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.

Extracellular matrix stiffness activates mechanosensitive signals but limits breast cancer cell spheroid proliferation and invasion

“…Much of the previous work studying ERK and YAP activity in response to stiffness-and functional consequences therein-have been conducted in cells on 2D surfaces, often at non-physiological stiffnesses (>100 kPa), and which do not recapitulate the 3D environment in solid tumors. Our data agrees with other studies demonstrating that increased 3D collagen stiffness can suppress proliferation and invasion in other cancer types (Berger et al, 2017;Staneva et al, 2018;Berger et al, 2020;Mahajan et al, 2021).…”

The role of cell-matrix adhesion and cell migration in breast tumor growth and progression

“…Furthermore, the spheroids also appeared to spread away from the tumor mass, presenting a less compact structure following a 5-day culture within the 3D collagen matrix. Our observations align with Mahajan and colleagues' study, where increased stiffness of degradable hydrogel (15-20 kPa) similarly reduces the growth of MCF7 spheroids, accompanied by lower invasion into the matrix (Mahajan et al, 2021). Our further investigation revealed that changes in spheroid sizes and morphologies appeared to be regulated by cell growth, cell cycle arrest, apoptosis, and EMT when embedded in collagen matrices.…”

Exploiting Matrix Stiffness to Overcome Drug Resistance

“…For imaging purposes of TME, nonlinear imaging like multiphoton microscopy and second harmonic generation can be used, especially for live imaging of composition and architecture change of the TME. − As a noncontact method, Brillouin microscopy is used to map the stiffness of different biological samples in 2D and 3D. Since Brillouin microscopy is a noninvasive measurement technique, it can potentially be adapted for obtaining in vivo measurements of tissue mechanical properties with subcellular resolution. − Other noninvasive methods such as magnetic resonance elastography, ultrasonography, and optical coherence tomography , are also widely used in measuring the mechanical properties of both healthy normal and cancer tissues in vivo since they are based on elastography. One of the most commonly used stiffness measurement techniques for biomaterials or synthetic materials in biomedical research is shear rheometry.…”