Background: Arterial thrombosis arises under high shear conditions and is a leading cause of ischemic stroke and myocardial infarction. The local hemodynamic environment during the formation of the thrombus influences its final composition, which in turn influences its mechanical characteristics and the associated risk of embolization. However, how arterial shear rates determine the local mechanical properties, structure, and composition of clots remains poorly understood. Objectives: This study investigated how physiological and pathologic arterial shear rates shape blood clot composition, formation dynamics, and regional mechanical properties in a microfluidic flow model. Methods: We used a microfluidic flow model that allows real-time imaging of blood clot formation under controlled arterial shear rates (300/s, 1000/s, and 3500/s). We visualized platelets and fibrin deposition using confocal microscopy and quantified composition-dependent spatial heterogeneities in clot stiffness with microindentation. Results: Pathologically high (3500/s) arterial shear rates promoted more rapid platelet aggregation and the formation of taller platelet-rich clots. Fibrin deposition started after ∼6 minutes in all shear conditions and coincided with platelet aggregate stabilization at higher shear rates. Microindentation revealed that platelet- and fibrin-rich regions were significantly stiffer than red blood cell–rich regions. Clots formed at 3500/s had a >3-fold higher average stiffness than those formed at 300/s, reflecting their higher platelet content. Conclusion: This study demonstrates that arterial shear rate governs clot growth and composition, translating into shear-dependent microscale mechanical properties. These findings provide new mechanistic insights into clot properties that are relevant for clot stability and vulnerability to embolization.

Collagen networks form the structural backbone of the extracellular matrix in both healthy and cancerous tissues, exhibiting nonlinear mechanical properties that crucially regulate tissue mechanics and cell behavior. Here, we investigate how the presence of invasive breast cancer cells (MDA-MB-231) influences the polymerization kinetics and mechanics of collagen networks using bulk shear rheology and rheo-confocal microscopy. We show that embedded cancer cells delay the onset of collagen polymerization due to volume exclusion effects. During polymerization, the cells (at 4% volume fraction) cause an unexpected time-dependent softening of the network. We show that this softening effect arises from active remodeling via adhesion and contractility rather than from proteolytic degradation. At higher cell volume fractions, the dominant effect of the cells shifts to volume exclusion, causing a two-fold reduction of network stiffness. Additionally, we demonstrate that cancer cells suppress the characteristic stress-stiffening response of collagen. This effect (partially) disappears when cell adhesion and contractility are inhibited, and it is absent when the cells are replaced by passive hydrogel particles. These findings provide new insights into how active inclusions modify the mechanics of fibrous networks, contributing to a better understanding of the role of cells in the mechanics of healthy and diseased tissues like invasive tumors. Statement of significance: Understanding how cells influence tissue mechanics is crucial to unravel disease progression. While fibroblasts are known to stiffen tissues, the role of invasive cancer cells is less clear. Using collagen-based tissue models, we reveal that cancer cells unexpectedly soften the collagen matrix and disrupt its stress-stiffening response. By comparing active cells to passive particles and selectively blocking cell functions, we show that volume exclusion, adhesion, and contractility each play distinct roles in shaping tissue mechanics. This work sheds light on the physical impact of cancer cells on their environment, advancing our understanding on how cells dynamically alter the mechanical properties of tissues.