Single-Cell Quantification of Viscoelastic Phase Transitions in 3D Tissues

Abstract

Transitions of biological tissues between solid-like and liquid-like phases have been of great recent interest. Here, the first successful cell-by-cell evaluation of tissue viscoelastic transition is presented. An in situ micro-mechanical perturbation is applied to a microtissue, and the resulting volumetric deformation is evaluated using 3D light-sheet microscopy and digital image correlation (DIC), quantifying both solid-like, well-aligned displacement and liquid-like swirling motion between individual cells. The viscoelastic transition of fibroblasts is crucial in fundamental physiological events, such as placentation, cancer dissemination, and wound healing. This study investigates 3D organoid systems modeling maternal-fetal and tumor-stroma interfaces, demonstrating established molecular and structural parallels. The analysis visualizes individual cells in stromal-epithelial interactions and how they collectively alter tissue viscoelastic properties. It also enables in-silico microdissection, linking single-cell viscoelasticity with multi-channel fluorescence. RNAseq analysis of endometrial stromal fibroblasts shows that decidualization activates mechano-transcriptional regulators, including myocardin-related transcription factors (MRTFs), associated with increased cellular contractility and actomyosin mobilization. Knocking down MRTFA in cancer-associated fibroblasts in the tumor-fibroblast co-culture 3D model induces significant changes in fibroblast properties, mirroring those observed in the maternal-fetal interface model, highlighting parallels between placentation and cancer invasion. This analysis confirms existing beliefs and discovers new insights broadly applicable to studying organoids, embryos, tumors, and other tissues.