Genetic variation drives cancer cell adaptation to ECM stiffness.

The progression of many solid tumors is accompanied by temporal and spatial changes in the stiffness of the extracellular matrix (ECM). Cancer cells adapt to soft and stiff ECM through mechanisms that are not fully understood. It is well known that there is significant genetic heterogeneity from cell to cell in tumors, but how ECM stiffness as a parameter might interact with that genetic variation is not known. Here, we employed experimental evolution to study the response of genetically variable and clonal populations of tumor cells to variable ECM stiffness. Proliferation rates of genetically variable populations cultured on soft ECM increased over a period of several weeks, whereas clonal populations did not evolve. Tracking of DNA barcoded cell lineages revealed that soft ECM consistently selected for the same few variants. These data provide evidence that ECM stiffness exerts natural selection on genetically variable tumor populations. Soft-selected cells were highly migratory, with enriched oncogenic signatures and unusual behaviors such as spreading and traction force generation on ECMs with stiffness as low as 1 kPa. Rho-regulated cell spreading was found to be the directly selected trait, with yes-associated protein 1 translocation to the nucleus mediating fitness on soft ECM. Overall, these data show that genetic variation can drive cancer cell adaptation to ECM stiffness.

Development of Antifouling Hyperbranched Polyglycerol Layers on Hydroxyl Poly-p-xylylene Coatings.

Antifouling surfaces that are resistant to protein adsorption and cell adhesion are desirable for many biomedical devices, such as diagnostic devices, biosensors, and implants. In this study, we developed an antifouling hyperbranched polyglycerol (hPG) surface on hydroxyl poly-p-xylylene (PPX-OH). PPX-OH was deposited via chemical vapor deposition (CVD), and an hPG film was then developed via the ring-opening reaction of glycidol. The hPG film greatly reduced the adhesion of L929 cells and platelets as well as protein adsorption. The addition of alkenyl groups in the hPG layer allows the conjugation of biomolecules, such as peptides and biotin, and elicits specific biological interactions. Since the CVD deposition of PPX-OH could be applied to most types of materials, our approach makes it possible to decorate an antifouling hPG film on most types of materials. Our method could be applied to biosensors, diagnostics, and biomedical devices in the future.

Nucleocytoplasmic transport rates are regulated by cellular processes that modulate GTP availability.

Nucleocytoplasmic transport (NCT), the facilitated diffusion of cargo molecules between the nucleus and cytoplasm through nuclear pore complexes (NPCs), enables numerous fundamental eukaryotic cellular processes. Ran GTPase uses cellular energy in the direct form of GTP to create a gradient across the nuclear envelope (NE) that drives the majority of NCT. We report here that changes in GTP availability resulting from altered cellular physiology modulate the rate of NCT, as monitored using synthetic and natural cargo, and the dynamics of Ran itself. Cell migration, cell spreading, and/or modulation of the cytoskeleton or its connection to the nucleus alter GTP availability and thus rates of NCT, regulating RNA export and protein synthesis. These findings support a model in which changes in cellular physiology that alter GTP availability can regulate the rate of NCT, impacting fundamental cellular processes that extensively utilize NCT.

Stiffness assisted cell-matrix remodeling trigger 3D mechanotransduction regulatory programs.

Engineered matrices provide a valuable platform to understand the impact of biophysical factors on cellular behavior such as migration, proliferation, differentiation, and tissue remodeling, through mechanotransduction. While recent studies have identified some mechanisms of 3D mechanotransduction, there is still a critical knowledge gap in comprehending the interplay between 3D confinement, ECM properties, and cellular behavior. Specifically, the role of matrix stiffness in directing cellular fate in 3D microenvironment, independent of viscoelasticity, microstructure, and ligand density remains poorly understood. To address this gap, we designed a nanoparticle crosslinker to reinforce collagen-based hydrogels without altering their chemical composition, microstructure, viscoelasticity, and density of cell-adhesion ligand and utilized it to understand cellular dynamics. This crosslinking mechanism utilizes nanoparticles as crosslink epicenter, resulting in 10-fold increase in mechanical stiffness, without other changes. Human mesenchymal stem cells (hMSCs) encapsulated in 3D responded to mechanical stiffness by displaying circular morphology on soft hydrogels (5 kPa) and elongated morphology on stiff hydrogels (30 kPa). Stiff hydrogels facilitated the production and remodeling of nascent extracellular matrix (ECM) and activated mechanotransduction cascade. These changes were driven through intracellular PI3AKT signaling, regulation of epigenetic modifiers and activation of YAP/TAZ signaling. Overall, our study introduces a unique biomaterials platform to understand cell-ECM mechanotransduction in 3D for regenerative medicine as well as disease modelling.

Dynamically Cross-Linked Granular Hydrogels for 3D Printing and Therapeutic Delivery.

Granular hydrogels have recently emerged as promising biomaterials for tissue engineering and 3D-printing applications, addressing the limitations of bulk hydrogels while exhibiting desirable properties such as injectability and high porosity. However, their structural stability can be improved with post-injection interparticle cross-linking. In this study, we developed granular hydrogels with interparticle cross-linking through reversible and dynamic covalent bonds. We fragmented photo-cross-linked bulk hydrogels to produce aldehyde or hydrazide-functionalized microgels using chondroitin sulfate. Mixing these microgels facilitated interparticle cross-linking through reversible hydrazone bonds, providing shear-thinning and self-healing properties for injectability and 3D printing. The resulting granular hydrogels displayed high mechanical stability without the need for secondary cross-linking. Furthermore, the porosity and sustained release of growth factors from these hydrogels synergistically enhanced cell recruitment. Our study highlights the potential of reversible interparticle cross-linking for designing injectable and 3D printable therapeutic delivery scaffolds using granular hydrogels. Overall, our study highlights the potential of reversible interparticle cross-linking to improve the structural stability of granular hydrogels, making them an effective biomaterial for use in tissue engineering and 3D-printing applications.

Dynamically crosslinked thermoresponsive granular hydrogels.

Granular hydrogels are a promising biomaterial for a wide range of biomedical applications, including tissue regeneration, drug/cell delivery, and 3D printing. These granular hydrogels are created by assembling microgels through the jamming process. However, current methods for interconnecting the microgels often limit their use due to the reliance on postprocessing for crosslinking through photoinitiated reactions or enzymatic catalysis. To address this limitation, we incorporated a thiol-functionalized thermo-responsive polymer into oxidized hyaluronic acid microgel assemblies. The rapid exchange rate of thiol-aldehyde dynamic covalent bonds allows the microgel assembly to be shear-thinning and self-healing, with the phase transition behavior of the thermo-responsive polymer serving as secondary crosslinking to stabilize the granular hydrogels network at body temperature. This two-stage crosslinking system provides excellent injectability and shape stability, while maintaining mechanical integrity. In addition, the aldehyde groups of the microgels act as covalent binding sites for sustained drug release. These granular hydrogels can be used as scaffolds for cell delivery and encapsulation, and can be 3D printed without the need for post-printing processing to maintain mechanical stability. Overall, our work introduces thermo-responsive granular hydrogels with promising potential for various biomedical applications.

Nucleocytoplasmic transport rates are regulated by cellular processes that modulate GTP availability.

Nucleocytoplasmic transport (NCT), the facilitated diffusion of cargo molecules between the nucleus and cytoplasm through nuclear pore complexes (NPCs), enables numerous fundamental eukaryotic cellular processes. Ran GTPase uses cellular energy in the direct form of GTP to create a gradient across the nuclear envelope (NE) that drives the majority of NCT. We report here that changes in GTP availability resulting from altered cellular physiology modulate the rate of NCT, as monitored using synthetic and natural cargo, and the dynamics of Ran itself. Cell migration, cell spreading and/or modulation of the cytoskeleton or its connection to the nucleus alter GTP availability and thus rates of NCT, regulating RNA export and protein synthesis. These findings support a model in which changes in cellular physiology that alter GTP availability can regulate the rate of NCT, impacting fundamental cellular processes that extensively utilize NCT.

Conjugation of Polysulfobetaine via Poly(pyrogallol) Coatings for Improving the Antifouling Efficacy of Biomaterials.

Antifouling treatment is critical to certain biomedical devices for their functions and patients' life. Facial, versatile, and universal coating methods to conjugate antifouling materials on a wide variety of biomaterials are beneficial for the fabrication of low-fouling biomedical devices. We developed a simple one-step coating method for surface conjugation of zwitterionic poly(sulfobetaine) via deposition of self-polymerized pyrogallol (PG). Poly(pyrogallol) could deposit copolymers of sulfobetaine methacrylate and aminoethyl methacrylate (pSBAE) on various biomaterials. pSBAE coatings inhibited as high as 99.8% of the adhesion of L929 cells and reduced protein adsorption significantly. The resistance against L929 cell adhesion was increased with increasing coating time and was positively correlated with the surface hydrophilicity and film thickness. Such a coating was robust to resist harsh sterilization conditions and stable for long-term storage in phosphate-buffered saline. We expect that the simple low-fouling pSBAE coating is applicable to the manufacture of medical devices.