 +870A>G were determined by PCR-RFLP methods. The JMP 5.01 statistical software was used to analyze the association. Results: After adjusting with age, gender, ethnicity, educational levels, smoking, drinking and betel quid chewing , an association of the cyclin D1 +870CC genotype with lower risk of OSCC (OR adjusted = 0.48, 95% CI = 0.23 - 0.98), particularly in non-chewers (ORadjusted = 0.30，95% CI = 0.07 - 0.98) and in non-drinkers (ORadjusted = 0.14，95% CI = 0.03 - 0.49). No association was found between VEGF -460C>T, +405C>G, and +936C>T polymorphisms and OSCC, OSF or OL. Stratification with the habits of substance usage or not, VEGF -460(CT+CC) genotype was associated with OSCC development in smokers (OR adjusted = 2.11, 95% CI = 1.05 - 4.43). Cyclin D1 +870GG genotype was a risk factor for malignant potential of OL in drinkers (OR adjusted = 2.20，95% CI = 1.03 - 4.98); however an association of the cyclin D1 +870CC genotype with lower risk for malignant potential of OL in non-drinkers (OR adjusted = 0.15，95% CI = 0.02 - 0.87). Further to analyze the risks of the VEGF -460C>T, +405C>G, and +936C>T polymorphisms in the OSCC development, malignant potential of oral precancerous lesion, and development of oral precancerous lesions. We stratified the participants by the VEGF genotypes to determine the association between the habits of three substances’ usage and oral lesions. The association between the habits of three substances’ usage and oral lesions could be differentiated by the different VEGF genotypes. Conclusion: We suggest that an effect of the interaction between the habits of betel quid chewing and alcohol drinking, and Cyclin D1 +870A>G polymorphisms on OSCC development. The association of Cyclin D1 +870A>G polymorphisms with malignant potential of OL was affected by the habit of alcohol consumption. The association between the habits of three substances’ usage and malignant potential of OSF, OSCC development, OSF development, OL development were differentiated by the VEGF -460C>T, +405C>G, and +936C>T polymorphisms.