Tumor Cell Plasticity and Angiogenesis in Human Melanomas

Daniela Mihic-Probst,Gianluca Civenni,Marianne Tinguely,Peter Schraml,Kristian Ikenberg,Reinhard Dummer,Burkhardt Seifert,Silvia Behnke,Holger Moch,Lukas Sommer

doi:10.1371/journal.pone.0033571

Abstract

Recent molecular studies provide evidence for a significant transcriptional plasticity of tumor cell subpopulations that facilitate an active contribution to tumor vasculature. This feature is accompanied by morphological changes both in vitro and in vivo. Herein, we investigated the morphological plasticity of tumor cells with special focus on vasculogenic mimicry and neovascularisation in human melanoma and mouse xenografts of human melanoma cell lines. In melanoma xenograft experiments, different vessel markers and green fluorescent protein expression were used to show how melanoma cells contribute to neovascularization. Additionally, we analyzed neovascularization in 49 primary melanomas and 175 melanoma metastases using immunostaining for blood (CD34) and lymphatic (D2–40) vessel-specific markers. We found significantly more lymphatic vessels in primary melanomas than in melanoma metastases (p<0.0001). In contrast to the near absence of lymphatic vessels within metastases, we found extensive blood micro-neovascularization. Blood micro-neovascularization was absent in micro metastases (less than 2 mm). A significant inverse correlation between Glut-1 expression (implying local hypoxia) and the presence of microvessels indicates their functional activity as blood vessels (p<0.0001). We suggest that the hypoxic microenvironment in metastases contributes to a phenotype switch allowing melanoma cells to physically contribute to blood vessel formation.

Highlights

Primary melanomas show a higher lymphatic vessel density than melanoma metastases We investigated 127 melanoma metastases (21 lymph nodes and 106 other sites) and 49 primary melanomas for vascularisation
15 of 127 (12%) metastases had lymphatic vessels whereas 39 of 49 (80%) primary melanomas were positive for lymphatic vessels
Several studies have shown that increased lymphatic density in primary melanomas is associated with a higher incidence of regional lymph node metastases [5,33]

Summary

Introduction

We have demonstrated two opposing gene expression profiles in melanoma short term cell cultures associated with proliferative or invasive cell phenotypes and provided evidence for phenotype switching favouring disease progression [12,13]. This transcriptional plasticity is accompanied by morphological changes in vitro, and involves many factors governing angiogenesis. In 1999, Maniotis and co-workers introduced the concept of vasculogenic mimicry (VM) in melanoma [14] He defined VM as melanoma cells with endothelial-like morphology embedded in a PAS and collagen IV positive extracellular matrix. Most recently, sophisticated investigations have shown that tumor cells directly participate in blood vessel formation [25,26]

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Conclusion