Materials Today Bio

Cell migration in three-dimensional (3D) environments is highly plastic and regulated by extracellular matrix (ECM) cues. Engineered biomaterials provide controllable platforms to investigate how specific matrix signals regulate cell behavior in 3D, yet how defined biochemical signals control migration modes remain unclear. Here, we present tunable fibrous polyisocyanide (PIC) hydrogels functionalized with integrin-binding RGD peptides, cadherin-mimetic HAVDI peptides, or no ligands to direct mesenchymal, hybrid, or amoeboid-like migration of human adipose-derived stem cells without altering matrix mechanics. Using live-cell tracking, 3D displacement microscopy, matrix remodeling analysis, and YAP nuclear localization, we show that ligand identity governs adhesion organization, force transmission, and mechanotransduction. RGD-functionalized matrices promote {beta}1-integrin clustering, extensive matrix remodeling, strong YAP activation and upregulation of migration-related genes. In contrast, non-adhesive matrices limit adhesion formation, resulting in weak force transmission and amoeboid-like behavior. HAVDI-functionalized matrices induce cadherin clustering and heterogeneous cellular responses, indicating that a hybrid migration mode arises from adhesion organization rather than a distinct transcriptional program. Together, these findings demonstrate that ligand identity alone is sufficient to program migration mode in a force-responsive 3D matrix and provide a versatile platform to dissect cell-matrix interactions in complex environments. Statement of significanceO_LICell migration in tissues is highly adaptable, yet precise control of migration modes in defined 3D biomaterials remains challenging. C_LIO_LIWe introduce fibrous PIC hydrogels presenting RGD, HAVDI, or no adhesive ligands to bias human stem cells toward mesenchymal-like, hybrid, or amoeboid-like migration states. C_LIO_LIBy linking ligand identity to adhesion organization, matrix remodeling, YAP mechanotransduction, and gene expression, this work provides a minimal platform to dissect and engineer 3D cell-matrix interactions C_LI

The combination of synthetic biology and additive manufacturing has driven major changes in production of biomaterials, especially through the use of three-dimensional (3D) bioprinting to create engineered living materials. However, current fabrication methods can be limited by prohibitive hardware costs and the inability to maintain structural fidelity in complex, free-form living architectures. This work demonstrates how to build a low-cost, open-source 3D bioprinting platform that can make complicated bacterial structures with complex geometry and high dimensional accuracy. A commercially available, conventional fused deposition modeling 3D printer was modified to create a bioprinting system that is simple to build. The modified bioprinter, which costs around $450, is less expensive than many commercial bioprinters. This 3D-printing technology uses slurry-based support bath methods featuring low-cost gelatin and agarose microparticles, resulting in structures with a high aspect ratio (>8:1) and feature sizes as small as 260 m. The optimization of critical printing settings, including the ability of the bioink to retract during non-print movements, resulted in a reduction of unwanted bacterial deposition by nearly two orders of magnitude. Long-term viability experiments showed that bacteria in the bioprints could survive for at least 28 days with nutrient supplementation. Additionally, 3D-printed engineered biofilms revealed that incubation conditions and extracellular matrix composition significantly impacted the mechanical properties of printed constructs, with tradeoffs between matrix production and mechanical integrity. This study showcases an accessible 3D bioprinting platform for advanced bioprinting technologies, enabling development of engineered living materials with potential applications in synthetic biology, biotechnology, and tissue engineering.

Collagen type II (Col-II) and collagen type I (Col-I) are major components of articular cartilage present at different ratios at its surface. Understanding how each of these components mediates the assembly of molecular films derived from synovial fluid (SF), the native lubricant of synovial joints, is critical to explain the loss of mechanical performance in pathological conditions, guide the design of biomaterial implants meant to be in contact with SF, and develop molecular therapies to restore SF properties. This work demonstrates that Col-II articular surface model assists in scaffolding of full SF-derived films, while Col-I model lacks SF film scaffolding capabilities. However, when Col-II and Col-I are exposed to recombinant lubricin (rLub) alone, the major boundary lubricant in SF, both adsorbed and retained similar amounts. These insights, deduced from quartz crystal microbalance with dissipation, diffuse reflectance circular dichroism, and atomic force microscopy, reveal possible mechanisms underlying the loss of mechanical performance of synovial joints in pathology, where Col-I becomes the major collagenous component of the articular cartilage surface, as well as considerations for designing functional biomaterial implants. Furthermore, this work reinforces the idea of rLub as an intra-articular osteoarthritis therapy with the ability to bind to Col-II and Col-I, irrespectively. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/726594v1_ufig1.gif" ALT="Figure 1"> View larger version (69K): org.highwire.dtl.DTLVardef@138a77borg.highwire.dtl.DTLVardef@7b8512org.highwire.dtl.DTLVardef@15d7060org.highwire.dtl.DTLVardef@17ccf20_HPS_FORMAT_FIGEXP M_FIG C_FIG

The inaccessibility of intracellular bacteria has long rendered the treatment of Staphylococcus aureus infections an challenge. Studies have demonstrated that the extracellular injection system PVC can accurately deliver proteins into cells, which would not need small molecules, and enables effective intracellular delivery of antimicrobial peptides for treatment. Accordingly, we selected antimicrobial peptides including Cecropin, LL37 and Indolicidin that possess potent bactericidal activity, and established the Directed Antimicrobial Assault platform (DAAT) by leveraging the intracellular delivery capacity of PVC. DAAT Cecropin, DAAT LL37 and DAAT Indolicidin inhibited intracellular bacteria in a dose-dependent manner, with DAAT LL37 reaching 86.76% inhibition; after 72 h of treatment, viable-cell numbers reduse to 66--82-fold those of the control. Tail-fibre retargeting enabled direct extracellular S. aureus killing, while combined DAAT therapy promoted wound healing in mice. These findings expand the utility of PVC-derived nanosyringes and establish DAAT as a modular platform for intracellular antimicrobial peptide therapy.

Tissue regeneration using bioactive biomaterials has made great progress in the field of wound healing. Biopolymers play a cardinal role in regenerative medicine by providing safe, biocompatible and bioresorbable support. The electrospinning fabrication technique has been used in creating suitable wound care materials. PHBV and PLLA are FDA approved polymers having important applications in biomedical field. In this study, to increase the wound healing potential, PHBV was functionalized with -COOH group and electrospun nano-fibrous mat was produced using PHBV-COOH and PLLA blended solution. Antibiofilm peptide (IDR-1018) with immunomodulatory activity was incorporated into the blended solution to improve infected wound treatment by actively fighting against bacterial infections. Furthermore, in-vitro experiments including cell cytotoxicity assay and scratch wound healing assay were done to evaluate the potential of the synthesized bioactive nanofibrous mat as a potential wound management aid.

Critical-sized bone defects represent a challenge in bone tissue engineering, due to insufficient vascularization that results in implant failure. Scaffold pre-vascularization is a promising strategy to create a functional microvascular network that integrates with host vasculature. In this study, we present a hybrid 3D construct comprising a hyaluronic acid-based hydrogel and a 3D printed polycaprolactone/{beta}-tricalcium phosphate scaffold, to support vascular network formation and osteogenic differentiation. Peptide-functionalized (i.e. RGD, YIGSR, IKVAV, QK) hydrogels were obtained via thiol-ene chemistry, using two crosslinkers (PEG-diSH or MMP-diSH). Preliminary biological experiments assessed human mesenchymal stromal cells (hMSCs), endothelial cells (hUVECs), and their co-culture, on different gel formulations. All cell conditions displayed enhanced spreading and metabolic activity on gel formulations comprising RGD; thus these (i.e. RGD only and a combination of RGD/YIGSR) were selected for further studies. Cells were then mixed with the hydrogel precursor solutions, which were injected to embed the scaffolds and crosslinked using a UV lamp. After 7 days, tubule formation was observed only in co-culture conditions, highlighting the importance of cellular crosstalk for the formation of a vascular network. Significant differences were found across the tested formulations. In the RGD-PEG constructs, hUVECs formed tubule-like structures, surrounded by hMSCs, exhibiting pericyte-like behavior, supported by the upregulation of SMA gene. Conversely, in the RGD/YIGSR-MMP conditions, hMSCs were mostly located on the scaffold fibers, and showed the highest expression of early osteogenic markers (RUNX2 and ALP). Overall, we demonstrated that the hybrid system with tailored hydrogel chemistry can support simultaneous microvascular organization and osteogenic commitment, offering a promising platform for bone tissue engineering applications. However, further studies involving longer culture periods will aim at clarifying the complex interplay between material composition, cell crosstalk and spatial organization and their influence on the maturation and stability of the vascular network.

Stroke remains a leading cause of adult disability, driven in part by the formation of a non-permissive extracellular matrix environment that limits endogenous repair. Injectable biomaterials that can modulate this microenvironment while enabling localised therapeutic delivery, represent a promising strategy for post-stroke brain regeneration. Here, we report the development of a thermoresponsive hybrid hydrogel composed of chitosan, {beta}-glycerophosphate, silk fibroin, polyvinyl alcohol and polyvinyl pyrrolidone, engineered to provide a tuneable physicochemical properties and enhanced biological functionalities for intracerebral delivery. Systemic optimisation identified a formulation (F6) that exhibited rapid gelation at physiological temperature, appropriate viscoelastic properties, a microporous architecture, and controlled biodegradation, conducive to cellular infiltration and molecular transport. In a mouse model of photothrombotic stroke, intracerebral delivery of the F6 hydrogel attenuated reactive astrogliosis and microglial activation in the peri-infarct region, while enhancing neurogenesis in the subventricular zone. Notably, incorporation of brain-derived neurotrophic factor within the hydrogel significantly improved functional recovery over 8 weeks, demonstrating the capacity of this system to act as a localised delivery platform for neuro-regenerative therapeutics. Together, this study establishes a tuneable thermoresponsive hydrogel platform that integrates structural support with controlled therapeutic delivery, highlighting its potential as a minimally invasive strategy for modulating the post-stroke microenvironment and promoting functional recovery.

The mechanical properties of the cellular microenvironment are key regulators of cellular physiology. Although the field of cancer mechanobiology has attracted attention, the availability of matrix systems with independently and precisely tunable mechanical properties remains limited. Our group previously developed a collagen gel with enhanced mechanical strength by cross-linking collagen molecules using a platinum complex. In this study, we investigated the tunability of the mechanical properties of the platinum cross-linked collagen gel (PCG) and demonstrated that mechanical parameters can be controlled by varying the amount of the platinum complex. In addition, we examined how matrix mechanical properties modulate the phenotypes of lung adenocarcinoma A549 cells using the collagen matrix. Although A549 cells exhibited significant morphological alterations on stiffer matrices, these changes were not accompanied by classical epithelial-to-mesenchymal transition (EMT). Instead, they were associated with the upregulation of diverse gene expression related to cancer malignancy. We focused on maternal embryonic leucine zipper kinase (MELK) whose gene expression increased on stiffer matrices. Consistently, A549 cells cultured on stiffer matrices displayed enhanced sensitivity to a MELK-targeting anticancer drug. These findings highlight the potential of the matrices with tunable mechanical parameters not only to provide variety of physiological microenvironment but also to advance anticancer drug screening when combined with gene expression analysis. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/720034v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@419074org.highwire.dtl.DTLVardef@72ef67org.highwire.dtl.DTLVardef@1c36e17org.highwire.dtl.DTLVardef@170dc33_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIPlatinum cross-linked collagen gel enables independent tuning of compressive and shear elasticities. C_LIO_LICellular functions may be regulated by matrix mechanical parameters through distinct mechanisms. C_LIO_LICorrelation analysis between matrix mechanical parameters and cancer cell gene expression provides a rational strategy for therapeutic drug screening. C_LI

Granular extracellular matrix (gECM)-based biomaterials commonly contain polymer components to improve scaffold cohesion and handling during fabrication and use. However, these polymer hydrogel components may dilute ECM content and increase fabrication and regulatory complexity. This study evaluated whether particle-only gECM wafers could serve as a simplified alternative to hydrogel-based gECM scaffolds while maintaining structural, mechanical, and biological performance. Decellularized human cartilage and skin tissues were processed and fabricated into three scaffold formats: gECM hydrogels, freeze-dried gECM hydrogel wafers, and freeze-dried particle-only gECM wafers. Across fabrication methods, scaffold swelling, volume fraction, and stiffness were strongly influenced by both tissue type and fabrication approach. gECM hydrogels exhibited the greatest swelling and lowest stiffness, while gECM wafers displayed higher volume fractions and greater mechanical stiffness. Notably, gECM particle-only wafers achieved performance comparable to gECM hydrogel wafers despite the absence of a secondary polymer network. Particle-only wafers also maintained swelling behavior and structural properties over 3 months of dry storage at room temperature, with only modest decreases in stiffness. In vitro studies showed sustained cell viability over 14 days on particle-only wafers, with chondrocytes infiltrating cartilage wafers and fibroblasts remaining primarily surface-localized on skin wafers. In addition, particle-only wafers remained cohesive during implantation into a bovine cartilage defect model. These findings demonstrate that particle-only gECM wafers can achieve structural integrity, mechanical performance, and cytocompatibility without the need for an additional polymer network, highlighting a simplified and ECM-rich biomaterial platform. By eliminating polymer carriers and enabling dry storage with preserved function, this approach supports the development of off-the-shelf, translationally accessible gECM particle-only wafers for tissue engineering applications.

Cultivated meat is an emerging biotechnology that aims to produce edible tissues in an ethical and sustainable manner. However, the recreation of skeletal muscle tissue that replicates the protein composition and sensory characteristics of traditional meat is a major challenge. Skeletal muscle tissue engineering requires non-animal-based scaffolds which are inexpensive and food-safe, while meeting specific mechanical requirements with respect to viscosity, stress-relaxation and stiffness. While many of these characteristics can be fulfilled by alginate-based biomaterials, a key limitation of alginate is its lack of intrinsic attachment sites for animal cells, preventing efficient adhesion, differentiation and tissue formation. Here, we established a screening platform to evaluate extracellular matrix (ECM)-mimicking peptides as functionalisations of alginate scaffolds in 2D. Our platform enables high-throughput assessment of cell/peptide interactions, serving as a predictive tool for 3D tissue constructs. Our screen identified two RGD-containing sequences (vitronectin- and fibronectin-mimicking peptides) as most effective in promoting attachment and myogenic fusion of bovine satellite cells. Notably, these peptides outperformed more complex mixtures containing up to seven different ECM-mimicking peptides. Our findings provide a streamlined approach for optimising biomaterial functionalisations for cultivated meat applications, and lay the groundwork for future advancements in scalable, sustainable skeletal muscle tissue engineering.

This study focuses on the development of 3D culture model dedicated to liquid cancers drug screening. The challenge addressed was to effectively retain non adherent small cells within a 3D-scaffold with tailorable mechanical properties, while proposing a fast and effective tool for drug screening. To that aim, we developed a macroporous alginate-chitosan polyelectrolyte complex (PEC) scaffold combined with a low-viscosity alginate (LVA) cell seeding solution. We hypothesized that LVA could undergo in situ pore gelation via calcium ions retained from the PEC fabrication process, enabling effective retention and homogeneous cell distribution, leading to an improved platform for drug screening and personalized medicine. First, we evaluated scaffold suitability for LVA infiltration and gelation. Microtomography revealed a highly porous architecture (98%) enabling LVA homogeneous penetration and complete gelation within 30 min, as confirmed by SEM, microscopy, rheology, and micro-rheology. Next, we assessed cell retention and biocompatibility using primary human chronic lymphocytic leukemia (CLL) cells. LVA-assisted seeding increased cell density 2.6-fold compared to medium alone, with homogeneous distribution, >80% viability over 7 days, and preserved differentiation into nurse-like cells. Finally, we demonstrated a proof of concept for drug screening. The Alginate-PEC scaffold (A-PEC scaffold) supported both qualitative live/dead imaging and rapid quantitative viability measurement with the Alamar Blue assay. Drug responses reproduced microenvironment-dependent protection effects observed in vivo. This integrated scaffold and seeding method provides a promising 3D platform for in vitro liquid cancer studies and drug screening on patient-derived hematological cancer cells. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=67 SRC="FIGDIR/small/722037v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@9b71d4org.highwire.dtl.DTLVardef@14e1dd0org.highwire.dtl.DTLVardef@1876a56org.highwire.dtl.DTLVardef@15656bc_HPS_FORMAT_FIGEXP M_FIG C_FIG

This study aimed to develop a BMSC-laden polyethylene glycol diacrylate/methacrylated hyaluronic acid (PEGDA/HAMA) dual-crosslinked hydrogel and evaluate its effects on osteochondral defect repair. Two PEGDA concentrations, 3.75% and 7.5% (w/v), were used to prepare representative soft and stiff hydrogel formulations, respectively. The hydrogels were characterized in terms of morphology, cytocompatibility, and compressive behavior, and their ability to support BMSC-associated matrix deposition was evaluated in vitro. Repair outcomes were further assessed in a rat osteochondral defect model at 4 and 8 weeks. The 7.5% PEGDA/HAMA hydrogel exhibited higher stiffness than the 3.75% formulation and supported BMSC viability and matrix deposition in vitro. In vivo, the stiff hydrogel group showed improved defect filling and subchondral bone remodeling compared with the soft hydrogel and defect groups. However, histological and immunohistochemical analyses revealed predominant collagen type I deposition and limited collagen type II expression in the repair region, indicating fibrocartilaginous rather than hyaline-like cartilage repair. These findings suggest that BMSC-laden PEGDA/HAMA hydrogels may provide a useful platform for osteochondral defect repair, while further optimization of degradation behavior, matrix maturation, and collagen type II deposition is required to improve hyaline cartilage-oriented repair.

Silk proteins, including sericin and fibroin, are natural biopolymers with broad applications in tissue engineering where angiogenesis plays an essential role. However, the pro-angiogenic effects of silk proteins with varying molecular weights (MWs) remain poorly understood. Here, silk proteins with MW distributions at 40-180 kDa or less than 25 kDa were obtained through alkaline hydrolysis to evaluate their effects on angiogenesis. Structurally, reducing MW induced a conformational transition in silk proteins, accompanied by a striking morphological shift in sericin from nanofibers to nanoparticles. Functionally, high-MW sericin (SSH) suppressed, whereas low-MW sericin (SSL) and both high- and low-MW silk fibroin (SFH/SFL) directly promoted endothelial angiogenic activity. Transcriptomic analysis revealed that angiogenesis-related genes such as Id1 and Smad6/9 may underlie the angiostatic effects of SSH. Notably, both SSH and SSL enhanced angiogenesis indirectly via macrophages; however, SSH induced mixed M1/M2-like polarization, while SSL preferentially drove an M2-like phenotype. In a subcutaneous implantation model, SSH promoted angiogenesis but yielded vessels with weak integrity and increased fibrosis, whereas SSL enhanced angiogenesis with improved vascular maturity and reduced fibrotic response. These findings elucidate how the MWs of silk proteins shape angiogenic behavior and highlight the importance of MW tailoring for optimized tissue engineering applications.

Biodegradable electrospun nanofiber (NF) scaffolds have emerged as promising materials for tissue engineering applications, including vascular grafts, because their mechanical properties and degradability can be tuned. However, their in vivo degradation behavior remains poorly understood. In this study, we characterized the in vivo degradation profiles of representative biodegradable NF materials widely used in small-caliber vascular graft research, namely polycaprolactone (PCL), poly(D,L-lactide) (PLA), polyglycolic acid (PGA), and a PCL/PLA blend, by monitoring molecular weight changes in subcutaneous and vascular environments. Electrospun NF sheets were implanted subcutaneously in mice, and tubular NF grafts were implanted into the abdominal aorta of rats. Samples were harvested for up to 48 weeks after implantation and analyzed primarily by size-exclusion chromatography (SEC) to assess time-dependent changes in molecular weight. Scanning electron microscopy (SEM) and solid-state 13C nuclear magnetic resonance (NMR) were additionally performed to evaluate ultrastructural and chemical changes associated with degradation. SEC analysis revealed distinct material-specific degradation patterns. PCL showed the slowest degradation and retained a relatively high weight-average molecular weight (Mw) in both environments. PLA exhibited marked environment dependence, with near-complete degradation in the subcutaneous environment by 48 weeks, whereas scaffold structure was maintained in the vascular environment. The PCL/PLA blend showed earlier reduction in the high-molecular-weight fraction than PCL, indicating faster scaffold breakdown. PGA degraded most rapidly and could not be evaluated beyond 2 weeks in the subcutaneous model or in the vascular model because of early graft rupture. SEM analysis further demonstrated that progressive loss of fibrous ultrastructure over time was a common feature across all materials. In addition, NF scaffolds became resistant to organic solvent after implantation in vivo, and solid-state 13C NMR analysis of the solvent-insoluble fractions detected polymer-derived signals together with additional signals consistent with biological constituents. These findings indicate that in vivo degradation of biodegradable NF scaffolds is material dependent, environment dependent, and more complex than simple hydrolytic chain cleavage alone. This study provides a quantitative framework for evaluating NF degradability and offers new insight into the design of biodegradable vascular grafts. HighlightsO_LISEC quantified long-term in vivo degradation of PCL, PLA, PGA, and PCL/PLA. C_LIO_LIDegradation was both material dependent and implantation environment dependent. C_LIO_LIIn vivo nanofiber degradation involved structural and chemical changes beyond hydrolysis. C_LI

While defined synthetic substrates can replace Matrigel for human induced pluripotent stem cell (hiPSC) culture and hiPSC-derived cardiomyocyte (hiPSC-CM) production, existing approaches culture cells on two-dimensional surfaces and yield structurally immature cardiomyocytes, limiting their use in disease modeling and regenerative medicine. Here, we developed a xeno-free, fully-defined cyclic RGD (cRGD)-functionalized alginate platform in which we encapsulated hiPSCs to support their expansion and in situ cardiac differentiation. cRGD functionalization was essential for hiPSC survival and pluripotency, with maximal support achieved at a low ligand density (25 M). In the presence of cRGD, hiPSC encapsulation into softer gels made from lower molecular weight alginates led to enhanced hiPSC expansion and improved cardiogenesis. Strikingly, differentiation in situ with 3D gels led to hiPSC-CM with higher structural maturity, including a markedly increased proportion of Desmin positive cardiomyocytes. Finally, after enzymatic retrieval from hydrogels, cardiomyocytes derived from softer gels formed tissue-engineered myocardium with superior contractile force compared to tissue fashioned from hiPSC-CM derived from more rigid gels. Together, these results demonstrate the promise of this defined, tunable platform for biomanufacturing of structurally mature cardiomyocytes from hiPSC.

Rapid revascularization is critical to tissue graft survival, as delayed reperfusion drives tissue ischemia and compromises cell viability and graft function. Although bulk hydrogels have been explored for promoting vessel formation, vascularization remains too slow to prevent ischemic injury to grafted tissues, highlighting the need for biomaterial platforms that accelerate graft revascularization and reperfusion. In this study, we present granular hydrogel composites (GHCs), where interstitial space is filled with fibrin and collagen to provide a vasculogenic matrix environment. GHCs supported the assembly of embedded endothelial cells into interconnected, lumenized networks in vitro which anastomosed with host vasculature and were systemically perfused 7 days after implantation. Careful optimization studies revealed that GHCs formed from covalently interlinked, RGD-functionalized microgels of 115 {micro}m diameter best supported vascular network formation in vitro and intravascular blood perfusion in vivo. To test the utility of GHCs for the vascular integration of a demanding and therapeutically relevant parenchymal tissue, GHC-based ovarian tissue grafts were implanted in a murine xenograft model and successfully connected to host vasculature, restoring blood flow to embedded human ovarian tissues within 10 days post-implantation. Notably, endothelial cells seeded within GHCs formed viable vasculature without pre-culture. This work establishes GHCs as a biomaterial platform to rapidly connect parenchymal tissues to host vasculature, with broad translational potential across engineered tissue grafting applications.

Biofilm extracellular matrix (ECM) varies with environmental conditions and substrate properties. Understanding the surface-biofilm relationship helps to perfect antibacterial strategies and to design new engineered living materials (ELMs). In this work, we studied how cationic and anionic polyelectrolyte coatings affect macroscopic features of Escherichia coli curli-producing biofilms, as well as the properties of their curli amyloid fibers. Cationic coatings limited biofilm spreading, increased their surface density and water absorption, which correlated with a higher yield of curli amyloid fibers with looser structure. In contrast, anionic surfaces allowed for standard biofilm spreading, with a lower fiber yield but a more compact and chemically stable fiber structure. Higher biofilm rigidity and adhesion were measured on both types of charged surfaces. Thus, we propose that the differences in biofilm macroscopic properties result from a trade-off between curli quantity and quality in the ECM, namely fiber density and molecular packing, as well as their interaction with water. Our findings provide insights on how the biophysical properties of the ECM can be controlled by tuning the substrate physico-chemical characteristics with charged coatings. This work opens up new avenues for developing antimicrobial strategies, as well as tailoring the properties of amyloid-based ELMs. TOC figure O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=82 SRC="FIGDIR/small/721109v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@191cd79org.highwire.dtl.DTLVardef@148f914org.highwire.dtl.DTLVardef@1d8c2f8org.highwire.dtl.DTLVardef@1e84eaf_HPS_FORMAT_FIGEXP M_FIG C_FIG

Tissue engineered skin models are important tools for the in vitro study of physiological and pathophysiological processes as well as the valuation of therapeutic strategies and the efficacy of pharmaceutical and cosmetic compounds. Replicating the functional anatomy of cutaneous tissue is a crucial aspect in ensuring that observations made using these models are translatable to the actual situation in native skin. However, most contemporary full-thickness skin models neglect the reconstruction of the undulated microtopography of the dermal-epidermal junction (DEJ), which not only contributes to the biological functionality of the skin (e.g. stem cell niches), but also affects tissue mechanics and drug diffusion. Herein, we fabricated bilayer skin models with DEJ-like microtopographies introduced by interfacial wrinkling between a hydrogel and a nanofibrous membrane through a controllable swelling-deswelling approach. The interfacial wrinkles contributed to the structural integrity of the bilayer models. Their formation could be induced in the presence of living cells through mechanical stress-driven buckling instabilities, thus differentiating the process from commonly used pre-patterning techniques. Bilayer models supported the co-culture of human dermal fibroblasts and human epidermal keratinocytes, and the formation of stratified epithelia. Our findings provide a potential alternative method to introduce DEJ-like anatomical features into full-thickness skin tissue models.

Targeted lipid nanoparticles (LNPs) for extrahepatic drug delivery are limited by apolipoprotein E (ApoE)-mediated hepatic accumulation. We developed NanoPilot, a modular fusion protein platform comprising antibodies and anchors blocking the low density lipoprotein receptor (LDLR) ApoE binding site, to block LNP liver uptake and redirect to target cells. NanoPilot can be applied to preformulated LNPs in 10 minutes with two pipetting steps. In vitro, an anti-CD3{varepsilon} NanoPilot increased T-cell transfection 40-fold and reduced monocyte transfection 10-fold in human peripheral blood mononucleocytes. In immunocompetent mouse models, NanoPilot-coated LNPs achieved 30-40% splenic and hepatic T-cell transfection whilst bulk liver accumulation was reduced 3-fold. An anti-c-Kit NanoPilot was further shown to enhance delivery to a haematopoietic stem cell-like cell line in an in-vitro co-culture assay. NanoPilot establishes a versatile framework for the systemic delivery of genetic therapies through concomitant cell-specific targeting and off-target blocking. Further research will assess potential clinical applications.

The transplantation of stem cell-derived extracellular vesicles (EVs) holds promise for tissue repair and regeneration, but scalable production and effective delivery to target tissue remain major challenges. Here, we present a biomaterial platform that combines high-yield, scalable nanovesicles (NVs) - EV mimetics derived from human induced pluripotent stem cells - with an adhesive silk hydrogel patch for localized and sustained delivery. We show that this platform enables efficient NV encapsulation via visible light crosslinking and supports controlled release over short (2 days), intermediate (7 days), and extended (up to 28 days) periods, while maintaining adhesion to heart tissue. Importantly, the sustained delivery of NVs for 3 days in vitro results in promoting anti-fibrotic cell remodeling and significant functional recovery of primary myofibroblast activation, modulating integrin signaling, actomyosin organization, and cell-matrix adhesion networks. Finally, we demonstrate biocompatibility, retention, and anti-fibrotic function of the patch in a murine ischemia-reperfusion injury model. Thus, we establish the proof-of-principle that di-tyrosine silk hydrogels can be used as a strategy to encapsulate and deliver NVs to the heart, thus offering an innovative delivery platform for NVs. Statement of significanceExtracellular vesicles (EVs) represent an emerging frontier in tissue engineering. Their cell-specific cargo contains biological information capable of repairing and regenerating injured tissues. However, their clinical translation is hindered by limited manufacturing scalability, undefined dosing and modes of administration, and low organ retention, particularly in the heart. This study addresses these challenges by combining stem cell-derived nanovesicles (NVs), which mimic biological EVs, with an adhesive hydrogel patch for localized and sustained delivery to the heart. We provide proof-of-principle that di-tyrosine photo-crosslinked silk hydrogels are a suitable delivery platform for cell-derived NVs, preserving NV bioactivity and their ability to remodel recipient cells following delivery both in vitro and in vivo. This study integrates three key advantages: (i) the use of scalable iPSC-derived nanovesicles as an EV-mimetic platform, addressing limitations in EV manufacturing; (ii) a mechanically robust and tunable silk fibroin hydrogel formed via visible light-induced di-tyrosine crosslinking without chemical modification; and (iii) an injection-free, adhesive patch-based delivery strategy enabling localized and sustained therapeutic administration to the heart. This innovative platform represents a significant advancement in the fields of nanomedicine and biomedical engineering. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/722555v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@fed253org.highwire.dtl.DTLVardef@1a270b0org.highwire.dtl.DTLVardef@19437c1org.highwire.dtl.DTLVardef@1d863ca_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO C_FIG