Data for: Rigidity sensing of inclusions directs differentiated cell elongation and force generation across phenotypes
Abstract
Fibrosis is driven in part by the transition of healthy fibroblasts to a contractile phenotype, called myofibroblasts. The mechanics of the extracellular matrix play a crucial role in regulating the cell fates and behaviors during this transition. However, most studies to date focus on cells grown on 2D surfaces and matrices with homogeneous properties. This leaves open how local rigidity differentially regulates the behaviors of both phenotypes in 3D environments, including polarization, contraction, and maintenance of phenotypes, during remodeling. Here, we engineer 3D microgel-in-collagen composites by embedding low-volume fractions of cell-scale microgels with two levels of rigidity, mimicking healthy and pathological tissues, that are stiffer than the surrounding collagen but do not significantly change the bulk modulus. We find that microgels serve as mechanical centers: both phenotypes polarize toward microgel inclusions. The polarization response decays as a power-law with distance ∼ r−n, decreasing more slowly for myofibroblasts (n ≈ 0.35) than fibroblasts (n ≈ 0.81), indicating that myofibroblasts are more sensitive to small mechanical variations. In-situ measurement finds that forces are highest for myofibroblasts near stiff microgels and lowest for fibroblasts near soft microgels. Local rigidity also stabilizes the myofibroblast phenotype: Both the ordering of the proinflammatory marker α-SMA and nuclear YAP localization persist for cells cultured with stiff microgels over several days, but diminish quickly for those cultured with soft microgels and in pure collagen. Together, these results reveal a rigidity- and phenotype-dependent feedback loop: stiff inclusions induce cell polarization and collagen remodeling via contractile force, which in turn, maintain the myofibroblast phenotype. Our study positions mechanical heterogeneity as a useful and sensitive handle to probe and potentially modulate early fibrotic progressions.
Overview
This repository (Dryad.zip) contains all data, scripts, and analyses supporting the results presented in the associated publication: 'Rigidity sensing of inclusions directs differentiated cell elongation and force generation across phenotypes' by Yuxin Luo and Yimin Luo. Each figure has a corresponding folder, which includes raw data, processed data, and analysis scripts used to generate the figure panels.
General Notes
Dataset Organization
The dataset is organized by figure, with each folder named according to its corresponding figure (e.g., Figure_3, Figure_4, etc.). Each figure folder has contents as follows:
Data Contents
- MATLAB Scripts: Primarily used for figure generation, statistical analysis, and visualization. These scripts are self-contained and reproduce the corresponding figure panels when run. They are prefixed with the figure number (e.g.,
Figure_7b.m,Figure_7d.m). - Processed Data (CSV files): Cleaned and prepared datasets for analysis and visualization are included directly alongside corresponding scripts. These files contain quantitative measurements such as rheology values, collagen contraction areas, anisotropy indices (DOA), and YAP ratios.
- Raw Data: Included where applicable, such as cell tracking trajectories from TrackMate, or Trios files from the rheometer.
Experimental Conditions
Descriptions of setups of experiemts such as bulk rheometry are briefly described in figure-specific README files.
Overview of Figures
Figure 3
- Content: Frequency sweep rheology of storage modulus (G′) for bulk PEGDA gels and collagen composites.
Figure 4
- Content: Polarization of fibroblasts and myofibroblasts relative to microgels, quantified by angle distribution and distance-dependent correlation.
Figure 5
- Content: Collagen contraction assays in 96-well plates, quantified by normalized pellet area over 72 hours.
Figure 6
- Content: Tracer bead displacement analysis for quantifying forces exerted by fibroblasts and myofibroblasts near stiff or soft microgels.
Figure 7
- Content: Examining the extent of myofibroblast activation through analyzing degree of anisotropy (DOA) of alpha-SMA staining and nuclear-to-cytoplasm YAP staining.
Figure S2
- Content: Joint analysis of cell alignment and spatial proximity to microgels. Includes polarization angle–distance datasets, power-law decay of ⟨cos(2θ)⟩, boxplots of alignment factor distributions, cell-count histograms per radial bin, and polar heatmaps visualizing the spatial distribution of alignment around microgels.
Figure S3
- Content: Local collagen fiber alignment relative to the cell–microgel axis. OrientationJ hue maps are combined with cell outlines and microgel centroids to compute fiber deviation Δθ, rectangular masked regions along the cell–microgel direction, and corresponding histograms and cos(2Δθ) maps quantifying fiber reorientation influenced by cell–microgel interactions.
Figure S4
- Content: Frequency sweep rheometry of the storage (G’) moduli measured for bulk pure collagen and composite with microgels of different stiffness after decellularizing the matrix.
Figure S5
- Content: Representative polar plots for trajectories of fibroblasts and myofibroblasts near microgels, and mean speed comparison across conditions.
Figure S6
- Content: Histograms and volume fraction analysis of PEGDA microgel size distributions.
Citation
If you use this dataset in your work, we request that you cite the corresponding publication:
Title: Rigidity sensing of inclusions directs differentiated cell elongation and force generation across phenotypes
Journal: ACS Biomaterials Science & Engineering
Contact
For questions regarding the data or scripts:
- Refer to the figure-specific README files for detailed information.
- Contact information is provided in the associated publication.