Human malignant pleura mesothelioma ‐ a three‐dimensional (3D) tumor model

Abstract

Human malignant pleura mesothelioma (MPM) is an extremely aggressive tumor of the mesothelium lining pleural cavity. Once diagnosed, it is uniformly fatal, with a median survival ranging from 4 to 12 month. The aim of our study is to design a 3D MPM model, to get knowledge of the biologic processes, which promote and maintain the uncontrolled proliferation of MPM cells, because the use of monolayer (2D) cultures does not accurately reflect the behaviour of cells in vivo. Human anchorage-depended MPM cells were seeded with macroporous gelatine microcarriers in the NASA-developed Rotary Cell Culture System™ (RCCS™). By simulating weightlessness (10−2 g) cells are maintained in a dynamic fluid suspension. Cells were examined at the state of viability, growth, morphology, differentiation and protein level in comparison to 2D controls. Different tumor marker were evaluated by immunohistochemistry, and the secretion of mucoglycoproteins by Alcian Blue staining. In addition, a comparative proteomic profile was established by 2D gel elektrophoreses and MALDI-TOF MS. Morphology, cell-cell junctional complexes and the well-developed formation of microvilli was shown by electron microscopy. The resulting morphological and functional differentiated tumor spheroids (5 mm in average) displayed a distinct protein pattern, than the control. Interestingly, the MPM marker Calretinin and Vimentin are expressed in a different manner. The organized 3D arrangement of mesothelioma cells with different phenotypes such as proliferating, non-proliferating, necrotic cells and extracellular matrix-interactions, represents the histological complexity of MPM tissue in-vivo. In conclusion, we find that this 3D in vitro culture is a credible and reliable preclinical tumor model.