A new method for the study of biophysical and morphological parameters in 3D cell cultures: Evaluation in LoVo spheroids treated with crizotinib

Abstract

Three-dimensional (3D) culture systems like tumor spheroids represent useful in vitro models for drug screening and more broadly for cancer biology research, but the generation of uniform populations of spheroids remains challenging. The possibility to properly characterize spheroid properties would increase the reliability of these models. To address this issue different analysis were combined: i) a new device and relative analytical method for the accurate, simultaneous, and rapid measurement of mass density, weight, and size of spheroids, ii) confocal imaging, and iii) protein quantification, in a clinically relevant 3D model. The LoVo colon cancer cell line forming spheroids, treated with crizotinib (CZB) an ATP-competitive small-molecule inhibitor of the receptor tyrosine kinases, was employed to study and assess the correlation between biophysical and morphological parameters in both live and fixed cells. The new fluidic-based measurements allowed a robust phenotypical characterization of the spheroids structure, offering insights on the spheroids bulk and an accurate measurement of the tumor density. This analysis helps overcome the technical limits of the imaging that hardly penetrates the thickness of 3D structures. Accordingly, we were able to document that CZB treatment has an impact on mass density, which represents a key marker characterizing cancer cell treatment. Spheroid culture is the ultimate technology in drug discovery and the adoption of such precise measurement of the tumor characteristics can represent a key step forward for the accurate testing of treatment’s potential in 3D in vitro models.

Sonoprinting nanoparticles on cellular spheroids via surface acoustic waves for enhanced nanotherapeutics delivery

“…The size of both spheroids was within the peak Bjerknes forces in the device. Furthermore, as a common indicator of the cell–cell compactness in spheroids, 48 the optical density measurement revealed a higher density in YUMM 1.7 (Fig. 2A).…”

Sonoprinting Nanoparticles on Cellular Spheroids via Surface Acoustic Waves for Enhanced Nanotherapeutics Delivery

“…The size of both spheroids was within the peak Bjerknes forces in the device. Furthermore, as a common indicator of cell-cell compactness in spheroids, 40 the optical density measurement revealed a higher density in YUMM 1.7 (Figure 2A).…”

Adipose Stromal Cell Spheroids for Cartilage Repair: A Promising Tool for Unveiling the Critical Maturation Point

“…By confocal microscopy, we analyzed the 3D cell architecture of spheroids cultured for 21 and 28 days on-chip or in ULA plates. We imaged cleared spheroids combined with a silicone immersion oil objective suitable for deep imaging analysis as previously described [27]. The cultured spheroids on-chip appeared smaller in size (as shown in Figures 3A,B) after 21 and 28 days, which could be due to the small size of the culture wells.…”

“…Mass density profoundly affects the cellular arrangement within the spheroid, the establishment of nutrient gradients, and the efficacy of cell-cell and cellextracellular matrix interactions [25]. Mass density and its correlation with size have been previously established as pivotal factors influencing various biological phenomena, such as cell permeation [26] and drug activity [27]. In this light, the comprehension of the complex relationship between mass density and dimension could be essential for understanding their role in the growth and differentiation of spheroids.…”