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目的:研究人乳头瘤病毒(HPV)16型LCR基因在湖北地区部分宫颈癌组织中的变异特点,通过系统发生学分析探讨宫颈癌患者感染的HPV16型变异株的流行型别及特征。方法:从22例宫颈癌手术切除标本中提取组织DNA,用HPV16LCR特异性引物进行PCR扩增,对扩增的LCR产物片段进行测序,用Paup 4.0软件使用邻接法对HPV16变异株进行系统发生学分析。结果:在22例宫颈癌组织中LCR基因存在着12个位点的基因变异。高频率突变位点有5个。其中第7 521位核苷酸由G变异为A,变异频率为100%;第7 730位核苷酸由A变异为C,变异频率为86.4%,第7 842位核苷酸由G变异为A,变异频率为77.3%,第7 714位由T变异为G,变异频率为31.8%;第7 930位核苷酸由C变异为T,变异频率为31.8%。HPV 16LCR中的5个高频位点,均位于可与多个转录因子结合的区域。对LCR基因进行的系统发生学分析表明,湖北地区流行的HPV16病毒株主要为亚洲型(As)变异体(77.2%),其次为欧洲型(E)(22.8%),没有发现非洲-1型(Af-1),非洲-2型(Af-2)和亚洲美洲型(AA)HPV16变异体。结论:湖北地区宫颈癌组织中LCR存在着一定程度的基因变异,其中一些高频位点与国内、外报道的结果有明显差异,可能是湖北地区LCR的突变热点。
Objective: To study the variation of human papillomavirus (HPV) 16 LCR gene in some cervical cancer tissues in Hubei Province. To investigate the prevalence and characteristics of HPV16 variant in cervical cancer patients by phylogenetic analysis. Methods: Tissue DNA was extracted from 22 cases of cervical cancer. The HPV16LCR specific primers were used for PCR amplification. The amplified fragment of LCR product was sequenced. Using the Paup 4.0 software, the phylogenetic analysis of HPV16 variants analysis. Results: There were 12 loci in LCR gene in 22 cases of cervical cancer. There are 5 high-frequency mutation sites. The nucleotide variation from nucleotide 7 to nucleotide 521 was A with a mutation frequency of 100%. The nucleotide from nucleotide 7 to nucleotide A was C with a mutation frequency of 86.4%. The nucleotide variation from nucleotide 7 to nucleotide 842 was A, the frequency of variation was 77.3%, the 7th 714th position was changed from T to G, the mutation frequency was 31.8%; the 7th 930th nucleotide changed from C to T, and the mutation frequency was 31.8%. The five high frequency sites in the HPV 16 LCR are located in areas that can bind to multiple transcription factors. Phylogenetic analysis of LCR genes showed that the predominant strains of HPV16 in Hubei were mainly As (As) variant (77.2%), followed by European (E) (22.8%). No African-1 (Af-1), Af-2 and Asian American Type AA (HPV16) variants. Conclusion: There are some genetic variations in LCR in Hubei Province. Some of the high frequency loci have significant differences with those reported in China and other countries, which may be the hot spot of LCR mutation in Hubei Province.