Mapping Tumor Spheroid Mechanics in Dependence of 3D Microenvironment Stiffness and Degradability by Brillouin Microscopy

Abstract

Altered biophysical properties of cancer cells and of their microenvironment contribute to cancer progression. While the relationship between microenvironmental stiffness and cancer cell mechanical properties and responses has been previously studied using two-dimensional (2D) systems, much less is known about it in a physiologically more relevant 3D context and in particular for multicellular systems. To investigate the influence of microenvironment stiffness on tumor spheroid mechanics, we first generated MCF-7 tumor spheroids within matrix metalloproteinase (MMP)-degradable 3D polyethylene glycol (PEG)-heparin hydrogels, where spheroids showed reduced growth in stiffer hydrogels. We then quantitatively mapped the mechanical properties of tumor spheroids in situ using Brillouin microscopy. Maps acquired for tumor spheroids grown within stiff hydrogels showed elevated Brillouin frequency shifts (hence increased longitudinal elastic moduli) with increasing hydrogel stiffness. Maps furthermore revealed spatial variations of the mechanical properties across the spheroids’ cross-sections. When hydrogel degradability was blocked, comparable Brillouin frequency shifts of the MCF-7 spheroids were found in both compliant and stiff hydrogels, along with similar levels of growth-induced compressive stress. Under low compressive stress, single cells or free multicellular aggregates showed consistently lower Brillouin frequency shifts compared to spheroids growing within hydrogels. Thus, the spheroids’ mechanical properties were modulated by matrix stiffness and degradability as well as multicellularity, and also to the associated level of compressive stress felt by tumor spheroids. Spheroids generated from a panel of invasive breast, prostate and pancreatic cancer cell lines within degradable stiff hydrogels, showed higher Brillouin frequency shifts and less cell invasion compared to those in compliant hydrogels. Taken together, our findings contribute to a better understanding of the interplay between cancer cells and microenvironment mechanics and degradability, which is relevant to better understand cancer progression.

Correlation of biomechanics and cancer cell phenotype by combined Brillouin and Raman spectroscopy of U87-MG glioblastoma cells

“…Comparing the Brillouin shifts of adherent cells and cell spheroids of the same cell line U87-MG revealed significantly higher values for the latter ones. The Brillouin shift difference of about 0.1 GHz is in line with previous reported values of single cells and spheroids of breast cancer cells (MCF-7) (28). Line scans through spheroids showed local variations of the Brillouin shift.…”

“…We attributed the difference to the culturing conditions. This finding demonstrates that the choice of brain tumor model (adherent cells vs. cell spheroids) is highly relevant for analysis of biomechanics and in agreement with other studies on breast cancer spheroids and single cells in hydrogels (28).…”

“…Moreover, it was shown that the osmolality of surrounding medium affects the biomechanics of ovarian cancer cell spheroids (22). Furthermore, the effect of micro-environmental stiffness and degradability of hydrogels on breast cancer spheroids was demonstrated (28).…”

Magnetic Compression of Tumor Spheroids Increases Cell Proliferation In Vitro and Cancer Progression In Vivo

“…The impact of mechanical stress on cancer progression has been recently explored in vitro for cancer cell suspensions [ 9 ] and tridimensional tumor models [ 10 , 11 , 12 ] subjected to mechanical compressions, with stresses ranging from 50 Pa to a few kPa, corresponding to the ones experienced in vivo during tumor growth. Isotropic or anisotropic confinements of small growing spheroids (less than 100 µm in diameter) led to a limitation of the final spheroid volume and were associated with an increase in cell apoptosis and a decrease in cell proliferation toward the center of the confined spheroids [ 13 , 14 ], the latter of which can be related to an inner pressure rise [ 15 , 16 ].…”

Longitudinal shear wave elasticity measurements of millimeter-sized biomaterials using a single-element transducer platform

“…Temporal measurements of the elasticity in a 3D culture or an engineering tissue reveal dynamic cell–ECM interactions. They are usually best performed using noncontact imaging-based methods, including Brillouin microscopy [ 11 , 12 ], magnetic resonance elastography (MRE), and ultrasound shear wave (SW) elastography [ 13 – 15 ]. Confocal Brillouin microscopy probes the sample’s bulk modulus by detecting frequency shifts of the transmitted laser light caused by thermally generated acoustic phonons.…”