Secretomes From Non-Small Cell Lung Cancer Cells Induce Endothelial Plasticity Through a Partial Endothelial-to-Mesenchymal Transition

Abstract

Aim

The tumor microenvironment (TME) of non-small cell lung cancer (NSCLC) is highly heterogeneous and is involved in tumorigenesis and resistance to therapy. Among the cells of the TME, endothelial cells are associated with the latter processes through endothelial-to-mesenchymal transition (EndMT). During EndMT, endothelial cells (ECs) progressively lose their endothelial phenotype in favor of a mesenchymal phenotype, which favors the production of cancer-associated fibroblasts (CAFs). Our study aimed to investigate the consequences of exposure to different lung tumor secretomes on EC phenotype and plasticity.

Materials and Methods

Conditioned media (CM) were prepared from the tumor cell lines A549, H1755, H23, H1437, and H1975. Proliferation and migration of ECs treated with these CMs were assessed by Cyquant and Incucyte technologies, respectively. The angiogenic capacity of ECs was assessed by following tubulogenesis on Matrigel. Phenotypic changes in treated ECs were detected by flow cytometry. Morphological analysis of actin fibers was performed by immunohistochemistry, while proteomic analysis by mass spectrometry was used to identify the protein content of secretomes.

Results

A change of the endothelial phenotype was found when human umbilical vein endothelial cells (HUVECs) were treated with different CMs. This phenotypic change was associated with a morphological change, an increase in both stress fiber expression and spontaneous migration. Furthermore, an increase in mesenchymal markers (α-SMA and CD44) confirmed the phenotypic changes. However, the secretomes did not modify the rate of double-labeled cells (vWF⁺/α-SMA⁺ or CD31⁺/CD44⁺). Proteomic analysis identified potential targets involved in the EndMT with therapeutic relevance.

Conclusion

Taken together, these data suggest that CMs can induce partial EndMT.