J3 - Intercellular flow dorminates the poroelasticity of the multicellular tissues

Introduction: : Multicellular tissues and organs consist of cells, extracellular matrices (ECM), and substantial amounts of interstitial fluid, collectively determining their mechanical characteristics, such as elasticity, viscosity, and surface tension. These mechanical properties are critically linked to physiological functions and pathological states including fibrosis, tumor metastasis, and tissue injury response. Traditionally, due to their observed long-term adaptive shape changes, multicellular tissues have been predominantly modeled as viscoelastic materials. However, rapid mechanical responses in biological processes such as heartbeats, muscle contractions, and acute injuries suggest that tissue mechanics at shorter timescales involve mechanisms beyond simple viscoelasticity. Recent evidence indicates that interstitial fluid flow, mediated by intercellular spaces within tissues, might significantly influence mechanical behavior under rapid deformation. Nonetheless, the role and underlying mechanics of interstitial fluid flow in multicellular systems remain poorly understood, largely due to the lack of suitable experimental platforms capable of simultaneously applying controlled bulk deformation and measuring mechanical responses in three-dimensional (3D) multicellular systems.

Materials and

Methods: : To address this gap, we developed a tailored micro-mechanics platform integrating parallel plate compression and optical imaging to directly measure the mechanical responses of multicellular tissues. The platform enabled precise deformation of cultured 3D cell spheroids (e.g., A549, MCF10A, MDA-MB-231, HeLa) and native mouse pancreatic islets, capturing real-time force relaxation data under controlled mechanical stimuli. Intercellular fluid flow was visualized using a cell-impermeable fluorescent tracer (FD4), while structural changes in intercellular spaces were examined using confocal microscopy with specific stains for cell nuclei and plasma membranes. We further manipulated the internal structure of these multicellular tissues chemically, through gap junction inhibition (carbenoxolone treatment), cytoskeletal disruption (cytochalasin D), ECM degradation (collagenase treatment), and mechanically through osmotic shocks (hypoosmotic and hyperosmotic conditions). The resulting mechanical properties and structural dynamics were quantified and compared across different experimental conditions.

Results, Conclusions, and Discussions:: Our results reveal that both cultured cell spheroids and native pancreatic islets display distinct poroelastic behavior at timescales of up to one minute, indicated by a characteristic size-dependent force relaxation. Contrary to classical viscoelastic predictions, larger tissue sizes exhibited longer force relaxation times, clearly implicating fluid transport within intercellular spaces as the dominant mechanism governing immediate mechanical responses. Direct visualization of interstitial fluid influx and efflux validated the existence and functional relevance of intercellular spaces for fluid transport. Furthermore, structural modulations that altered intercellular spaces or skeleton stiffness—via gap junction inhibition, ECM degradation, cytoskeletal disruption, or osmotic compression—systematically influenced poroelastic responses. Larger intercellular spaces or stiffer multicellular skeletons facilitated faster interstitial fluid flow and quicker force relaxation.

These findings clarify the critical yet previously underappreciated role of interstitial fluid transport in determining the short-timescale mechanical behavior of multicellular systems. Importantly, this poroelastic mechanism is directly linked to physiological and pathological conditions, including diabetes-induced tissue stiffening and aging-related structural degradation, highlighting potential mechanical biomarkers for disease diagnosis. Overall, our micro-mechanics platform provides valuable insights into the fundamental physics of tissue mechanics and offers new opportunities for exploring disease mechanisms and designing therapeutic strategies by precisely characterizing tissue poroelasticity.

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