Chamulitrat Walee, Schmidt Rainer, Chunglok Warangkana, Kohl Annette, Tomakidi Pascal
Deutsches Krebsforschungszentrum, Department of Applied Tumorvirology, Heidelberg, Germany.
Eur J Cell Biol. 2003 Jun;82(6):313-22. doi: 10.1078/0171-9335-00317.
Epithelial-mesenchymal transition (EMT) may be critical for neoplastic progression and its eventual tumorigenicity of epithelia. In this context, we investigated whether EMT and EMT-associated features occurred after chronic ethanol treatment of human gingival keratinocytes immortalized with the E6/E7 oncogenes of human papillomavirus (HPV) type 16. Following a nine-week treatment of cells with 30 mM ethanol in keratinocyte growth medium, they were cultured in normal DMEM with 10% serum. These cell populations were able to proliferate in this medium gradually exhibiting elongated morphology indicating that these cells underwent EMT. Control cells without ethanol treatment did not survive subcultures in DMEM. Upon long-term subcultures of ethanol-treated cells, two phenotypes were obtained exhibiting epithelium-like and spindle-shape fibroblast-like morphology (respectively, termed as EPI and FIB cells), the latter indicating EMT. In comparison to EPI cells, the phenotypic transition to FIB cells was concomitant with a decrease in the expression of keratins, desmoplakins and a complete loss of K14. Moreover, FIB cell transition strongly correlates with an increase in the expression of vimentin and simple epithelial keratin K18. These alterations in FIB cells were associated with the ability of these cells to exhibit anchorage-independent growth, while EPI cells exhibited anchorage-dependent growth. Concerning the transformation stage, FIB cells represent a progressively more advanced transformed phenotype which may reflect an early step during HPV- and ethanol-dependent multi-step carcinogenesis.
上皮-间质转化（EMT）可能对上皮细胞的肿瘤进展及其最终的致瘤性至关重要。在此背景下，我们研究了用16型人乳头瘤病毒（HPV）的E6/E7癌基因永生化的人牙龈角质形成细胞经慢性乙醇处理后是否发生EMT及与EMT相关的特征。在用角质形成细胞生长培养基中的30 mM乙醇处理细胞九周后，将它们培养在含10%血清的正常DMEM中。这些细胞群体能够在这种培养基中增殖，逐渐呈现出细长的形态，表明这些细胞经历了EMT。未经乙醇处理的对照细胞在DMEM中传代培养时无法存活。对经乙醇处理的细胞进行长期传代培养后，获得了两种表型，分别呈现上皮样和纺锤形成纤维细胞样形态（分别称为EPI细胞和FIB细胞），后者表明发生了EMT。与EPI细胞相比，向FIB细胞的表型转变伴随着角蛋白、桥粒斑蛋白表达的降低以及K14的完全丧失。此外，FIB细胞转变与波形蛋白和简单上皮角蛋白K18表达的增加密切相关。FIB细胞中的这些改变与这些细胞表现出的不依赖贴壁生长的能力相关，而EPI细胞表现出依赖贴壁生长。关于转化阶段，FIB细胞代表一种逐渐更高级的转化表型，这可能反映了HPV和乙醇依赖性多步骤致癌过程中的早期步骤。
