Advanced Materials Technologies

Sensitive and low-cost protein biomarker detection is critical for disease diagnosis. Advanced microfluidic systems can generate miniature reaction compartments for a high-sensitivity assay. However, these platforms often require external instruments, skilled operators, and complex setups. Here, we develop a Lab on a Capillary (LabCap) platform that integrates photopatterned hydrogel rings within a glass capillary using a reconfigurable stop-flow lithography system. During sample loading and unloading steps, nanoliter-scale aqueous droplets (torodrops) are spontaneously formed around the hydrogel rings, creating isolated reaction compartments without the need for external instruments or an immiscible oil phase. The LabCap platform enables quantitative detection of clinically relevant biomarkers, including C-reactive protein (CRP) and N-terminal pro-B-type natriuretic peptide (NT-proBNP). By adjusting the incubation protocol, assay speed and sensitivity can be tuned to meet different analytical requirements. A periodic medium exchange protocol enables biomarker detection at concentrations as low as 1 ng/mL, whereas prolonged static incubation extends detection to 0.1 ng/mL. In addition, LabCap offers practical advantages, including low fabrication cost (< EUR 1 per device), low reagent consumption (<100 microlitres per assay step), and minimal wash-buffer usage (1 mL). These results demonstrate that LabCap is a simple, cost-effective, and versatile platform for biomarker detection.

Conventional extrusion-based 3D bioprinting encounters limitations in fabricating intricate tissue architectures due to fixed nozzle diameters and fixed deposition orientations. These constraints restrict conformal printing on curved or non-planar surfaces and often necessitate support-intensive fabrication strategies. This work introduces a mechanically simplified extrusion platform inspired by the swivel jet nozzle, featuring a free-degree-of-orientation extrusion head termed the universal extrusion head (Univ-Ex head), coupled with a modular nozzle architecture. The Univ-Ex head employs a swivel-like mechanical design that enables orientation freedom without external actuation in its current implementation, thereby minimizing mechanical complexity while supporting deposition on physiologically relevant, non-planar geometries. Multiple nozzle concepts were developed through comparative CAD iterations, with two representative geometries--a flat nozzle and a conical nozzle--selected for experimental validation. The platform is evaluated through parametric CAD design, stereolithography-printed prototypes, proof-of-concept extrusion experiments, and fluid dynamics simulations performed using FLOW-3D software. Numerical and experimental results demonstrate stable filament formation and clear diameter-dependent extrusion behavior, while simulations further confirm the feasibility of angled and non-planar deposition. A variable-diameter nozzle concept is proposed as a forward design direction to enable real-time adjustment of bioink flow rate and deposition resolution in principle; however, the present study intentionally validates the system using fixed-diameter nozzle variants to maintain stable numerical and experimental boundary conditions. A gear-integrated Univ-Ex head is also presented as a forward upgrade and demonstrated as a single-piece prototype. Collectively, this work establishes a scalable, hardware-focused pathway toward conformal bio-additive manufacturing. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=96 SRC="FIGDIR/small/734010v1_ufig1.gif" ALT="Figure 1"> View larger version (63K): org.highwire.dtl.DTLVardef@8833caorg.highwire.dtl.DTLVardef@33dforg.highwire.dtl.DTLVardef@14d8d11org.highwire.dtl.DTLVardef@685ef0_HPS_FORMAT_FIGEXP M_FIG C_FIG

Non-obstructive azoospermia (NOA) represents the most severe form of male infertility, severely limiting a patient's prospects for biological fatherhood when surgical retrieval fails. However, the true biological limits of NOA remain obscured by the inherent limitations of conventional gamete recovery protocols: standard centrifugation frequently causes substantial cell loss, masking extremely rare sperm, while surgical interventions are constrained by spatial sampling biases. Here we report SpermSeek, an integrated AI-guided microfluidic platform for real-time, non-destructive isolation of single sperm directly from semen. Operating at scalable throughput (0.36 mL/h), the system achieves 98.3% detection precision and a 95.5% target encapsulation efficiency, suppressing background debris. In a 59-patient NOA cohort, SpermSeek detected morphologically identifiable sperm in 64.4% (38/59) of cases, spanning diverse genetic etiologies, including AZFb/c microdeletions, and severe histopathological phenotypes, such as Sertoli-cell-only syndrome (SCOS). Notably, among a sub-cohort of 41 patients who remained consistently sperm-negative despite prior medical or micro-TESE interventions, our platform identified gametes in 53.7% (22/41) of these cases. Comprehensive safety profiling in healthy human donors and wild-type mice confirmed that processed sperm retain high DNA integrity and epigenomic concordance (r=0.98), supporting transgenerational developmental stability in mice. Furthermore, in a 26-patient validation cohort, SpermSeek recovered rare sperm in 11 cases. Utilizing gametes from a subset (n=5), we demonstrated their capacity to support early human embryogenesis, yielding high-quality cleavage-stage embryos with confirmed genomic euploidy. This work establishes a highly sensitive framework for re-examining the biological limits of human spermatogenesis, laying the foundation to expand autologous reproductive options for patients refractory to conventional retrieval protocols.

Foreign body response is a common yet serious challenge for biomedical implants. It can trigger inflammation and eventually lead to the formation of a fibrotic capsule, which compromises device function. Although significant efforts have been made to develop antifibrotic surface coatings for implantable materials, developing broadly applicable solutions remains challenging due to the diversity of materials used in biomedical implants. Here, we propose a simple and versatile strategy to develop antifibrotic coatings for biomedical implants. Photoreactive benzophenone groups are incorporated into designer polymers to enable covalent attachment to various substrates. The effect of benzophenone group density within polymer chains on surface coating efficiency was investigated, and an optimal BP incorporation ratio was identified. Polymers incorporating varying ratios of an anti-fibrotic small molecule and anti-fouling zwitterionic moieties were synthesized and successfully attached to silicone implants. In vivo evaluation of these implants in C57BL/6 mice identified an optimized polymer composition that reduced fibrotic capsule thickness by around 60%. Coating of commercial medical catheters with this optimized polymer reduced collagen deposition by over 3.5-fold following 4 weeks of implantation in the peritoneal space of C57BL/6 mice. Finally, we demonstrated that the optimized polymer coating can be readily applied to a variety of commonly used biomedical materials using this straightforward method, highlighting the versatility of the approach. This work provides a facile and broadly applicable strategy for developing antifibrotic coatings, which has the potential to expand the design of surface modifications aimed at improving the performance of biomedical implants.

Two-photon polymerization (2PP) enables fabrication of hydrogel constructs with submicron, cell-scale resolution, but hydrogel-based bioinks are markedly more sensitive to process variability than conventional photoresists, and this sensitivity is further amplified when living cells are embedded in the resin. Post-processing evaluation, performed only after development, occurs too late to enable any corrective action. A full-factorial design of experiments across laser power and scan speed shows that fabrication outcome depends on both parameter choice and cell presence, with cells shifting and broadening the range of conditions yielding structurally sound constructs. However, substantial variability persists within each nominal condition and cannot be resolved by parameter refinement alone, indicating that outcome is governed by what occurs during each individual print rather than by the parameters set. To capture this, a layer-wise polymerization score is derived from pairwise comparisons of same-layer coaxial images, grounded in the psychophysics of relative judgment, and assembled into a Layer-wise Image Trajectory (LIT) for each print. Applied to both acellular and cell-laden formulations, LIT curves separate cleanly by post-processing outcome without any outcome label used in training, showing that fabrication quality can be predicted early in the build. Building on this signal, individual LIT curves are compared against statistical control limits derived from confirmed successful prints, enabling early detection of anomalous fabrication behavior at early-to-mid layers, well before development. To the best of the authors knowledge, this is the first application of in situ quality prediction and anomaly detection to cell-laden two-photon polymerization.

Control of fluids is a hallmark of microfluidic systems and fundamental for the successful application of microfluidic devices. Trigger valves use geometric features to autonomously control the release of fluids in microfluidic devices. Our previous work has adapted geometries used in closed trigger valve systems to enable use in open systems, allowing for open microfluidic devices with up to three trigger valves. Here, we focus on the parallel co-flows produced by sequential release of trigger valves and present a model that predicts their layer widths as a function of the geometric characteristics of the different side channels of each trigger valve. We show layered co-flows with widths as low as 50 microns. Additionally, we expand the use of trigger valves in open microfluidic devices by incorporating 1) varied step heights, 2) devices with up to seven trigger valves, and 3) use of varied fluids and plastics. To validate the implementation and use of these trigger valves in open systems, we have developed a theoretical framework to compare predicted outcomes (i.e., fluid travel distance, velocity, and layering width) with our experimental values. This theoretical work offers applications in various fields, including hydrogel patterning for 3D cell culture, organ-on-a-chip models, at-home sample preparation, and autonomous microfluidic systems for biosensing.

Synthetic optical skin models offer reproducible, tunable optical properties but lack biological responsiveness, while tissue engineered skin models provide cellular authenticity but suffer from optical variability and limited controllability. The growing demand for alternatives to animal models in the development and validation of optical biomedical technologies highlights the need for a new class of test system that combines the strengths of both approaches while addressing their respective limitations. Here, we introduce the concept of a modular biohybrid skin model, a new testing concept that integrates an optically defined artificial epidermal layer, incorporating polydopamine nanoparticles for changes in skin tone, with living human keratinocytes in two and three-dimensional configurations. In the Optical Protection Model, UV-B-induced apoptosis in primary keratinocytes is quantitatively modulated by model pigmentation level, demonstrating a relationship between optical attenuation and caspase 3/7 activity across three artificial skin tone conditions. In a Structured Dermal Model, keratinocytes seeded onto a hydrogel scaffold localize within follicle-like microcavities, as confirmed by live/dead staining and confocal z-stack imaging. Together, these experiments lead to a new category of test system in the space between inert optical models and variable tissue models that may contribute to reducing the reliance on animal models in biomedical optics.

Engineered epithelial models require three-dimensional extracellular matrix environments that support organized cell growth and allow independent access to luminal and basal compartments. However, many organ-on-chip (OoC) fabrication strategies rely on planar geometries, non-native materials, or multi-step assembly workflows that limit architectural complexity and experimental control. Here, we report a direct in-chip volumetric printing strategy for fabricating stretchable and perfusable collagen-I scaffolds inside custom OoC devices. A vitamin C-regulated ruthenium/sodium persulfate photocrosslinking system enabled high-fidelity printing of collagen-I into open-lumen architectures with ductal- and alveolar-inspired features. By generating scaffolds directly within the final culture device, this workflow eliminates post-print transfer and integrates defined collagen architectures with compartmentalized fluidic access and a mechanically actuable chip format. To support chip-based culture, printed collagen constructs were stabilized after fabrication using EDC/NHS chemistry, which limited thermally induced collagen densification, improved shape retention, and maintained scaffold anchorage during perfusion. The chip design provided separate access to the printed lumen and surrounding basal compartment, which enabled compartment-specific fluid handling while preserving scaffold integrity during inflation, stretching, and perfusion of the printed construct. On the collagen-I scaffolds, human milk-derived mammary epithelial cells formed epithelial layers with tight junctions and lactation associated markers. The platform further supported perfusion culture, in situ staining, and whole-chip volumetric imaging. Together, this work establishes direct in-chip collagen-I volumetric printing as a biofabrication strategy for creating perfusable epithelial tissue chips with native matrix architecture and compartmentalized fluidic control.

Myocardial infarction highlights an urgent need for strategies to regenerate functional cardiac tissue. Cardiac tissue engineering offers a promising approach; however, fabricating scaffolds that simultaneously integrate precise architectural anisotropy, mechanical compliance, and electrical conductivity remains an open challenge. In this work, we utilized melt electrowriting (MEW) to construct well-defined, 20-layer anisotropic rhomboidal polycaprolactone (PCL) scaffolds. We characterised them by tensile testing and by micro- and nanoscale microscopy. While introducing electrical conductivity via bulk blending with fillers (polypyrrole (PPy), polyaniline, or graphene oxide) compromised MEW print fidelity and failed to achieve physiological conductivity, surface coating strategies effectively combined conductivity from structural mechanics. Electrical and mechanical testing revealed that gold sputter coating and in situ PPy polymerization both imparted robust electrical conductivity while preserving the microfibrous architecture. However, when seeded with human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in fibrin hydrogels, only the gold-coated scaffolds supported synchronized, robust, and sustained contractile activity. PPy-coating resulted in functionally restricted constructs, suggesting that excessive structural rigidity limited tissue deformability. Gene expression analysis further revealed that elevated electrical conductivity alone does not drive hiPSC-CM maturation. Our data indicates that successful cardiac patch design relies on the integrated optimization of mechanics and architecture rather than treating conductivity as an isolated parameter, offering foundational guidelines for developing translational bioengineered heart patches.

Implantable microdevices capable of autonomous operation over extended lifetimes are promising enablers for minimally invasive diagnostics and therapy. Microelectromechanical systems (MEMS)-based piezoelectric ultrasonic energy harvesters (PUEH) have emerged as a compelling approach for powering implantable microdevices, where both miniaturization and efficient wireless energy transfer are essential. Here, we present a highly miniaturized (5 x 5 x 5 mm3) ultrasonic energy-harvesting platform enabling sustained in vivo operation of implantable microdevices. The platform integrates a MEMS-PUEH, a high-efficiency power management system, an energy storage element, and representative functional electronics. We first investigate the effect of backside cavity boundary conditions on MEMS-PUEH performance and show that a sealed air-filled chamber significantly outperforms an open water-filled cavity, yielding a 46% increase in root-mean-square output voltage and a 117% increase in average output power across a 2 k{Omega} resistive load under identical incident acoustic intensity at the respective optimal operating frequencies. We then demonstrate system-level integration and characterization. In a tissue-mimicking phantom, under an incident acoustic intensity of approximately 257 mW/cm2, the device charges an 11.5 mF supercapacitor, a 5 {micro}Ah solid-state microbattery, and a 100 {micro}F capacitor to their nominal voltages in less than 5 min, 3 min, and 20 s, respectively. Finally, in vivo validation demonstrates fully autonomous operation of representative functional electronics following ultrasonic charging of the onboard energy storage element. These results establish a highly miniaturized and fully integrated ultrasonic energy-harvesting platform that advances MEMS-based power solutions for implantable biomedical microdevices.

Bioxolography enables high-resolution fabrication of geometrically complex, cell-laden constructs for tissue engineering. However, tissue-relevant cell densities conflict with the optical transparency required for efficient dual-color volumetric printing. In this work, we extend the Bioxolography toolbox to include refractive index (RI) matching for cell-laden bioresins using iodixanol (IDX). Remarkably, IDX enhances optical transparency and boosts reactivity -- a phenomenon unique to Xolography. Yet, excessive IDX compromises dual-color efficiency through increased absorption and undesired UV-only curing, underscoring a central trade-off between optical clarity and photochemical performance. Systematic tuning of resin compositions along an iso-refractive index line demonstrated the versatility of Bioxolography, with IDX enhancing polymerization and 4-Hydroxy-TEMPO providing biocompatible inhibition. Optimizing composition and printing parameters yielded GelMA hydrogels with cell densities up to 5{middle dot}106 cells{middle dot}mL-1. Cell-laden prints achieved sub-100 {micro}m resolution and complex geometries such as channels and gyroids. Using skeletal muscle tissue as a model, we validated RI matched Bioxolography as a promising strategy for tissue engineering by demonstrating cell alignment along printed grooves and formation of mature muscle fibers characterized by MyoHC+ staining and fusion index. By integrating physical, chemical, and biological perspectives, this work advances Xolography toward biomaterials development and reinforces its position as an emerging volumetric (bio)printing technology. Table of Contents O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=175 SRC="FIGDIR/small/729865v1_ufig1.gif" ALT="Figure 1"> View larger version (86K): org.highwire.dtl.DTLVardef@407d86org.highwire.dtl.DTLVardef@1f116b4org.highwire.dtl.DTLVardef@109a437org.highwire.dtl.DTLVardef@1fc4ece_HPS_FORMAT_FIGEXP M_FIG C_FIG For printing higher cell density bioresins with Xolography, iodixanol (IDX) is added for refractive index-matching. The addition leads to an unexpected additional effect with increased reactivity in the dual-color photopolymerization. With careful adjustment of the resin composition and the printing parameters, Bioxolography is proven as a viable tool for tissue engineering.

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.

Four-dimensional (4D) materials incorporating functional gradient designs offer a powerful platform for engineering dynamic structures capable of programmed shape transformations in response to environmental stimuli. However, most gradient-based 4D systems rely on uniaxial gradients, which typically generate simple, symmetric deformations with uniform curvature, limiting their ability to recreate biomimetic architectures that require spatially coordinated morphogenesis. Here, we report a biaxial gradient-engineered 4D hydrogel system capable of programmable, non-uniform shape morphing within a single construct. A one-step photocrosslinking strategy integrates vertical light attenuation and horizontal grayscale photomask patterning to establish orthogonal crosslinking gradients along two directions, producing spatially heterogeneous swelling stresses that drive controlled multi-directional deformation. The resulting hydrogels exhibit tunable swelling and mechanical properties, enabling precise regulation of curvature distribution and shape transformation. This biaxial gradient platform generates diverse biomimetic architectures, including swan-neck, fiddlehead fern, sea star, and Euonymus europaeus-like structures. Importantly, the system supports cell-laden biofabrication, where human mesenchymal stem cell-encapsulated constructs maintain high viability and undergo chondrogenic differentiation while preserving programmed morphologies. This work establishes biaxial gradient-programmed 4D hydrogels as a robust strategy for integrating morphogenesis with tissue formation, advancing biomimetic biofabrication and morphogenetic tissue engineering.

Controlled biomineralization is central to engineering physiologically relevant hard-tissue models, yet achieving spatially organized, three-dimensional (3D) mineral deposition in microfluidic on-chip systems remains challenging. While cell-based bone-on-chip platforms offer biological complexity, they intrinsically couple mineral initiation to confounding factors such as matrix remodeling and paracrine signaling, obscuring the earliest biochemical drivers of nucleation. Drawing inspiration from bottom-up synthetic biology, we engineered an enzymatically active granular matrix that recapitulates a key osteogenic function within a perfusable 3D microenvironment. Alkaline phosphatase (ALP), the key driver of native bone formation, was covalently conjugated to poly(ethylene glycol)-based (PEG) microgels via thiol-ene photochemistry, retaining over 90% enzymatic activity after 48 h. These monodisperse microgels were assembled into a jammed, perfusable matrix within an on-chip chamber, enabling independent control over enzyme loading and substrate delivery. The system supported rapid in situ mineralization (24-48 h), yielding a carbonated, calcium-deficient, apatite-like phase characteristic of early-stage bone mineral. We demonstrate that the spatial 3D localization of enzymatic activity to discrete microscale compartments, coupled with interstitial perfusion, enables localized and near-physiological mineral formation. This mechanistically defined, acellular platform provides a programmable foundation for investigating ALP-driven 3D mineralization and establishes a modular route toward hybrid biosynthetic models of (patho)physiological tissue mineralization.

Automation of organoid and cell culture processes is essential for achieving scalable and standardized experimentation in regenerative medicine and stem cell research. However, existing microfluidic platforms often rely on complex setups, limiting their integration within standard incubator environments. To address these challenges, we developed a compact, scalable multi-well platform featuring 3D-printed, servo-actuated disposable microvalves for fully automated media and drug exchange. This design eliminates the need for external pressure sources and control channels, providing a simplified and cost-effective solution for organoid culture. The platform integrates an internet-connected microscopy module with a motorized XYZ stage, allowing continuous, real-time imaging of individual wells directly within the incubator. It supports precise and reliable fluid handling under physiological conditions, improving throughput, reproducibility, and accessibility. We validate the platform through bench-top testing and in both mouse and human organoid models. Morphological analysis, immunohistochemistry (IHC), and qPCR demonstrate comparable viability, growth, and gene expression profiles between automated and manual culture conditions. These results establish a robust and scalable framework for fully automated organoid culture, offering a simplified and accessible alternative to conventional microfluidic systems with broad applications in regenerative medicine, drug discovery, and scalable biological screening. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=101 SRC="FIGDIR/small/732526v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@5efd07org.highwire.dtl.DTLVardef@3600d0org.highwire.dtl.DTLVardef@16f85f5org.highwire.dtl.DTLVardef@c39fbd_HPS_FORMAT_FIGEXP M_FIG C_FIG

Conducting polymers are widely used in bioelectronic interfaces because of their mixed ionic-electronic conductivity, mechanical compliance, and compatibility with biological systems. However, their electrochemically driven structural dynamics have received little attention as a mechanism for mechanical cell stimulation. Here, we show that electrochemical actuation of poly(3,4-ethylenedioxythiophene):polystyrenesulfonate (PEDOT:PSS) generates mechanical cues capable of activating endogenous mechanosensitive pathways in HEK293T cells. Transparent PEDOT:PSS films deposited on ITO exhibited a heterogeneous granular morphology and underwent potential-dependent microscopic deformation during electrochemical modulation. Direct optical tracking revealed displacement of the polymer boundary, with structural changes occurring preferentially in polymer-dense regions and propagating toward the film edge. When HEK293T cells were cultured directly on PEDOT:PSS, repeated electrochemical stimulation at -240 mV produced reproducible intracellular Ca2+ responses. Pharmacological inhibition with GsMTx4 attenuated the calcium response, whereas blockade of voltage-gated sodium channels with tetrodotoxin largely preserved it, supporting the involvement of mechanosensitive pathways in the cellular response. These findings identify PEDOT:PSS as an electromechanical biointerface in which electrochemical modulation can introduce a mechanical component alongside the established electrical function of the interface. This mechanical contribution should therefore be considered when interpreting cellular responses to conducting polymer- based electrical stimulation and provides a basis for engineering bioelectronic interfaces that deliberately couple electrical control with mechanotransduction.

Intracranial aneurysm (IA) rupture is catastrophic, yet current models of rupture-risk inadequately capture underlying IA remodelling mechanisms. Endothelial-haemodynamic interactions are central to these processes, but in vitro flow platforms often lack vessel-relevant geometry or long-term perfusion. Here, temporal and spatial endothelial responses to haemodynamic stress were investigated across idealised and patient-specific vascular models. Polydimethylsiloxane models were endothelialised with human aortic endothelial cells then perfused at up to 1.6 Pa wall shear stress for five days. IA models were exposed to steady or cardiovascular flow waveforms, with endothelial phenotype assessed by immunofluorescence and cytokine profiling. Flow initiation induced a transient inflammatory response, with elevated MCP-1 and TNF- at day two, followed by a resolution of cytokine levels by day five, including a [~]7.5-fold reduction in MCP-1, despite increased haemodynamic loading. Endothelial cells retained a cobblestone-like morphology with eNOS undetected, resembling a partially activated phenotype. Compared with steady flow, cardiovascular flow reduced TGF-{beta}1 and IL-8 secretion and decreased FGF-b consumption ([~]2.5 fold), suggesting enhanced phenotypic stability. This study presents the first in vitro IA model incorporating a cardiovascular flow waveform and identifies cytokine signatures with potential utility as biomarkers of IA remodelling, highlighting the importance of long-term perfusion for modelling chronic vascular disease. Table of Contents FigureAn in vitro model of an intracranial aneurysm was developed to investigate how fluid flow dynamics impact endothelial remodelling and inflammation. Pulsatile cardiac flow promoted stabilisation of inflammatory signalling, which was sustained under a steady flow regime. Cytokine signatures emerged with potential utility as biomarkers of IA remodelling, highlighting the importance of long-term perfusion for modelling chronic vascular disease. The schematic of the cytokine release dynamics used in the graphical abstract below was generated with the assistance of AI-based tools including ChatGPT (v5.5) and M365 Copilot to align with key results from this manuscript. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=182 SRC="FIGDIR/small/733289v1_ufig1.gif" ALT="Figure 1000"> View larger version (80K): org.highwire.dtl.DTLVardef@c00d12org.highwire.dtl.DTLVardef@9a3afaorg.highwire.dtl.DTLVardef@1961b66org.highwire.dtl.DTLVardef@1e0fec5_HPS_FORMAT_FIGEXP M_FIG C_FIG

Synthetic grafts remain ineffective for small-caliber vascular applications due to thrombosis and intimal hyperplasia. To address these limitations, our lab designed a multilayer graft consisting of a hydrogel coating that promotes post-implantation endothelialization and an electrospun mesh that matches arterial mechanical properties. Damage-resistant hydrogels were engineered using a double-network system composed of polyether urethane diacrylamide and N-acryloyl glycinamide to enhance fracture resistance through hydrogen bonding. In this study, we utilized redox initiation to apply conformal, durable hydrogels to electrospun grafts. Bioactivity wa introduced using streptococcal collagen-like proteins containing 1{beta}1 and 2{beta}1 integrin-binding motifs, enabling selective cell-material interactions that support endothelialization while preserving acute thromboresistance. To establish the feasibility of these grafts as off-the-shelf devices, we evaluated coating integrity and bioactivity retention following sterilization and dynamic physiological loading. Sterilized composites exhibited surgically-associated damage resistance, indicating that sterilization did not compromise hydrogel durability. Coating integrity and bioactivity were also preserved after six weeks of physiological loading. Acut thromboresistance was supported by both static platelet adhesion assays and dynamic whole-blood bioreactor studies using heparinized blood, with low platelet adhesion observed relative to ePTFE. Finally, a pilot ovine carotid model demonstrated successful surgical handling and sustained graft patency. Collectively, these results highlight the promise of multilayer vascular grafts as durable, thromboresistant conduits for small-diameter vascular applications. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=60 SRC="FIGDIR/small/741222v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@14ec910org.highwire.dtl.DTLVardef@17259e8org.highwire.dtl.DTLVardef@6cb611org.highwire.dtl.DTLVardef@1251d7d_HPS_FORMAT_FIGEXP M_FIG C_FIG

Embedded extrusion printing can process collagen-rich bioinks, but their low viscosity and slow fibrillogenesis compromise print fidelity and post-deposition stability. Here, we developed a collagen fibril-inducing support bath (FIB) that combines mechanical support for embedded printing with biochemical induction of collagen assembly. Microfibrillated or nanofibrillated cellulose was incorporated into a fibril-inducing buffer, and formulations were screened at 37 degrees Celsius for rheological behaviour and optical transparency. The selected FIB was evaluated by printing 1% and 5% (w/v) articular cartilage-derived extracellular matrix (ECM) inks at 10-20 mm/s and compared with a cellulose-only control bath. FIB exhibited yield-stress, shear-thinning and rapid recovery behaviour that supported reproducible filament deposition. Unlike the control bath, FIB enabled intact construct retrieval following stabilisation and promoted the formation of fibrillar collagen within the printed strands. Scanning electron microscopy revealed D-banded collagen fibrils preferentially oriented along the deposition direction, with dominant orientation peaks within +/- 10-15 degrees. The platform supported the fabrication of 15 x 15 x 1.5 mm sheets and 6 x 6 x 6 mm scaffolds whose macroscopic dimensions were retained after processing. Constructs produced from 5% ECM inks exhibited approximately fourfold higher ramp and relaxation moduli than those produced from 1% ECM inks. Extracts from both formulations caused no detectable reduction in cell metabolic activity after 24 h or 72 h. Mesenchymal stem/stromal cells (MSCs) seeded onto printed sheets became markedly elongated and aligned by day 3, with approximately 80% of cells having an aspect ratio exceeding 1.5, significantly greater than cells seeded onto casted ECM controls, with a mean deviation of ~9 degrees from the filament print direction. These findings establish FIB as a bioactive support bath that couples embedded printability with collagen fibrillogenesis, enabling recoverable collagen-rich constructs with aligned fibrillar architecture that directs early cellular organisation.

Proper alignment of cells is crucial for functioning of various tissues such as skeletal muscle tissues, neural cells, adipose-derived stem cells, etc. Current in-vitro fabrication methods to replicate the cellular environment, e.g., photolithography and 3D printing, are not cost-effective and cannot capture the complexity of the surfaces to which these cells are exposed to. In this work, we used bio-mimicked leaf templates to closely resemble the in-vivo environment the muscle cells and cultured C2C12 cells, myoblast cell lines, on modified PDMS substrates fabricated using these leaf templates. Using image analysis software, we analyzed the degree of alignment of cells, aspect ratio and the area projected by individual cells cultured on these surfaces. The C2C12 cells cultured on the PDMS substrates formed utilizing the front and back sides of the leaves of Musaceae Banana were found to have an Aspect Ratio of 6.3 and 8.3, the highest among the surfaces studied in this paper. C2C12 cells cultured had the highest degree of alignment on the negative replica of the back side of Dracaena Sanderiana. Due to the availability of a wide range of leaf templates and bio-mimicked surface structures to measure cell response, it is difficult to find the optimal design. Hence, we have also tried to create a catalog using 15 different leaf surfaces and characterized these surfaces into various categories based on the grooves on the surfaces to provide a more comprehensive set of surface designs for studying cell behavior. To quantitatively analyze the groove pattern, 2D FFT analysis was also performed to find the dominant wavelength of the grooves. In surface characterization, hydrophobicity is also a parameter that needs to be considered; hence, the water contact angle of these surfaces was also measured. Our findings highlight the importance of surface topography and hydrophobicity in influencing cell alignment and can contribute to developing biomimetic surfaces for tissue engineering applications.