Lung mesenchymal cells function as an inductive microenvironment for human lung cancer propagating cells†

Abstract

The aim of the present study was to characterize the biological properties and in vivo tumourigenic potential of mesenchymal cells (MCs) obtained from non-small-cell lung cancer (NSCLC) samples.NSCLC samples (53 adenocarcinomas and 24 squamous-cell carcinomas) surgically removed from 46 males and 31 females were processed to identify mesenchymal cells from human lung cancer (hLc-MCs). hLc-MCs were separated from neoplastic epithelial cells, expanded and extensively characterized in vitro. Subsequently, female BALB/c nude mice were subcutaneously injected with either 10(6) or 2.5 × 10(6) Calu-3 (human adenocarcinoma cell line able to reproducibly induce xenografted tumours) alone or in combination with equal doses of hLc-MCs. Control animals were injected with the two doses of hLc-MCs only.Primary cultures of hLc-MCs were obtained from >80% of NSCLC specimens. The typical MCs immunophenotype was documented by the expression of CD90, CD105, CD73, CD13 and CD44 at fluorescence-activated cell sorting analysis. CD45, CD14, CD34 and epithelial antigens were negative while CD117 (c-kit) and CD133 (prominin) were partially expressed. Interestingly, nuclear transcription factors octamer-binding transcription factor 3/4 and sex determining region Y-box 2 involved in stemness, thyroid transcription factor 1 in bronchoalveolar commitment, and ETS1 in carcinogenesis, were expressed in hLc-MCs isolated from NSCLC. Specific conditioned media and cocultures confirmed the supportive role of hLc-MCs for cancer cells. In vivo experiments showed that at both doses Calu-3 xenografts doubled in size when hLc-MCs were coinjected. Cell tracking in xenografted tumours, by immunofluorescence combined with fluorescence in situ hybridization analysis, documented hX-chromosome-labelled, Calu-3-derived cytokeratin-positive adenocarcinoma structures surrounded by hLc-MCs.Tumour-propagating cells require the inductive interaction of resident mesenchymal cells to foster lung cancer development.