Prof. Dr. Aránzazu del Campo

Photocrosslinkable formulations based on the radical thiol-ene reaction are considered better alternatives than methacrylated counterparts for light-based fabrication processes. This study quantifies differences between thiol-ene and methacrylated crosslinked hydrogels in terms of precursors stability, the control of the crosslinking process, and the resolution of printed features particularized for hyaluronic acid (HA) inks at concentrations relevant for bioprinting. First, the synthesis of HA functionalized with norbornene, allyl ether, or methacrylate groups with the same molecular weight and comparable degrees of functionalization is presented. The thiol-ene hydrogel precursors show storage stability over 15 months, 3.8 times higher than the methacrylated derivative. Photorheology experiments demonstrate up to 4.7-times faster photocrosslinking. Network formation in photoinitiated thiol-ene HA crosslinking allows higher temporal control than in methacrylated HA, which shows long post-illumination hardening. Using digital light processing, 4% w/v HA hydrogels crosslinked with a dithiol allowed printing of 13.5 × 4 × 1 mm3 layers with holes of 100 µm resolution within 2 s. This is the smallest feature size demonstrated in DLP printing with HA-based thiol-ene hydrogels. The results are important to estimate the extent to which the synthetic effort of introducing –ene functions can pay off in the printing step.

The potential of bioactive coatings as an innovative biotechnology to overcome the mass-transfer limitations of conventional technologies when treating air pollutants, especially hydrophobic volatile organic compounds, was herein assessed. Bioactive coatings consist of active microorganisms entrapped in a polymer matrix, which needs to be porous to facilitate an effective gas pollutant exchange. To increase porosity, two additives, sucrose and glycerol mixtures (Suc/Gly) and halloysite nanotubes (HNTs), were included in the bioactive coatings at two concentration levels. The toluene removals of the different bioactive coatings were studied in batch mode at low (∼300 mg m−3) and high (∼3000 mg m−3) toluene concentrations. Overall, low HNTs concentration coatings supported optimum toluene removals (>95 %), comparable to biofilm controls at both toluene concentrations. High HNTs concentration coatings and low Suc/Gly concentration coatings achieved toluene removals over 95 % after 7 toluene injections at low toluene concentration. At high toluene concentrations, these coatings eventually outperformed the biofilm controls. High Suc/Gly concentration coatings supported a limited toluene removal (4 and 1 injection at low and high toluene concentrations, respectively), attributed to a preferential consumption of sucrose over toluene. These findings were corroborated by ESEM/conventional SEM imaging, revealing porosity in the HNTs bioactive coatings, visible at both the surface and internal levels. On the contrary, more homogeneous surfaces were observed in the Suc/Gly bioactive coatings, where total polymer coalescence was partially hindered by the addition of Suc/Gly. These results paved the way towards the implementation of bioactive coating in larger bioreactors for real-life air purification.

The persistent accumulation of plastic waste, particularly polystyrene (PS), poses significant environmental challenges because of its extensive use and low recycling rates. Addressing these challenges necessitates innovative and sustainable solutions. This study presents a strategy to upcycle PS waste into valuable chemical products, including adipic acid, hexanediol, hexamethylenediamine, and nylon-6,6, using metabolically engineered Pseudomonas putida KT2440. This process involves the photolytic degradation of PS into benzoic acid, followed by microbial conversion into cis,cis-muconate (MA) and chemical synthesis of the final products. The engineered strains withstood 30 mM concentrations of PS-derived aromatics and converted them stoichiometrically into MA in the presence of glucose as a growth substrate. 13C metabolic flux analysis revealed energy and redox limitations in the presence of 25 mM benzoate and 300 mM MA. The cells responded to stress by enhancing the flux for periplasmic glucose oxidation and fluxes through the NADPH-forming dehydrogenases; this process caused more than 40 % glucose‑carbon loss into byproducts. Fine-tuned dynamic glucose and benzoate feeding enabled high-level MA production. Energy-optimized genome-reduced strains were used to increase carbon efficiency. A final MA titer of over 65 g L−1 was achieved in fed-batch fermentation. This process was demonstrated using the glucose derived from a viscose textile waste blend as the growth substrate and resulted in fully waste-based products. The resulting adipic acid and hexamethylenediamine were polymerized into nylon-6,6 with properties comparable to those of petrochemical-derived polymers, revealing a sustainable pathway for PS upcycling. This research provides a proof-of-concept for bacterial upgrading of PS-derived substrates and a viable method for managing plastic waste and producing valuable chemical products.

Focal adhesions (FAs) are dynamic structures central to cell migration, serving as mechanotransduction sites linking the extracellular matrix (ECM) to intracellular signaling pathways such as FA kinase (FAK). How FAK becomes activated in response to cell-ECM adhesive forces at single FAs to facilitate directional motion is poorly understood. Using micropillar-based force microscopy and FA-targeted FRET biosensors, we monitored real-time traction forces and FAK activity at individual FAs during assembly and disassembly. Our results demonstrate oscillatory temporal coupling of traction force and FAK activity in high-tension FAs before FA disassembly. Cross-correlation analyses revealed that force precedes FAK activation, guiding FA turnover. Atomistic molecular simulations unveiled a force-induced mechanism where traction forces disrupt autoinhibitory FERM-kinase interactions in FAK, enabling catalytic activity without structural unfolding. Our findings provide mechanistic insights into the spatiotemporal integration of mechanical forces and biochemical signaling in cell migration.

Pluronic (Plu) hydrogels mixed with variable fractions of Pluronic diacrylate (PluDA) have become popular matrices to encapsulate bacteria and control their growth in engineered living materials. Here we study the rheological response of 30 wt.% Plu/PluDA hydrogels with PluDA fraction between 0 and 1. We quantify the range of viscoelastic properties that can be covered in this system by varying in the PluDA fraction. We present stress relaxation and creep-recovery experiments and describe the variation of the critical yield strain/stress, relaxation and recovery parameters of Plu/PluDA hydrogels as function of the covalent crosslinking degree using the Burgers and Weilbull models. The analyzed hydrogels present two stress relaxations with different timescales which can be tuned with the covalent crosslinking degree. We expect this study to help users of Plu/PluDA hydrogels to estimate the mechanical properties of their systems, and to correlate them with the behaviour of bacteria in future Plu/PluDA devices of similar composition.

The hierarchical design of the toe pad surface in geckos and its reversible adhesiveness have inspired material scientists for many years. Micro- and nano-patterned surfaces with impressive adhesive performance have been developed to mimic gecko's properties. While the adhesive performance achieved in some examples has surpassed living counterparts, the durability of the fabricated surfaces is limited and the capability to self-renew and restore function—inherent to biological systems—is unimaginable. Here the morphogenesis of gecko setae using skin samples from the Bibron´s gecko (Chondrodactylus bibronii) is studied. Gecko setae develop as specialized apical differentiation structures at a distinct cell–cell layer interface within the skin epidermis. A primary role for F-actin and microtubules as templating structural elements is necessary for the development of setae's hierarchical morphology, and a stabilization role of keratins and corneus beta proteins is identified. Setae grow from single cells in a bottom layer protruding into four neighboring cells in the upper layer. The resulting multicellular junction can play a role during shedding by facilitating fracture of the cell–cell interface and release of the high aspect ratio setae. The results contribute to the understanding of setae regeneration and may inspire future concepts to bioengineer self-renewable patterned adhesive surfaces.

Biofunctionalized polyacrylamide (PAAm) hydrogels are important 2D substrates for studying cell physics and mechanobiology. In this work, an arylmethylsulfone (MS) comonomer is developed that can be incorporated into PAAm gels under aqueous radical polymerization conditions. The resulting hydrogels show similar properties to unmodified PAAm gels, indicating that the comonomer is incorporated without affecting PAAm physical properties. The MS-containing PAAm hydrogels allow efficient conjugation of thiol derivatized biomolecules and require very low comonomer content (2 mM, 0.18 mol% relative to AAm) and thiol incubation amounts (≥ 0.15 µg per gel) to achieve functional densities that elicit cell responses. Compared to carboxyl-functionalized PAAm hydrogels, a 10-fold lower comonomer concentration and a 10-fold lower ligand feed concentration are sufficient to achieve comparable cell adhesion responses. The new comonomer opens up possibilities for efficient and straightforward biofunctionalization of PAAm hydrogels used in cell biophysical studies.

Natural killer (NK) cells play a vital role in eliminating tumorigenic cells. Efficient locating and killing of target cells in complex three-dimensional (3D) environments are critical for their functions under physiological conditions. However, the role of mechanosensing in regulating NK-cell killing efficiency in physiologically relevant scenarios is poorly understood. Here, we report that the responsiveness of NK cells is regulated by tumor cell stiffness. NK-cell killing efficiency in 3D is impaired against softened tumor cells, whereas it is enhanced against stiffened tumor cells. Notably, the durations required for NK-cell killing and detachment are significantly shortened for stiffened tumor cells. Furthermore, we have identified PIEZO1 as the predominantly expressed mechanosensitive ion channel among the examined candidates in NK cells. Perturbation of PIEZO1 abolishes stiffness-dependent NK-cell responsiveness, significantly impairs the killing efficiency of NK cells in 3D, and substantially reduces NK-cell infiltration into 3D collagen matrices. Conversely, PIEZO1 activation enhances NK killing efficiency as well as infiltration. In conclusion, our findings demonstrate that PIEZO1-mediated mechanosensing is crucial for NK killing functions, highlighting the role of mechanosensing in NK-cell killing efficiency under 3D physiological conditions and the influence of environmental physical cues on NK-cell functions.

The increasing prevalence of dry eye syndrome in aging and digital societies compromises long-term contact lens (CL) wear and forces users to regular eye drop instillation to alleviate discomfort. Here a novel approach with the potential to improve and extend the lubrication properties of CLs is presented. This is achieved by embedding lubricant-secreting biofactories within the CL material. The self-replenishable reservoirs autonomously produce and release hyaluronic acid (HA), a natural lubrication and wetting agent, long term. The hydrogel matrix regulates the growth of the biofactories and the HA production, and allows the diffusion of nutrients and HA for at least 3 weeks. The continuous release of HA sustainably reduces the friction coefficient of the CL surface. A self-lubricating CL prototype is presented, where the functional biofactories are contained in a functional ring at the lens periphery, outside of the vision area. The device is cytocompatible and fulfils physicochemical requirements of commercial CLs. The fabrication process is compatible with current manufacturing processes of CLs for vision correction. It is envisioned that the durable-by-design approach in living CL could enable long-term wear comfort for CL users and minimize the need for lubricating eye drops.

Polyacrylamide (PAAm) hydrogels are widely adopted as 2D-model soft substrates for investigating cell-material interactions in a controlled in vitro environment. They offer facile synthesis, tunable physico-chemical properties, diverse biofunctionalization routes, optical transparency, mouldability in a range of geometries and shapes, and compatibility with living cells. PAAm hydrogels can be engineered to reconstruct physiologically relevant biointerfaces, like cell-matrix or cell–cell interfaces, featuring biochemical, mechanical, and topographical cues present in the extracellular environment. This Review provides a materials science perspective on PAAm material properties, fabrication, and modification strategies relevant to cell studies, highlighting their versatility and potential to address a wide range of biological questions. Current routes are presented to integrate cell-instructive features, such as 2D patterns, 2.5D surface topographies, or mechanical stiffness gradients. Finally, the recent advances are emphasized toward dynamic PAAm hydrogels with on-demand control over hydrogel properties as well as electrically conductive PAAm hydrogels for bioelectronics.