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Cells in tissues navigate fibrous environments fundamentally differently than they do on flat substrates, but the establishment of cell forces in physiological fibrous settings remains poorly understood. Although factors such as the stiffness of the extracellular matrix (ECM) are known to drive behaviors, including cell motility on flat nonfibrous substrates, the interplay between fiber architecture and stiffness in fibrous ECM is not known. Here, we find that in fibrous environments, the directionality of mechanical forces overrides ECM stiffness as the primary regulator of contractility in migrating cells. Using an approach combining phase microscopy with deep learning to map forces in real time, termed deep learning-enabled live-cell fiber-force microscopy (DLFM), we reveal that when cells transition between anisotropic and isotropic stress fields, their contractility significantly drops despite encountering stiffer ECM, contrary to the behavior of cells on flat nonfibrous substrates. Unlike the peripheral adhesions observed on flat nonfibrous substrates, cells in fibrous matrices form force-generating adhesions throughout their body, stabilized by out-of-plane mechanical components unique to fiber geometry. Cells exhibit distinct force signatures during migration, division, and differentiation, with temporal signatures that predict stem cell fate. These findings, enabled by combining deep learning and the mechanics of cells and fibers, explain long-standing paradoxical behavior of cells navigating deformable fibrous environments, how they can pull and tug at them, and identify tension anisotropy as a master regulator of cell behavior, with implications for cancer invasion, tissue engineering, and regenerative medicine.
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This content will become publicly available on December 1, 2026
Combinatorial extracellular matrix tissue chips for optimizing mesenchymal stromal cell microenvironment and manufacturing
Abstract Despite the therapeutic potential of mesenchymal stromal cells (MSC), there is limited understanding of optimal extracellular matrix (ECM) environments to manufacture these cells. We developed tissue chips to study the effects of multi-factorial ECM environments under manufacturable stiffness ranges and multi-component ECM compositions. Manufacturing qualities of cell expansion potential, immunomodulation, and differentiation capacity were examined. The results show stiffness effects, with 900 kPa substrates supporting higher proliferation and osteogenic differentiation, along with anti-inflammatory IL-10 expression, whereas 150 kPa substrates promoted adipogenic differentiation at 150 kPa, suggesting that optimal ECM environments may differ based on manufacturing goals. ECM biochemistries containing fibronectin and laminin further modulated MSC manufacturing qualities across various stiffnesses. Proteomic and transcriptomic analyses revealed unique ECM combinations that induced higher levels of angiogenic and immunomodulatory cytokines, compared to single factor ECMs. These findings demonstrate that optimized ECM environments enhance MSC manufacturing quality.
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- PAR ID:
- 10651181
- Publisher / Repository:
- Springer Nature
- Date Published:
- Journal Name:
- npj Regenerative Medicine
- Volume:
- 10
- Issue:
- 1
- ISSN:
- 2057-3995
- Page Range / eLocation ID:
- 21
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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