高危型HPV基因型检测、分流及HPV16型变异体在宫颈癌及癌前病变中的分布研究

发布时间：2018-05-15 06:31

本文选题：人乳头状瘤病毒 + 宫颈癌　；参考：《新疆医科大学》2015年博士论文

【摘要】：目的:评价高危型HPV基因型检测在宫颈癌筛查中的可行性及应用价值。P16/Ki-67细胞学双染在HPV阳性、宫颈不同病变脱落细胞中的表达和意义。探讨新疆地区感染HPV16变异体谱系分布及E6、E7、LCR基因突变研究,分析HPV16变异体在宫颈癌及癌前病变发生发展中的作用。方法:应用高危型HPV基因检测系统,对2014年09月~2015年8月在新疆自治区人民医院住院及门诊就诊行HC2、TCT检测、并有病理结果的564位女性的宫颈细胞标本进行HPV基因型检测,对其中59例HC2阳性患者的液基细胞学涂片用免疫细胞化学法检测P16/Ki-67蛋白在宫颈脱落细胞中的表达。对HPV16阳性的宫颈癌及癌前病变患者,提取基因组DNA,利用PCR扩增HPV16 DNA E6、E7基因及LCR区核苷酸片段,正反向测序。与HPV16基因序列分析比对,确定HPV16谱系分布,分析核苷酸突变位点。结果:(1)高危型HPV基因型检测在诊断CIN2+病变时优于HC2、TCT、Hybr Miax分型检测,差异具有统计学意义(P0.05),不同液基细胞学分级及组织病理分级中HR-HPV感染率不同,差异具有统计学意义(P0.05),随细胞学及病理学诊断级别升高,HR-HPV感染率上升(P0.05),HPV16型感染率上升(P0.05)。各级别宫颈病变以HPV16型及合并HPV16型感染为主,HP16型分布在维吾尔族和汉族妇女之间无统计学差异(P0.05)。各年龄段高危型HPV的构成比无统计学差异(P0.05)。高危型HPV基因型检测对ASCUS分流优于HC2。(2)P16/Ki-67细胞学双染检测诊断CIN2+病变时具有高灵敏度(92%)和总符合率(78%),P16/Ki-67细胞学双染检测与组织病理学结果符合率较一致(Kappa=0.569,P0.05),在对ASCUS/ASC-H进行P16/Ki-67细胞学双染检测更能发现宫颈高级别病变(P0.05),不同宫颈病变P16/Ki-67细胞学双染检测阳性率:炎症39.13%(9/23)CIN1 20%(2/10),CIN2-3 91.30%(21/23),宫颈癌100%(2/2)差异具有统计学意义(P0.05),随病变程度的加重,P16/Ki-67细胞学双染阳性率呈上升的趋势。(3)E6基因突变率为80.00%(92/115)主要突变位点T350G(58.78%),T178G(18.47%),E7突变率为54.78%(63/115)主要突变位点A647G(33.33%),T846C(26.98%),LCR突变率为23.48%(27/115),主要突变位点为C24T(74.07%),C13T(25.92%)。维吾尔族妇女与汉族妇女之间比较,维吾尔族T350G突变率显著高于汉族,汉族A647G、T846C、C24T突变率显著高于维吾尔族,差异具有统计学意义(P0.05)。维吾尔族宫颈癌组T350G突变率显著高于炎症组(P0.05),且随病变严重程度增加突变率上升,汉族T350G、A647G、T846C、C24T突变率炎症组、宫颈病变组显著高于宫颈癌组(P0.05),维吾尔族C24T突变率炎症组显著高于宫颈癌组(P0.05),差异均具有统计学意义(P0.05)。新疆汉族人中感染的HPV16主要为亚洲型,新疆维吾尔族感染的HPV16主要为欧洲型。结论:高危型HPV感染是宫颈癌前病变和宫颈癌的主要致病因素;宫颈各级别病变中以HPV16型及合并HPV16型感染为主;随细胞学及病理学诊断级别升高HR-HPV感染率上升;高危型HPV基因型检测对ASCUS诊断分流优于HC2。P16/Ki-67细胞学双染阳性率随着宫颈病变程度加重呈上升趋势;P16/Ki-67细胞学双染检测可以有效对ASCUS及HPV阳性者进行诊断分流,具有临床应用价值。HPV16E6、E7突变可能与宫颈病变进展有关,T350G突变可能是维吾尔族宫颈癌高发的原因之一;新疆汉族人中感染的HPV16型主要为亚洲型,新疆维吾尔族感染的HPV16型主要为欧洲型。
[Abstract]:Objective: To evaluate the feasibility and application value of high risk HPV genotyping in screening cervical cancer and its value of.P16/Ki-67 cytology in HPV positive and cervical diseased cells. The distribution of HPV16 variants in Xinjiang area and the mutation of E6, E7, LCR gene and HPV16 variant in cervical cancer and cancer were analyzed. Methods: the role of high risk HPV gene detection system was used to detect the HC2, TCT test and HPV genotypes of 564 women's cervical cells in the hospital of Xinjiang Autonomous Region People's Hospital in Xinjiang Autonomous Region People's Hospital in August 2014, and 59 cases of HC2 positive patients with liquid based cytology. The expression of P16/Ki-67 protein in cervix exfoliated cells was detected by immunocytochemical method. Genomic DNA was extracted from HPV16 positive cervical cancer and precancerous lesions, HPV16 DNA E6, E7 gene and LCR region nucleotide fragment were amplified by PCR, and the sequence analysis of HPV16 gene was compared to determine the HPV16 lineage distribution and analysis of nucleoside. Acid mutation sites. Results: (1) high risk HPV genotypes were superior to HC2, TCT, Hybr Miax typing in the diagnosis of CIN2+ lesions, and the difference was statistically significant (P0.05). The rate of HR-HPV infection in different liquid based cytological classification and histopathological classification was different, the difference had the significance of total count (P0.05), and the diagnostic grade of cytology and pathology increased, H. The rate of R-HPV infection increased (P0.05) and the rate of HPV16 type infection increased (P0.05). There was no statistical difference between the Uygur and the Han women (P0.05) in all levels of cervical lesions (P0.05). The composition of high-risk HPV in all ages was less than that of the Han and Han women (P0.05). The high risk type HPV genotypic detection of ASCUS shunt (P0.05). High sensitivity (92%) and total coincidence rate (78%) were superior to HC2. (2) P16/Ki-67 cytological double staining for diagnosis of CIN2+ lesions. The coincidence rate of double staining and histopathology of P16/Ki-67 cytology was consistent (Kappa=0.569, P0.05). In ASCUS/ASC-H, double staining of P16/Ki-67 cytology was more capable of detecting high grade cervical lesions (P0.05). The positive rate of P16/Ki-67 cytology double staining for cervical lesions: 39.13% (9/23) CIN1 20% (2/10), CIN2-3 91.30% (21/23), and 100% (2/2) of cervical cancer (P0.05), with the aggravation of the lesion, the positive rate of double staining of P16/Ki-67 cytology was upward trend. (3) E6 gene mutation rate was 80% (92/115) main mutation site T350G (58). .78%), T178G (18.47%), E7 mutation rate is 54.78% (63/115) major mutation site A647G (33.33%), T846C (26.98%), LCR mutation rate is 23.48% (27/115), the main mutation site is C24T (74.07%), C13T (25.92%). Uygur women and Han women compared to the Han, the Uygur T350G mutation rate is significantly higher than the Han, the Han A647G, significant mutation rate is significant Higher than Uygur, the difference was statistically significant (P0.05). The mutation rate of T350G in Uygur cervical cancer group was significantly higher than that in the inflammatory group (P0.05), and the mutation rate increased with the severity of the lesion, the Han T350G, A647G, T846C, C24T mutation rate in the inflammatory group, the cervical lesion group was significantly higher than the cervical cancer group (P0.05), and the Uygur C24T mutation rate inflammation group showed a significant difference. The difference was statistically significant (P0.05) higher than the cervical cancer group (P0.05). The infection of HPV16 in Xinjiang Han people was mainly Asian type and the HPV16 of Xinjiang Uygur infection was mainly European type. Conclusion: high risk HPV infection is the main cause of cervical cancer and precancerous lesion and cervical cancer in all levels of the cervix with HPV16 and HPV16 HR-HPV infection rate increased with cytological and pathological diagnosis level, and high risk HPV genotyping was superior to HC2.P16/Ki-67 cytology with double staining of ASCUS diagnostic shunt as the degree of cervical lesions increased; double staining of P16/Ki-67 cytology could be used to diagnose and shunt of ASCUS and HPV positive patients. With the clinical value of.HPV16E6, E7 mutation may be associated with the progression of cervical lesions. T350G mutation may be one of the reasons for the high incidence of cervical cancer in Uygur nationality; the HPV16 type of infection in Xinjiang Han people is mainly Asian type, and the HPV16 type of Xinjiang Uygur infection is mainly European type.

【学位授予单位】：新疆医科大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R737.33

【参考文献】