Toussaint-Smith Esra, Donner David B, Roman Ann
Department of Microbiology and Immunology and Walther Oncology Center, Indiana University School of Medicine and Walther Cancer Institute, IN 46202-5120, USA.
Oncogene. 2004 Apr 15;23(17):2988-95. doi: 10.1038/sj.onc.1207442.
Human papillomavirus (HPV) 16 is involved in causing cervical cancer. The E6 and E7 proteins of HPV 16 immortalize human keratinocytes and this is due, at least in part, to inactivation of the tumor suppressor proteins p53 and pRB. These tumor suppressor proteins also regulate the expression of pro- and antiangiogenic factors by cells. For this reason, experiments were conducted to determine whether the expression of E6 and E7 in primary keratinocytes alters the phenotype of these cells such that they express diminished levels of antiangiogenic factors and/or increased levels of proangiogenic factors. To avoid variances in experimental observations, pools of human foreskin keratinocytes from multiple sources were infected with recombinant retrovirus expressing HPV 16 E6 and E7 or control retrovirus. Gene array analysis, RT-PCR, ELISAs and Western blotting showed that in cells expressing HPV 16 E6 and E7, expression levels of two potent angiogenesis inhibitors, thrombospondin-1 and maspin, were lower compared to controls. Additionally, major angiogenesis inducers, interleukin-8 and vascular endothelial growth factor (VEGF), were increased relative to controls. VEGF can be produced as multiple splice variants, all of which are required for the formation of patent blood vessels. The expression of HPV 16 E6 and E7 in keratinocytes augmented expression of VEGF 121, 145, 165 and 189. These observations show that HPV 16 E6 and E7 alter the phenotype of primary keratinocytes, diminishing expression of inhibitors and increasing expression of inducers of angiogenesis. This altered phenotype may permit keratinocytes infected by HPV 16 to play a role in the progression of cancer by permitting tumors to acquire a blood supply permissive of growth and spread.
人乳头瘤病毒（HPV）16型与宫颈癌的发生有关。HPV 16型的E6和E7蛋白可使人类角质形成细胞永生化，这至少部分归因于肿瘤抑制蛋白p53和pRB的失活。这些肿瘤抑制蛋白还调节细胞促血管生成因子和抗血管生成因子的表达。因此，开展了实验以确定E6和E7在原代角质形成细胞中的表达是否会改变这些细胞的表型，使其抗血管生成因子表达水平降低和/或促血管生成因子表达水平升高。为避免实验观察结果出现差异，用表达HPV 16 E6和E7的重组逆转录病毒或对照逆转录病毒感染来自多个来源的人包皮角质形成细胞池。基因阵列分析、逆转录聚合酶链反应（RT-PCR）、酶联免疫吸附测定（ELISA）和蛋白质免疫印迹法显示，在表达HPV 16 E6和E7的细胞中，两种强效血管生成抑制剂血小板反应蛋白-1和组织蛋白酶抑制剂maspin的表达水平低于对照。此外，主要的血管生成诱导因子白细胞介素-8和血管内皮生长因子（VEGF）相对于对照有所增加。VEGF可产生多种剪接变体，所有这些变体都是形成有功能的血管所必需的。角质形成细胞中HPV 16 E6和E7的表达增加了VEGF 121、145、165和189的表达。这些观察结果表明，HPV 16 E6和E7改变了原代角质形成细胞的表型，减少了血管生成抑制剂的表达并增加了血管生成诱导因子的表达。这种改变的表型可能使感染HPV 16的角质形成细胞通过使肿瘤获得允许生长和扩散的血液供应，在癌症进展中发挥作用。
