An official website of the United States government
The extracellular matrix provides macroscale structural support to tissues as well as microscale mechanical cues, like stiffness, to the resident cells. As those cues modulate gene expression, proliferation, differentiation, and motility, quantifying the stiffness that cells sense is crucial to understanding cell behavior. Whereas the macroscopic modulus of a collagen network can be measured in uniform extension or shear, quantifying the local stiffness sensed by a cell remains a challenge due to the inhomogeneous and nonlinear nature of the fiber network at the scale of the cell. To address this challenge, we designed an experimental method to measure the modulus of a network of collagen fibers at this scale. We used spherical particles of an active hydrogel (poly N-isopropylacrylamide) that contract when heated, thereby applying local forces to the collagen matrix and mimicking the contractile forces of a cell. After measuring the particles’ bulk modulus and contraction in networks of collagen fibers, we applied a nonlinear model for fibrous materials to compute the modulus of the local region surrounding each particle. We found the modulus at this length scale to be highly heterogeneous, with modulus varying by a factor of 3. In addition, at different values of applied strain, we observed both strain stiffening and strain softening, indicating nonlinearity of the collagen network. Thus, this experimental method quantifies local mechanical properties in a fibrous network at the scale of a cell, while also accounting for inherent nonlinearity.
more »
« less
Biocompatible micro tweezers for 3D hydrogel organoid array mechanical characterization
This study presents novel biocompatible Polydimethylsiloxane (PDMS)-based micromechanical tweezers (μTweezers) capable of the stiffness characterization and manipulation of hydrogel-based organoids. The system showed great potential for complementing established mechanical characterization methods such as Atomic Force Microscopy (AFM), parallel plate compression (PPC), and nanoindentation, while significantly reducing the volume of valuable hydrogels used for testing. We achieved a volume reduction of ~0.22 μl/sample using the μTweezers vs. ~157 μl/sample using the PPC, while targeting high-throughput measurement of widely adopted micro-mesoscale (a few hundred μm-1500 μm) 3D cell cultures. The μTweezers applied and measured nano-millinewton forces through cantilever’ deflection with high linearity and tunability for different applications; the assembly is compatible with typical inverted optical microscopes and fit on standard tissue culture Petri dishes, allowing mechanical compression characterization of arrayed 3D hydrogel-based organoids in a high throughput manner. The average achievable output per group was 40 tests per hour, where 20 organoids and 20 reference images in one 35 mm petri dish were tested, illustrating efficient productivity to match the increasing demand on 3D organoids’ applications. The changes in stiffness of collagen I hydrogel organoids in four conditions were measured, with ovarian cancer cells (SKOV3) or without (control). The Young’s modulus of the control group (Control—day 0, E = 407± 146, n = 4) measured by PPC was used as a reference modulus, where the relative elastic compressive modulus of the other groups based on the stiffness measurements was also calculated (control-day 0, E = 407 Pa), (SKOV3-day 0, E = 318 Pa), (control-day 5, E = 528 Pa), and (SKOV3-day 5, E = 376 Pa). The SKOV3-embedded hydrogel-based organoids had more shrinkage and lowered moduli on day 0 and day 5 than controls, consistently, while SKOV3 embedded organoids increased in stiffness in a similar trend to the collagen I control from day 0 to day 5. The proposed method can contribute to the biomedical, biochemical, and regenerative engineering fields, where bulk mechanical characterization is of interest. The μTweezers will also provide attractive design and application concepts to soft membrane-micro 3D robotics, sensors, and actuators.
more »
« less
- PAR ID:
- 10330888
- Editor(s):
- Jabbari, Esmaiel
- Date Published:
- Journal Name:
- PLOS ONE
- Volume:
- 17
- Issue:
- 1
- ISSN:
- 1932-6203
- Page Range / eLocation ID:
- e0262950
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
Cells encapsulated in 3D hydrogels exhibit differences in cellular mechanosensing based on their ability to remodel their surrounding hydrogel environment. Although cells in tissue interfaces feature a range of mechanosensitive states, it is challenging to recreate this in 3D biomaterials. Human mesenchymal stem cells (MSCs) encapsulated in methacrylated gelatin (GelMe) hydrogels remodel their local hydrogel environment in a time-dependent manner, with a significant increase in cell volume and nuclear Yes-associated protein (YAP) localization between 3 and 5 days in culture. A finite element analysis model of compression showed spatial differences in hydrogel stress of compressed GelMe hydrogels, and MSC-laden GelMe hydrogels were compressed (0–50%) for 3 days to evaluate the role of spatial differences in hydrogel stress on 3D cellular mechanosensing. MSCs in the edge (high stress) were significantly larger, less round, and had increased nuclear YAP in comparison to MSCs in the center (low stress) of 25% compressed GelMe hydrogels. At 50% compression, GelMe hydrogels were under high stress throughout, and this resulted in a consistent increase in MSC volume and nuclear YAP across the entire hydrogel. To recreate heterogeneous mechanical signals present in tissue interfaces, porous polycaprolactone (PCL) scaffolds were perfused with an MSC-laden GelMe hydrogel solution. MSCs in different pore diameter (~280–430 μm) constructs showed an increased range in morphology and nuclear YAP with increasing pore size. Hydrogel stress influences MSC mechanosensing, and porous scaffold-hydrogel composites that expose MSCs to diverse mechanical signals are a unique biomaterial for studying and designing tissue interfaces.more » « less
-
Abstract Mechanically tunable hydrogels are attractive platforms for 3D cell culture, as hydrogel stiffness plays an important role in cell behavior. Traditionally, hydrogel stiffness has been controlled through altering either the polymer concentration or the stoichiometry between crosslinker reactive groups. Here, an alternative strategy based upon tuning the hydrophilicity of an elastin‐like protein (ELP) is presented. ELPs undergo a phase transition that leads to protein aggregation at increasing temperatures. It is hypothesized that increasing this transition temperature through bioconjugation with azide‐containing molecules of increasing hydrophilicity will allow direct control of the resulting gel stiffness by making the crosslinking groups more accessible. These azide‐modified ELPs are crosslinked into hydrogels with bicyclononyne‐modified hyaluronic acid (HA‐BCN) using bioorthogonal, click chemistry, resulting in hydrogels with tunable storage moduli (100–1000 Pa). Human mesenchymal stromal cells (hMSCs), human umbilical vein endothelial cells (HUVECs), and human neural progenitor cells (hNPCs) are all observed to alter their cell morphology when encapsulated within hydrogels of varying stiffness. Taken together, the use of protein hydrophilicity as a lever to tune hydrogel mechanical properties is demonstrated. These hydrogels have tunable moduli over a stiffness range relevant to soft tissues, support the viability of encapsulated cells, and modify cell spreading as a consequence of gel stiffness.more » « less
-
A short peptide, FHHF-11, was designed to change stiffness as a function of pH due to changing degree of protonation of histidines. As pH changes in the physiologically relevant range, G′ was measured at 0 Pa (pH 6) and 50,000 Pa (pH 8). This peptide-based hydrogel is antimicrobial and cytocompatible with skin cells (fibroblasts). It was demonstrated that the incorporation of unnatural AzAla tryptophan analog residue improves the antimicrobial properties of the hydrogel. The material developed can have a practical application and be a paradigm shift in the approach to wound treatment, and it will improve healing outcomes for millions of patients each year.more » « less
-
Rizzo, Piervincenzo; Su, Zhongqing; Ricci, Fabrizio; Peters, Kara J (Ed.)Anisotropic collagen-based biomaterials have gained significant attention in the fields of tissue engineering and regenerative medicine. They have shown great potential for wound dressing, corneal grafting, and exploring the mechanism of cancer cell invasion. Various external physical field-based methods for the fabrication of anisotropic collagen-based biomaterials have been developed, including electrospinning, microfluidic shearing, mechanical loading, and so on. In this study, we put forward an acoustic streaming-based method that uses acoustic wave-induced fluid streaming to control collagen self-assembly and fiber arrangement. Our acoustic device leverages a piezoelectric transducer to generate traveling acoustic waves in fluids, and the wave-fluid interaction further induces fluid streaming, known as acoustic streaming. If the fluid contains collagen macromolecules, the acoustic streaming is able to affect the collagen self-assembly process to create biomaterials containing directionally arranged collagen fibers along the streaming velocity direction. Therefore, this acoustic streaming-based method allows for manufacturing collagen hydrogel layers that contain acoustically arranged collagen fibers and have controlled anisotropic material properties. We performed a series of proof-of-concept experiments by using a fabricated acoustic device to control the self-assembly process of collagens loaded in a Petri dish. Our results show the effectiveness of arranging collagen fibers that follow the flow direction of acoustic streaming. To better understand the collagen manipulation mechanism, we used particle image velocimetry to characterize the acoustic wave-induced fluid streaming. We expect this study can contribute to the fabrication of collagen-based anisotropic biomaterials for biomedical applications.more » « less
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1371/journal.pone.0262950
