Li Hui, Zhao Min, Qiu Xiaoping, Ding Xiaohua, Tan Yun, Wu Xinxing
Department of Molecular Biology, Institute of Virology, Wuhan University Medical School, Wuhan 430071, P.R. China.
Eksp Onkol. 2004 Mar;26(1):48-54.
To investigate the variation and biological properties of HPV16 E7 isolated from cervical cancer biopsy samples from highest incidence area in HuBei province of China.
HVP16 E7 sequences isolated from the cervical cancer biopsies of 10 local patients were amplified, sequenced and compared with prototype E7 gene. Then the variant gene was cloned into different vectors to study the antigenicity, expression and immunogenicity of its protein by Western blot, immunofluorescence and genetic immunization in vitro or in vivo.
The results showed that 7 of 10 samples had the same mutations which led to a nonsense mutation at codon 43 of E7 sequence. The truncated E7 protein could be recognized by standard E7 monoclonal antibody in Western blot and expressed in NIH3T3 cells. In the blood sera of mice immunized intramuscularly by the plasmid DNA expressing the variant E7 gene specific E7 antibodies could be detected at week 2, 3, 5 and 6 after inoculation. However, no specific lymphoproliferation after E7 protein stimulation in vitro was detected by MTT colorimetric assay in comparison to the prototype E7 protein.
HPV16 E7 gene may show variation in China and the variant protein could be expressed and induce host humoral immune response, but could not elicit special cellular-immune response against it. These data might hold the key for future development of HPV16 vaccine in HuBei province of China.
研究从中国湖北省宫颈癌高发区活检样本中分离出的人乳头瘤病毒16型E7（HPV16 E7）的变异情况及其生物学特性。
对从10例当地患者宫颈癌活检样本中分离出的HPV16 E7序列进行扩增、测序，并与E7基因原型进行比较。然后将变异基因克隆到不同载体中，通过蛋白质免疫印迹法、免疫荧光法以及体内外基因免疫法研究其蛋白的抗原性、表达情况和免疫原性。
结果显示，10个样本中有7个具有相同的突变，这些突变导致E7序列第43位密码子发生无义突变。截短的E7蛋白在蛋白质免疫印迹法中可被标准E7单克隆抗体识别，并在NIH3T3细胞中表达。在通过表达变异E7基因的质粒DNA肌肉注射免疫的小鼠血清中，接种后第2、3、5和6周可检测到特异性E7抗体。然而，与E7蛋白原型相比，通过MTT比色法在体外未检测到E7蛋白刺激后的特异性淋巴细胞增殖。
HPV16 E7基因在中国可能会出现变异，变异蛋白能够表达并诱导宿主产生体液免疫反应，但不能引发针对它的特异性细胞免疫反应。这些数据可能是中国湖北省未来开发HPV16疫苗的关键所在。
