共同引物PCR/RFLP方法对直肠癌组织中人乳头瘤病毒的检测及分型

Detecting and Typing of Human Papillomaviruses in Rectal Carcinoma Tissues by Polymerase Chain Reaction with Consensus Primers and Restriction Fragment Length Polymorphism

为探讨人乳头瘤病毒（ＨＰＶ）与直肠癌的关系，应用共同引物聚合酶链式反应扩增ＨＰＶ高保守区Ｌ１区序列，扩增产物经限制性内切酶ＤｄｅＩ，ＸｂａＩ，ＡｃｃＩ，ＰｓｔＩ酶切后，进行限制性片段长度多态性分析（ＲＦＬＰ），可对ＨＰＶ６，１１，１６，１８等常见的型别进行快速检测及鉴定。用该法对１１２例直肠癌和６３例正常直肠粘膜组织中ＨＰＶ进行检测。结果：直肠癌ＨＰＶ检出率（１８．７５％）与正常直肠粘膜（７．９３％）差异无显著性（Ｐ＞０．０５），但直肠癌ＨＰＶ１６检出率（１６．０７％）高于正常直肠粘膜（１．５８％），差异有高度显著性（Ｐ＜０．０１），表明ＨＰＶ１６可能与直肠癌的发生有密切关系，是直肠癌的高危致癌因素。该检测系统敏感性高，可检出每个二倍体细胞内０．１拷贝的ＨＰＶ１６ＤＮＡ，操作简便，适用于大批量标本ＨＰＶ的筛检。

In order to discuss the association between human papillomaviruses (HPV) and rectal carcinomas, many types of HPVs were detected by polymerase chain reaction (PCR) with a pair of consensus primers for L 1 region. The amplified HPV DNA could be typed by subsequent restriction mapped with Dde I, Xba I, Acc I, Pst I. HPV6,11,16,18 were easily typed by this method. The four types of HPVs above are very common. Detected were 112 cases of rectal carcinomas and 63 cases of normal rectal mucosa. HPV was found in 18.75% of rectal carcinomas (18 positive for HPV 16,2 for HPV18, 1 for HPV6/11) and in 7.93% of normal mucosa (1 positive for HPV16,3 for HPV6/11,1 for unknown). There was no significant difference ( P >0.05). HPV16 DNA sequences were found in 16.07% of the rectal carcinoma group, and in 1.58% of the normal group. There was significant difference in the detective rate of HPV 16 between the two groups( P <0.01). This study shows that HPV16 may have close relation to occurrence of rectal carcinoma and may be the high risk factor of carcinoma. This PCR system can detect 0.1 copy per cell of HPV16 DNA. The system is easy, sensitive and very useful for detecting HPV DNA among a large number of samples, and worth using widely.