Periodic Reporting for period 4 - STARNEL (supracellular contractility of myofibroblasts in gut homeostasis and cancer invasion)
Reporting period: 2023-04-01 to 2024-09-30
While the role of chemical cues in the intestinal stem cell niche and tumor progression is well established, how stromal cells—such as fibroblasts and cancer-associated fibroblasts (CAFs)—use mechanical forces to remodel the extracellular matrix (ECM) and influence epithelial behavior remains poorly understood. This gap in knowledge, especially at the single-cell level, is largely due to imaging limitations. In this project, we employed a multidisciplinary approach at the interface of cell biology and physics to investigate how fibroblast contractility regulates epithelial cell function in both homeostatic and cancerous contexts, using the gut as a model system.
We discovered that CAFs form a contractile capsule around tumors that actively compresses cancer cells via actomyosin-driven forces, rather than serving as a passive barrier. Disruption of CAF contractility reduces compression and impairs capsule formation, revealing the intrinsic, supracellular nature of force generation coordinated by fibronectin scaffolds (Barbazan et al., Nat Commun, 2023). CAF-induced compression suppresses cancer cell proliferation and may enhance chemoresistance.
To study how the CAF capsule forms, we co-cultured CAFs and tumor cells in elastic alginate shells. We found that compressive stress and fibronectin remodeling are required for CAFs to spread and encapsulate tumor aggregates (Bertillot et al., Commun Biol, 2024). Moreover, we showed that CAF alignment drives the formation of a nematic fibronectin network, which, in turn, guides further CAF organization. Over time, this feedback loop leads to a slowdown in network dynamics and creates local defects—potential weak points in the CAF capsule that may facilitate cancer cell escape (Jacques et al., BioRxiv, 2024).
We further demonstrated that cancer cell clusters, even without intrinsic polarity, can migrate persistently by generating self-induced collagen gradients and mechanical asymmetries. This reveals a novel, polarity-independent mechanism for directed migration on viscoelastic substrates (Clark et al., Nat Mater, 2022).
In the colon, we unexpectedly observed that a distinct population of macrophages extends "balloon-like" protrusions (BLPs) into the epithelial layer. Given the colon’s critical role in absorbing fluids and its exposure to microbial and fungal metabolites, we investigated the function of these structures. We found that BLPs sample absorbed fluids and block uptake when fungal toxins are detected. In the absence of these macrophages or BLPs, epithelial cells continue absorbing toxic fluids, leading to cell death and loss of barrier integrity. These findings reveal an essential, previously unrecognized role of macrophages in maintaining colonic homeostasis (Chikina et al., Cell, 2020).
We also explored how intestinal stem cells respond to mechanical cues in their niche. We showed that PIEZO ion channels act as mechanosensors required for stem cells state regulation. Activated by increased ECM stiffness and tissue tension, PIEZO channels influence stem cell maintenance and fate decisions through a defined mechanotransduction cascade (Baghdadi et al., Science, 2024).
To study how the CAF capsule forms, we co-cultured CAFs and tumor cells in elastic alginate shells. We found that compressive stress and fibronectin remodeling are required for CAFs to spread and encapsulate tumor aggregates (Bertillot et al., Commun Biol, 2024). Moreover, we showed that CAF alignment drives the formation of a nematic fibronectin network, which, in turn, guides further CAF organization. Over time, this feedback loop leads to a slowdown in network dynamics and creates local defects—potential weak points in the CAF capsule that may facilitate cancer cell escape (Jacques et al., BioRxiv, 2024).
We further demonstrated that cancer cell clusters, even without intrinsic polarity, can migrate persistently by generating self-induced collagen gradients and mechanical asymmetries. This reveals a novel, polarity-independent mechanism for directed migration on viscoelastic substrates (Clark et al., Nat Mater, 2022).
In the colon, we unexpectedly observed that a distinct population of macrophages extends "balloon-like" protrusions (BLPs) into the epithelial layer. Given the colon’s critical role in absorbing fluids and its exposure to microbial and fungal metabolites, we investigated the function of these structures. We found that BLPs sample absorbed fluids and block uptake when fungal toxins are detected. In the absence of these macrophages or BLPs, epithelial cells continue absorbing toxic fluids, leading to cell death and loss of barrier integrity. These findings reveal an essential, previously unrecognized role of macrophages in maintaining colonic homeostasis (Chikina et al., Cell, 2020).
We also explored how intestinal stem cells respond to mechanical cues in their niche. We showed that PIEZO ion channels act as mechanosensors required for stem cells state regulation. Activated by increased ECM stiffness and tissue tension, PIEZO channels influence stem cell maintenance and fate decisions through a defined mechanotransduction cascade (Baghdadi et al., Science, 2024).
To support these investigations, we developed two complementary model systems in collaboration with physicists: 2D and 3D gut-on-chip platforms. These models enabled us to map three-dimensional force patterns in intestinal epithelium and revealed how actomyosin-generated tension drives crypt folding, zonation, and coordinated epithelial migration (Perez-Gonzalez et al., Nat Cell Biol, 2021). A complementary gut-on-chip model replicating the stromal compartment showed that fibroblasts promote the correct spatial segregation of proliferative and differentiated cells (Verhulsel et al., Lab Chip, 2020).
Finally, we established a robust and rapid protocol for orthotopic tumor modeling in the colonic mucosa. This platform allows real-time monitoring of primary tumor growth and metastasis in vivo using colonoscopy or IVIS imaging, followed by tissue analysis for downstream applications (Richon et al., STAR Protoc, 2023).
Finally, we established a robust and rapid protocol for orthotopic tumor modeling in the colonic mucosa. This platform allows real-time monitoring of primary tumor growth and metastasis in vivo using colonoscopy or IVIS imaging, followed by tissue analysis for downstream applications (Richon et al., STAR Protoc, 2023).