Discriminating expression of differentiation markers evolves in transplants of benign and malignant human skin keratinocytes through stromal interactions

Accumulating evidence indicates a decisive role for the adjacent stroma in tumour growth and dissemination. However, it is not clear how far altered differentiation such as expression of aberrant keratins and vimentin, common in invasive human carcinomas, may reflect intrinsic cell properties or a response to the tumour environment. We have addressed this by transplanting benign and malignant human HaCaT-ras keratinocytes, seeded on collagen matrix, onto nude mice. Initially, epithelia derived from benign and malignant cells, being separated from host stroma by collagen, were poorly organized and exhibited the same differentiation markers, as identified by immunofluorescence and in situ hybridization. Epidermal basal and suprabasal keratins were expressed persistently even upon contact with newly formed stroma and malignant cell invasion. In contrast, non-epidermal keratins (K4/K13, K8/18, K19), which were similarly synthesized by benign and malignant cells in culture and in early transplants, were differentially regulated with increasing stromal vicinity. While both proteins and mRNAs were downregulated in benign epithelia, the malignant, invasive tumour cells continuously expressed these non-epidermal keratins throughout (K19), suprabasally (K4/13) or at invasive sites (K8/18). Furthermore, the mesenchymal protein vimentin was expressed de novo in invasive areas confronting tumour stroma. Thus, atypical tissue markers, similarly synthesized in isolated cells in vitro, are downregulated in benign but maintained and upregulated in malignant epithelia. This is presumably caused by the neighbouring stroma being permanently activated by malignant epithelia.