Bosch F X, Dürst M, Schwarz E, Boukamp P, Fusenig N E, zur Hausen H
Hals-Nasen-Ohren-Klinik der Universität Heidelberg, Germany.
Behring Inst Mitt. 1991 Jul(89):108-21.
We have transplanted HPV 18 positive nontumorigenic hybrid cells and tumorigenic hybrid segregants as well as nontumorigenic and tumorigenic human keratinocytes immortalized by HPV 16-DNA transfection (HPK cells and HPK-ras cells) into nude mice and grown them under in vivo conditions for different periods of time. By analyzing gene expression at the mRNA level of the early viral genes E6/E7 and a number of cytoskeletal cellular genes we wanted to test the hypothesis that the nontumorigenic phenotype of these cells in vivo may be determined by the specific downregulation of expression of the oncogenic E6/E7 genes by the product(s) of tumor suppressor gene(s). The results obtained showed that: 1) The nontumorigenic hybrid cells (in contrast to tumorigenic segregants) stopped to proliferate about 3 days after transplantation. At this time E6/E7 gene expression was already drastically reduced, whereas at day 2 expression was still high. This suppression specifically affected the HPV 18 E6/E7 genes and preceded cell death by at least 10 days. 2) The same specific suppression of HPV E6/E7 gene expression occurred during in vivo growth of the nontumorigenic HPK cells. These cells gave rise to transiently growing cysts, but had retained the in vivo differentiation potential of normal keratinocytes (orderly expression of cytokeratins 1 and 10, involucrin and filaggrin). HPV 16 E6/E7 gene expression was very low and clearly restricted to a small subset of basal cells, thus showing an inverse relationship to terminal differentiation. In contrast, in the tumors induced by HPK-ras cells E6/E7 gene expression extended from the basal cells into suprabasal cells and terminal differentiation was retarded. These results support the hypothesis that during in vivo growth tumor suppressor gene(s) negatively regulated HPV E6/E7 gene expression resulting in the stop of proliferation of the nontumorigenic cells.
我们已将人乳头瘤病毒18型（HPV 18）阳性的非致瘤性杂交细胞、致瘤性杂交分离株以及通过HPV 16 - DNA转染永生化的非致瘤性和致瘤性人角质形成细胞（HPK细胞和HPK - ras细胞）移植到裸鼠体内，并在体内条件下培养不同时间。通过分析早期病毒基因E6/E7以及一些细胞骨架细胞基因在mRNA水平的基因表达，我们想要验证一个假说，即这些细胞在体内的非致瘤表型可能由肿瘤抑制基因产物对致癌性E6/E7基因表达的特异性下调所决定。所得结果表明：1）非致瘤性杂交细胞（与致瘤性分离株不同）在移植后约3天停止增殖。此时E6/E7基因表达已大幅降低，而在第2天时表达仍很高。这种抑制特异性地影响了HPV 18 E6/E7基因，并且至少在细胞死亡前10天就已出现。2）在非致瘤性HPK细胞的体内生长过程中，同样出现了对HPV E6/E7基因表达的特异性抑制。这些细胞形成了短暂生长的囊肿，但保留了正常角质形成细胞在体内的分化潜能（细胞角蛋白1和10、内披蛋白和丝聚蛋白的有序表达）。HPV 16 E6/E7基因表达非常低，且明显局限于一小部分基底细胞，因此与终末分化呈负相关。相比之下，在HPK - ras细胞诱导的肿瘤中，E6/E7基因表达从基底细胞扩展到基底上层细胞，终末分化受到抑制。这些结果支持了这样一个假说，即在体内生长过程中，肿瘤抑制基因对HPV E6/E7基因表达产生负调控，导致非致瘤性细胞停止增殖。
