Bioengineering & Translational Medicine

Precision-cut lung slices (PCLS) retain the native cells and extracellular matrix that contribute to the structural and functional integrity of lung tissue. This technique enables the study of cell-matrix interactions and is particularly useful for pre-clinical pharmacological studies. More specifically, PCLS are widely used to model the complex pathophysiology of pulmonary fibrosis, an uncurable and progressive interstitial lung disease. Current ex vivo pulmonary fibrosis models expose PCLS to pro-fibrotic biochemical cues over a short timeframe (hours to days) and quickly collect samples for analysis due to viability concerns. This condensed timeline is a limitation to understanding chronic disease mechanisms. To extend the utility of ex vivo pulmonary fibrosis models, PCLS were embedded in engineered hydrogels and exposed to pro-fibrotic biochemical and biophysical cues. Hydrogel-embedded PCLS maintained greater than 80% total cell viability over 3 weeks in culture. Gene expression patterns in samples exposed to pro-fibrotic cues matched trends measured in human fibrotic lung tissue. Finally, treatment with Nintedanib, a Food and Drug Administration approved pulmonary fibrosis drug, moderately reduced fibroblast activation and influenced epithelial cell differentiation. Collectively, these results show that hydrogel-embedded PCLS models of pulmonary fibrosis extend our ability to study fibrotic processes ex vivo and, when applied to human tissues, present a new approach methodology for studying lung disease and treatment.

Pancreatic ductal adenocarcinoma (PDAC) remains difficult to treat with nucleic acid therapeutics because efficient intratumoral delivery is limited and off-target liver accumulation is common. Here, we developed a structure-activity map for intraperitoneally administered mRNA lipid nanoparticles (mRNA-LNPs) to identify formulation features that improve delivery to pancreatic tumors while reducing liver expression. A full-factorial library of 48 mRNA-LNP formulations was generated by varying ionizable lipid, sterol, phospholipid, and PEG-lipid components. Formulations were characterized for size, polydispersity, zeta potential, and encapsulation, then evaluated in an orthotopic KPC8060 pancreatic tumor model after intraperitoneal administration of firefly luciferase mRNA-loaded LNPs. Biodistribution was assessed by Rhodamine B fluorescence and functional delivery by luciferase expression 12 h after dosing. Lipid composition strongly influenced both physicochemical properties and in vivo performance. G0-C14-based formulations produced the smallest and most homogeneous particles, whereas FTT5-containing formulations were generally larger. Across the 48-formulation library, mRNA expression and nanoparticle biodistribution varied significantly among tumor, pancreas, liver, and spleen. Statistical, decision-tree, and predictive modeling analyses identified composition rules associated with organ-selective delivery. High tumor expression was associated primarily with G0-C14 combined with DSPC and {beta}-sitosterol, whereas liver expression was favored by C12-200 or DLin-MC3-DMA with DOPE and DSPE-PEG. Notably, a G0-C14/DSPC/DSPE-PEG formulation emerged as a lead candidate, producing a greater than 6-fold increase in tumor luciferase signal relative to the library median while reducing liver exposure by approximately 60%. Histopathology showed no treatment-related liver or lung toxicity. These findings define actionable formulation rules for tuning intraperitoneal mRNA-LNP delivery in PDAC and support further development of tumor-selective mRNA therapeutics for pancreatic cancer.

Intratumoral delivery of immunotherapy offers a means to enhance efficacy while limiting systemic toxicity, yet rapid diffusion from the tumor constrains dosing levels. Extracellular matrix-targeted anchoring strategies have emerged to improve tumor retention, but the influence of matrix target choice remains poorly understood. Here, we engineered a hyaluronic acid-anchoring platform and directly compared it to a well-established collagen-binding strategy for the delivery of IL-12/IL-15 combination therapy, assessing pharmacokinetic, efficacy, and toxicity endpoints. Hyaluronic acid anchoring markedly enhanced intratumoral retention and tumor loading relative to both unanchored and collagen-anchored constructs. While all anchored cytokine therapies achieved comparable curative tumor control, hyaluronic acid anchoring was associated with improved tolerability, including attenuated systemic inflammation, reduced liver toxicity, and diminished local tissue damage. Analysis of intratumoral immune signaling further indicated that the anchoring strategy modulates local cytokine exposure and immune cell infiltration, despite similar therapeutic outcomes. These findings demonstrate that extracellular matrix target selection significantly shapes the pharmacologic and safety profiles of intratumoral biologics, and identify hyaluronic acid anchoring as an alternative retention strategy with potential advantages.

In vitro models are increasingly critical for interrogating cancer biology and therapeutic response, however, accurately recapitulating the tumor microenvironment (TME) remains a persistent challenge, particularly in head and neck cancers (HNC) characterized by complex cell-matrix interactions and heterogeneity. Current models often lack independent tunability of biochemical and biophysical cues, limiting systematic investigation of microenvironmental cues in a high-throughput format. Here, we establish a 3D droplet-based bioprinting platform for the fabrication of customizable in vitro TME models using poly(ethylene glycol) (PEG) hydrogels. Human HNC cell lines (FaDu and 2A3) with differing HPV statuses were bioprinted into PEG matrices spanning physiologically relevant stiffnesses (0.7-4.8 kPa) and compositions, including non-functionalized PEG and peptide-functionalized PEG (PEGfnc: RGD, YIGSR, CNYYSNS) and cultured for 7 days. Cluster growth, cell viability, and cluster morphology were assessed across multiple time points, matrix compositions, and matrix stiffnesses. Proliferation and endpoint phenotype expression were visualized using confocal microscopy through immunofluorescence. Results indicated enhanced cell viability in PEGfnc matrices, compared to non-functionalized matrices, while effect of matrix stiffness was less prominent. Median cluster size reached 40-50 m by day 7, and linear mixed-effects modeling identified how changes in cluster surface area, volume, and tumoroid complexity varied with cell type, matrix, and stiffness. By decoupling and systematically varying key TME parameters, this approach provides a robust and scalable framework for dissecting tumor-matrix interactions and advancing physiologically relevant in vitro models for cancer research and therapeutic screening.

Extracellular vesicles (EVs) are lipid spheres released from cells. Research utilizing EVs has met several hurdles owing to the small size of the majority of EVs and other nanoparticles (<150 nm) and the lack of detection technologies capable of providing high-throughput single particle measurements at this scale. The use of high-throughput single particle measurements is critical for the assessment of EV heterogeneity and abundance which are features often used to assess the development of isolation protocols or particle characterization. The Coulter principle, known in the field as resistive pulse sensing (RPS), has been used for several decades to size and count cells. More recently, this technology has evolved to accommodate nanoparticle analysis. In the last decade a platform utilizing microfluidic resistive pulse sensing (MRPS) has been demonstrated for nanoparticles, offering ergonomic characterization of nanoparticles along with utilizing open format data. To date, assessment of MRPS accuracy and reporting standards have not been assessed. With the aim of increasing data accuracy, ergonomics, and reporting transparency, we developed a microfluidic resistive pulse sensing post-acquisition analysis software (RPSPASS) application for automated cohort calibration, population gating, statistical output, QC plot generation, alternative data file outputs, and standardized reporting templates.

Medulloblastoma (MB) is an aggressive central nervous system (CNS) malignancy that primarily affects children and frequently exhibits metastasis to the leptomeninges of the brain and spinal cord. We developed a {beta}-Cyclodextrin-poly({beta}-Amino Ester) nanoparticle system to deliver the histone deactylase inhibitor (HDACi) Panobinostat to MB by the intrathecal route. Various imaging methods were utilized to study nanoparticle and payload fate following infusion into the cerebrospinal fluid (CSF) of mice via cisterna magna or lumbar access points. Nanoparticles dramatically improved penetration of hydrophobic small molecules into distal regions of the spinal cord. Panobinostat-loaded nanoparticles were effective at treating patient-derived MB, activating pharmacodynamic targets, slowing growth of the primary tumor, decreasing incidence of metastasis at the time of death, and ultimately prolonging survival. These studies provide insight into the mechanisms mediating transport of colloids and therapeutic molecules in the subarachnoid space and highlight new approaches for treating metastatic disease in the CNS.

Scaling up microvessel culture systems is essential for producing vascularized clinically relevant tissues, yet current platforms offer little guidance on how to preserve flow conditions during scale-up. Here, we present a computational-experimental framework using computational fluid dynamics (CFD) to guide the design and scaling of microvessel bioreactors. Interstitial flow distributions were pre-dicted in two perfusion-based platforms-a permeable insert and a rhomboidal microfluidic chamber-across multiple scaling factors and hydrostatic pressures. CFD identified IF ranges conducive to vascu-logenesis and quantified how geometry and pressure modulate flow uniformity. Scaled-up bioreactors generated microvessel networks with consistent morphology and connectivity over a 30-fold increase in culture volume, confirming that maintaining equivalent IF ensures reproducible outcomes. The permeable insert platform maintained uniform IF across scales, while the rhomboidal chamber produced spatially varying IF resulting in heterogeneous but physiologically relevant networks. These findings establish CFD as a predictive tool for rationally scaling perfusion bioreactors, enabling microvessel production at clinically relevant scales with controllable morphology. Significance StatementScaling up microvessel bioreactors is critical for engineering large pre-vascularized tissues. However, larger scales may disrupt flow conditions that drive vessel formation. This study demonstrates that computational fluid dynamics (CFD) can predict interstitial flow and guide the rational scale-up while preserving the vasculogenic microenvironment. Experiments across 30+-fold size increase confirmed that matching inter-stitial flow results in morphologically identical microvessel networks. By linking simulation-based design with experimental validation, this work establishes CFD as design tool for scalable perfusion bioreactors for production of microvessel networks at clinically relevant scales.

Non-front-fanged snakes are abundant, diverse and represent approximately 70% of extant snakes. However, there is limited knowledge about most species and their venoms, in part due to the technical and welfare challenges associated with venom extraction, low venom yields, and the lack of cellular models available. Organoids represent an excellent opportunity to overcome these challenges. Here, we establish, for the first time, venom gland organoids from snakes of the Colubridae family and demonstrate the in vitro production of toxins.

Respiratory infectious diseases are among the leading causes of global morbidity and mortality and remain a major public health concern. Progress in understanding early host-pathogen interactions has been hampered by the limited physiological relevance of existing experimental systems. Different models mimicking the human respiratory epithelium have been developed to study viral infections in vitro, such as tridimensional (3D) tissue models and organoids. However, many lack key features of human tissue architecture, particularly the lamina propria or immune cells. To address these limitations, we established an immunocompetent 3D model of the human respiratory mucosa by combining nasal epithelial cells isolated from nasal brushings, fibroblasts from mid-turbinate nasal biopsies, and macrophages derived from blood monocytes. These cells were sequentially seeded into collagen-chitosan scaffolds, resulting in a reconstructed respiratory mucosa closely resembling the in vivo nasal tissues. To further confirm the physiological relevance of the model, we infected it with influenza A virus. The mucosa model supported viral replication in the epithelium and consequently showed increased secretion of inflammatory cytokines and upregulation of type I interferon related genes, enabling the monitoring of early antiviral innate immune responses in a physiologically relevant context.

Transforming growth factor-beta (TGF-{beta}), a potent promoter of extracellular matrix deposition and suppressor of infiltrating immunity, has arisen as an attractive target for improving outcomes in tissue fibrosis and cancer immune therapy. Despite the promise of TGF-{beta} inhibitors for attenuating the progression of fibrotic disorders or as adjuncts for cancer immunotherapy, current systemically administered inhibitors that target the ligand or receptors have significant on-target liabilities, including cardiotoxicity and development of pre-malignant cutaneous squamous lesions. Recently, an engineered mini monomer of TGF-{beta} (mmTGF-{beta}), which potently and specifically inhibits TGF-{beta} activity, was shown to strongly synergize with checkpoint inhibitors to suppress cancer progression in an aggressive model of melanoma when genetically delivered using an engineered form of vaccinia virus that preferentially infects cancer cells. Despite these promising results, however, a significant fraction of the mmTGF-{beta} was found to misfold, likely due to mispairing of the cysteines that comprise its cystine knot. Here, we demonstrate that inclusion of a modified form of the TGF-{beta} pro-domain that lacks its dimerization motif, the bowtie knot, dramatically improves both the folding and inhibitory activity upon secretion by mammalian cells, thus overcoming one of the major limitations of genetically delivering mmTGF-{beta}. Furthermore, we show that fusion of mmTGF-{beta} to a CD44 binding domain enhances the inhibitory potential of mmTGF-{beta} on immune cells, and on other cell types which express CD44, by more than 30-fold compared to cells negative for CD44. Together, these modifications provide a framework for further enhancing the efficacy and safety of mmTGF-{beta} for cancer immune therapy, and possibly also tissue fibrosis, when delivered genetically using vaccinia, or other related approaches.

Pluripotent stem cells represent a potentially unlimited cell source for the fabrication of human bioartificial tissues to study and treat degenerative conditions such as type 1 diabetes. Alginate is widely used for mammalian cell immobilization and the primary hydrogel studied for pancreatic islet encapsulation. Rheological properties of alginate solutions or fully gelled forms are unsuitable as support matrix for embedded 3D printing. We describe partially gelled self-healing alginate formulations tuned for embedded 3D printing. Perfusable multi-plane hierarchical networks branching into 10 parallel channels, obtained by 3D printing of Pluronic F127 into the alginate support, show high fidelity to computer-assisted models. Therapeutic {beta}-cell doses (40x106 cells/mL) within centimeter-thick perfusable constructs remained viable for at least 1 week of culture under flow, with rapid insulin secretion detected upon glucose challenges. Stem cell-derived islet clusters cultured in 5-channel contructs for 25 days differentiated towards functional insulin-expressing cells. We describe a novel approach to generate cm-scale perfusable endocrine pancreatic constructs using sacrificial embedded 3D printing into alginate. This approach offers an adaptable platform to engineer perfusable cm-scale functional endocrine pancreatic tissues and potentially other vascularized bioartificial tissues.

Small airways are the primary sites of airflow obstruction in chronic obstructive pulmonary disease. Effective delivery of aerosolized drug particles to these regions is crucial to maximize treatment efficacy while minimizing side effects. However, conventional inhalation therapy approaches (i.e., full-mouth particle release and inhalation (FMD)) typically result in insufficient drug deposition in the small airways and an uneven distribution across the five lung lobes. To address such deficiencies, the goals of this study are triple folds: (1) to develop a fast and accurate framework to secure target drug delivery (TDD) nozzle diameter and location based on the conventional computational fluid particle dynamics (CFPD)-FMD simulations, (2) to develop a CFPD-informed machine learning (ML) inverse-design framework that predicts optimal inhaler nozzle parameters based on patient-specific breathing patterns and drug properties, and (3) to demonstrate the feasibility of embedding this framework into a user-centered smart inhaler prototype to improve uniform TTD to the small airways across all five lung lobes. Specifically, a subject-specific mouth-to-generation-10 human respiratory system was employed, and 108 high-fidelity CFPD-FMD simulations were performed under varied physiological and design parameters, including tidal volume, particle diameter, release location, and release timing. Particle release maps generated from those CFPD-FMD simulations via backtracking identified optimal nozzle diameters and locations that promote uniform multi-lobe drug delivery while limiting off-target deposition. Accordingly, a dataset was compiled with inputs (i.e., flow rate, particle size, release z-coordinate, release time) and targets (i.e., nozzle center x- and y-coordinates, nozzle diameter). These inputs and targets form the CFPD-TDD dataset, on which 16 ML models were trained to learn inverse mapping from patient- and drug-specific inputs to optimal nozzle design parameters. Performance was evaluated using mean squared error (MSE) and mean absolute error (MAE) overall and per target feature. Parametric analysis using CFPD-FMD simulations was conducted to determine how patient-specific and drug-specific factors affect pulmonary air-particle transport dynamics and to explain why achieving CFPD-TDD in small airways with CFPD-FMD strategies remains challenging. Furthermore, the ML evaluation in this feasibility study demonstrated robust learning of the inverse mapping from patient-specific inputs to optimal nozzle parameters. Four top-performing models showed consistently low MSE/MAE across cases, and an ensemble (i.e., mixed model (MixModel)) combining their strengths was formulated. Independent CFPD-TDD simulations beyond the training and testing datasets were used as the ground truth to validate ML-predicted nozzle configurations. Compared with conventional CFPD-FMD strategies, ML-guided nozzle designs significantly improved inter-lobar deposition uniformity and reduced off-target deposition in the upper airways, demonstrating the feasibility of ML-enabled TDD to the small airways. Overall, this study establishes a CFPD-informed ML inverse-design framework as a viable algorithmic foundation for user-centered smart inhalers, enabling adaptive, patient-specific TDD to the small airways with improved deposition uniformity across all five lung lobes. By integrating first-principle-based CFPD with ML, this work provides a methodological pathway toward next-generation smart inhalers for more effective treatment of small airway diseases.

Understanding how airborne particulates disrupt the human alveolar barrier requires in vitro systems that accurately replicate its composition and function. We present a biodegradable lung alveoli-on-a-chip that reproduces the architecture and physiology of the human air-blood interface using a porous poly(lactic-co-glycolic acid) (PLGA) membrane positioned between epithelium and endothelium under air-liquid interface (ALI) culture. The membrane, fabricated by porogen-assisted nonsolvent-induced phase separation, exhibited >50 % porosity, [~]2 {micro}m thickness, and mechanical compliance over 100-fold higher than conventional Transwell inserts, closely resembling the native interstitium. During co-culture, gradual PLGA degradation was compensated by cell-secreted extracellular-matrix (ECM) proteins such as collagen IV and laminin, forming a self-remodeling barrier that maintained integrity for at least 11 days. The platform supported stable epithelial-endothelial co-culture, high transepithelial electrical resistance, and physiologically relevant permeability. To demonstrate its utility, the chip was used to assess pulmonary toxicity of four types of waste-combustion-derived particulates, including rubber, plastic bags, plastic bottles, and textile fibers, delivered apically under ALI conditions. All combustion products reduced cell viability, increased hydrogen-peroxide release, and elevated {gamma}-H2AX expression, indicating oxidative and genotoxic stress, while disrupting barrier permeability. Rubber combustion particles elicited the most severe toxicity, causing the greatest loss of viability, accumulation of reactive oxygen species, and formation of DNA double-strand breaks. Together, these results establish a biodegradable, ECM-remodeling lung alveoli-on-a-chip as a physiologically relevant platform for investigating source-specific particulate toxicity and alveolar-barrier pathophysiology. By bridging environmental exposure models with human-relevant lung biology, this system provides a quantitative and translatable tool for evaluating respiratory risks and therapeutic interventions.

Cochlear implants partially restore sound sensation for individuals with severe-to-profound hearing loss. This can significantly improve the recipients quality of life, largely through improved communication with spoken language. However, surgical trauma from electrode insertion also instigates biological responses that can compromise device performance and longevity. To address these limitations, we designed a novel polymer-based coating that provides controlled release of selonsertib, an apoptosis signal-regulating kinase 1 (ASK1) inhibitor with well described anti-inflammatory, anti-fibrosis and cytoprotective properties. We describe the physical and molecular evaluation of this coating, including the application of next generation mass spectrometry-based proteomics in mammalian cochlear explants and their culture medium. These analyses provide new insights into the acute tissue response in the damaged cochlea and demonstrate that selonsertib eluted from the polymer coating has a significant pro-survival, anti-inflammatory, and anti-fibrosis effect. This study establishes proof-of-concept for selonsertib-eluting polymers, demonstrating both feasibility for targeted drug delivery and efficacy for mitigating acute cochlear damage responses.

Central nervous system (CNS) disease or injury might be treated by implanted devices, tissue regenerative scaffolds, or drug delivery platforms. However, inflammatory CNS responses limit these interventions and may worsen outcomes following damage to the CNS. Via the foreign body reaction (FBR), macrophages and glial cells trigger a "glial scar" around implants, reducing device performance, scaffold regenerative ability, or drug delivery potential. Previous studies have shown that stiffness of CNS implants significantly affects glial encapsulation, but few studies have investigated materials that truly match brain tissue stiffness. Porous precision-templated scaffolds (PTS) with uniform, interconnected, 40 {micro}m pores have shown favorable healing outcomes and a reduced FBR in numerous soft and hard tissue applications. To quantify the effects of both hydrogel compliance (stiffness) and pore size on glial encapsulation, we implanted poly(2-hydroxyethyl methacrylate-co-glycerol methacrylate) (pHEMA/GMA) PTS of varying stiffness and pore size for 4 weeks in rat brain. We observed reduced astrocyte encapsulation around PTS compared to solid hydrogel rods, reduced pro-inflammatory macrophage polarization for softer hydrogels versus stiffer hydrogels, and the presence of neuronal markers and neurogenesis within the pores. Utilizing soft, precision-porous hydrogels could provide a strategy for mitigating glial scarring and improving implant-based CNS treatments.

Clinical outcomes in aggressive breast cancer vary widely, in part because the tumor microenvironment is structured to exclude immune infiltration. Low antigen load, dysfunctional antigen-presenting cells, T cell exclusion and exhaustion, and a stiff extracellular matrix that physically restricts immune cell trafficking work together to form a suppressive barrier that current immunotherapies struggle to overcome. We addressed this barrier using ultrasound (US)-activated nanobubbles (NBs), a drug-free intervention based on perfluoropropane-filled nanoparticles. The size and deformable phospholipid shell enable NBs to achieve deep tumor penetration and a uniform distribution throughout the entire tumor. Upon ultrasound activation, NBs generate localized mechanical forces that restore extracellular matrix elasticity, disrupt tumor transport barriers, and drive HMGB1 release, re-engaging endogenous antitumor immunity without pharmacological agents. In a syngeneic triple-negative breast cancer model, US-NB treatment depleted immunosuppressive myeloid cells 3-fold within 3 hours, followed by a greater than 5-fold increase in the ratio of antigen-experienced to suppressive T cells at 48 hours. US-NB drives rapid infiltration of CD4+ and CD8+ T cells within 48 hours. US-NB treatment achieved an 85% cure rate in the D2A1 model; cured animals maintained durable systemic immune memory, rejecting both local and systemic tumor rechallenge. Consistent therapeutic benefit was observed in a luminal B-like mammary tumor model (E0771), supporting activity across breast cancer subtypes. These results establish US-NB mechanical immunomodulation as a drug-free therapeutic strategy capable of generating robust and durable antitumor immunity, acting through biophysical tissue properties rather than tumor-specific molecular targets. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=90 SRC="FIGDIR/small/714247v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@b1ed5forg.highwire.dtl.DTLVardef@1572a98org.highwire.dtl.DTLVardef@1ad6906org.highwire.dtl.DTLVardef@1ca1b36_HPS_FORMAT_FIGEXP M_FIG C_FIG

Nanoplastics generated from plastic waste in our ecosystems are becoming increasingly prevalent as bulk plastics exposed to natural factors like water and sunlight fragment to the nanoscale over time. These incidental nanoplastics span a wide range of physicochemical properties, which makes studying nanoplastic interactions in biological systems difficult. Here, we characterized the behavior of incidental nanoplastics generated through mechanical abrasion within coacervate droplets to probe the surface properties of the nanoplastics. We used elastin-like polypeptides (ELPs) to create hydrophobic or charged coacervate microenvironments. Using optical microscopy and fluorescence quantification, we observed that nanoplastics made from polyethylene terephthalate (nPET), nylon 6 (nPA), and polystyrene (nPS) exhibited distinct partitioning behavior with more favorable interactions with hydrophobic droplets. This indicated that the hydrophobic polymer backbone was the predominate surface feature despite exposed functional groups of the incidental nanoplastics, in contrast to findings with model carboxylated latex nanospheres (nPS-COOH). Furthermore, the selective partitioning of incidental nanoplastics into the hydrophobic droplets was able to capture over 80% of nPET in solution, and after recovery of the protein droplet, was able to cumulatively capture over 75% of the nPET feedstock across multiple cycles. This work explores the nuanced surface characteristics of incidental nanoplastics, expands the application of coacervates as chemical probes, and demonstrates a biopolymer approach for effective nanoplastic removal.

Autologous cell suspension (ACS)-based therapies are an established strategy to enhance wound repair, yet limitations in preparation workflows and donor skin requirements remain barriers to wider clinical implementation. We have previously developed VeritaCell, a rapid enzymatic disaggregation-based approach that generates highly viable skin cell populations, including epidermal stem cell-enriched fractions, and demonstrated their pro-regenerative biological properties in vitro. Here, we have evaluated the in vivo efficacy of VeritaCell-derived ACS using a rat full-thickness excisional wound model. ACS preparations were applied at donor-to-wound area ratios of 1:1, 1:10, and 1:20, and wound progression was monitored through longitudinal image-based quantification alongside histological assessment of tissue architecture. ACS-treated wounds exhibited enhanced early wound closure dynamics, with significant within-group improvements evident by Day 6. Histological analysis demonstrated improved neo-epithelial organisation and reduced epidermal thickening in the 1:10 and 1:20 groups, with the 1:10 condition showing tissue architecture most closely resembling unwounded skin. Notably, beneficial effects were observed even at low estimated cell numbers, suggesting that cell viability and biological activity may be key determinants of therapeutic efficacy. Collectively, these findings provide in vivo validation of VeritaCell-derived ACS and support the use of biologically informed donor-to-wound coverage ratios. This approach may enable effective wound repair while minimising donor skin requirements, with potential relevance for the treatment of extensive injuries such as burns.

Long-term exposure to high-energy visible (HEV) blue light and infrared-A (IR-A) radiation accelerates oxidative stress, inflammation, and transepidermal water loss (TEWL), leading to photoaging and damage to the skin barrier. In this study, we developed Raybloc(R), a marine bioactive silica microsponge formulation, and evaluated its protective effects against combined high-energy visible (HEV; 410-480 nm) and infrared-A (IR-A; 700-1400 nm) exposure in a preclinical model. We divided 36 nude BALB/c-nu/nu mice into six groups: one that didnt get any treatment, one that got Raybloc(R) (no radiation), one that got Raybloc(R) 5%, one that got Raybloc(R) 8%, one that got HA 0.5%, and one that got HA 0.8%. Animals underwent topical treatment for 14 days under regulated exposure to HEV (410-480 nm, 100 J/cm2/day) and IR-A (700-1400 nm, 30 mW/cm2). We examined transepidermal water loss (TEWL), skin hydration, oxidative stress, inflammatory cytokines (IL-1{beta}, IL-6, TNF-, IL-10), and histological indicators of collagen preservation through biophysical, biochemical, and histopathological techniques. In the Raybloc(R) 8% group, TEWL dropped by 48.3 {+/-} 4.6% (p < 0.001), and skin hydration went up by 62.7 {+/-} 5.1%. The levels of ROS and MMP-1 expression decreased by 63.4% and 57.2%, respectively, while collagen I increased by 2.1 times compared to HA 0.8%. There was a big drop in the pro-inflammatory cytokines IL-1{beta}, IL-6, and TNF- (-54%, -49%, and -46%), and a big rise in IL-10 (+38%). Histological analysis demonstrated well-preserved epidermal integrity and dense collagen bundles in Raybloc(R)-treated mice, whereas irradiated controls exhibited dermal disorganization and inflammatory infiltration. Raybloc(R) showed better photoprotective, antioxidant, and moisturizing effects than HA-based products. It also helped reduce oxidative and inflammatory skin damage caused by blue light and IR-A. These results support Raybloc(R) as a next-generation multifunctional dermocosmetic that can help stop photoaging caused by digital and solar radiation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/713389v1_ufig1.gif" ALT="Figure 1"> View larger version (70K): org.highwire.dtl.DTLVardef@54e046org.highwire.dtl.DTLVardef@502f87org.highwire.dtl.DTLVardef@6088daorg.highwire.dtl.DTLVardef@1b8c241_HPS_FORMAT_FIGEXP M_FIG C_FIG

PurposeLarge quantities of human pluripotent stem cells (hPSCs) are required for clinical applications. 3D suspension cultures are suitable for large scale manufacturing of hPSCs but yield, viability and quality are affected by the hydrodynamic environment. This paper characterizes the hydrodynamic environment inside vertical wheel bioreactors (VWBRs) as a function of size and agitation rates, measures its effect on cell aggregation and proliferation, and proposes the use of Lagrangian-based shear stress and energy dissipation rate (EDR) exposures to support scale-up. MethodsIn silico: Transient, 3D, turbulent flow simulations are conducted for two VWBR sizes (100, 500 mL) at five agitation rates between 20 and 80 rpm. Trajectories of cell aggregates of sizes from 200 to 1,000 microns are calculated, and shear stress and EDR exposures are collected along these trajectories. In vitro: ESI-017 hPSCs were cultured in VWBRs for 6 days. Aggregation efficiency and daily fold ratios were calculated based on cell counts and initial inoculation density. ResultsAggregate size, agitation rate and bioreactor size modulate cell aggregate exposures to EDR and shear stress, which significantly depart from maximum or volume average metrics used for scale-up. Combined in vitro/in silico results show EDR affects aggregation efficiency, cell counts and aggregate size, and has a small effect on daily fold ratios but a significant effect on total fold ratio. ConclusionHistory of trajectory-based cell aggregate exposures to EDRs provide a better scale-up basis for VWBRs than volume-averaged EDR. Shear stress does not significantly affect hPSC aggregation, proliferation and expansion in VWBRs under the tested conditions.