白介素17基因启动子区CpG岛甲基化与宫颈癌的相关性研究

Correlation between CpG Island Methylation in IL-17 Gene Promoter and Cervical Cancer

目的探讨白介素17（IL-17）基因启动子区Cp G岛甲基化与宫颈癌的相关性,为临床诊断和免疫治疗宫颈癌提供新思路。方法选取2011年3月—2012年5月在唐山工人医院住院部就诊的宫颈癌患者43例（宫颈癌组）、癌前病变患者62例〔其中宫颈上皮内瘤变（CIN）Ⅰ级23例（CINⅠ组）、CINⅡ~Ⅲ级39例（CINⅡ~Ⅲ组）〕,另选取本院同期因子宫肌瘤行全子宫切除术的患者43例作为对照组。4组患者宫颈组织标本均经手术获得。提取4组宫颈组织DNA并进行高危型人乳头瘤病毒（HPV）16 DNA检测,采用Cp G岛甲基化特异性PCR（MSP）法检测IL-17基因启动子区Cp G岛甲基化状态,采用半定量反转录PCR（RT-PCR）法检测IL-17 mRNA表达水平,分析IL-17基因启动子区Cp G岛甲基化率与IL-17 mRNA表达水平的相关性及其与HPV16 DNA阳性率的关联性,并分析不同年龄、临床特征宫颈癌患者IL-17基因启动子区Cp G岛甲基化率。结果 4组HPV16 DNA阳性率比较,差异有统计学意义（P〈0.001）。CINⅡ~Ⅲ组、宫颈癌组IL-17基因启动子区Cp G岛甲基化率低于对照组（P〈0.01）;CINⅠ组与对照组、CINⅡ~Ⅲ组、宫颈癌组IL-17基因启动子区Cp G岛甲基化率比较,差异无统计学意义（P〉0.05）;CINⅡ~Ⅲ组与宫颈癌组IL-17基因启动子区Cp G岛甲基化率比较,差异无统计学意义（P〉0.05）。CINⅠ组、CINⅡ~Ⅲ组和宫颈癌组IL-17 mRNA表达水平均高于对照组（P〈0.05）;CINⅠ组与CINⅡ~Ⅲ组、宫颈癌组IL-17 mRNA表达水平比较,差异无统计学意义（P〉0.05）;CINⅡ~Ⅲ组与宫颈癌组IL-17 mRNA表达水平比较,差异无统计学意义（P〉0.05）。根据IL-17基因启动子区Cp G岛甲基化检测结果将所有患者分为IL-17基因启动子区Cp G岛甲基化患者（93例）和非甲基化患者（55例）,IL-17基因启动子区Cp G岛非甲基化患者IL-17 mRNA表达水平高于甲基化患者（P〈0.05）。所有患者宫颈组织中IL-17基因启动子区Cp G岛甲基化率与IL-17 mRNA表达水平呈负相关（rs=-0.627,P〈0.05）。所有患者宫颈组织中IL-17基因启动子区Cp G岛甲基化率和HPV16 DNA感染阳性率存在关联性（χ^2=33.209,P〈0.05,r=-0.474）。临床分期为Ⅱ期、Ⅲ期的宫颈癌患者IL-17基因启动子区Cp G岛甲基化率低于临床分期为Ⅰ期的宫颈癌患者（P〈0.05）;不同年龄、病理分级、肿瘤直径及有无淋巴结转移的宫颈癌患者IL-17基因启动子区Cp G岛甲基化率比较,差异无统计学意义（P〉0.05）。结论 IL-17基因启动子区Cp G岛甲基化与高危型HPV16感染具有关联性,IL-17基因启动子区Cp G岛低甲基化可能与宫颈癌发生、发展存在相关性。

Objective To investigate the correlation between Cp G island methylation of IL-17 gene promoter and cervical cancer,and to provide new ideas for clinical diagnosis and immunotherapy of cervical cancer. Methods 43 cervical cancer patients（ cervical cancer group） and 62 patients with precancerous lesions 〔23 cases with CIN Ⅰ（ CINⅠ group）,39 cases with CINⅡ～Ⅲ（ CINⅡ～Ⅲ group） 〕who received treatment in Inpatient Department of Tangshan Gongren Hospital from March 2011 to May 2012 were selected,and another 43 patients who had undergone total abdominal hysterectomy in this hospital at the same period due to uterine fibroids were enrolled as control group. Cervical tissue specimens of patients in four groups were obtained by surgery. DNA in cervical tissue of patients in four group was extracted to make a detection of HPV16 DNA,Cp G island methylation status in IL-17 gene promoter was detected by using Cp G island methylation specific PCR（ MSP）, IL-17 mRNA expression levels were detected by semi- quantitative reverse transcription PCR（ RT- PCR）,correlation between Cp G island methylation rate of IL-17 gene promoter and IL-17 mRNA expression levels,and the relevance of this rate to the positive rate of HPV16 DNA were analyzed, and Cp G island methylation status of IL-17 gene promoter in cervical cancer patients of different ages and clinical features were also analyzed. Results HPV16 DNA positive rates in four groups were significantly different（ P〈0. 001）. Cp G island methylation rate of IL-17 gene promoter in CINⅡ～Ⅲ group and cervical cancer group was significantly lower than that in control group（ P〈0. 01）; Cp G island methylation rates of IL-17 gene promoter in CINⅠ group and control group,CINⅠ group,CINⅡ～Ⅲ group,cervical cancer group were not significantly different（ P〈0. 05）; Cp G island methylation rates of IL-17 gene promoter between CINⅠgroup and CINⅡ～Ⅲ group were not significantly different（ P〈0. 05）. The expression levels of IL-17 mRNA in CINⅠ group,CINⅡ ~ Ⅲ group and cervical cancer group were higher than those in control group（ P〈0. 05）; the expression levels of IL-17 mRNA in CINⅠgroup,CINⅡ～Ⅲ group and cervical cancer group was not significantly different（ P〈0. 05）; the expression level of IL-17 mRNA between CIN Ⅱ～Ⅲ group and cervical cancer group was not significantly different（ P〈0. 05）. According to the detected results of Cp G island methylation in IL-17 gene promoter,all patients were divided into patients with Cp G island methylation of IL-17 gene promoter（ 93 cases） and non-methylation patients（ 55 cases）,IL-17 mRNA expression level of patients with Cp G island non- methylation in IL-17 gene promoter was higher than that in methylation patients（ P〈0. 05）. In all patients＇ cervical tissues,Cp G island methylation rate of IL-17 gene promoter was negatively correlated with its mRNA expression level（ rs=- 0. 627, P〈0. 05）. There was correlation between Cp G island methylation rate of IL-17 gene promoter in all patients＇ cervical tissue and infection positive rates of HPV16 DNA（ χ~2= 33. 209,P〈0. 05,r =- 0. 474）. Cp G island methylation rate of IL-17 gene promoter in cervical cancer patients at Ⅱ and Ⅲ clinical stages were lower than that of patients at Ⅰ clinical stage（ P〈0. 05）; Cp G island methylation rates of IL-17 gene promoter among cervical cancer patients of different ages, pathological grading, tumor diameter, and with or without lymph node metastasis were not significantly different（ P〈0. 05）. Conclusion Cp G island methylation of IL-17 gene promoter is related with high- risk HPV16 infection,Cp G island methylation of IL-17 gene promoter may be associated with the occurrence and development of cervical cancer.