Biomaterials Science

Hydrogels have emerged as attractive vaccine delivery platforms because they enable controlled modulation of antigen availability. However, how different hydrogel environments affect the release and functionality of mRNA-loaded lipid nanoparticles (mRNA-LNPs) remains poorly understood. Here, we investigated the release, stability, cellular uptake, and transfection capability of LNPs released from four hydrogel systems representing distinct crosslinking mechanisms: covalently crosslinked poly(ethylene glycol) (PEG), ionically crosslinked alginate, thermoresponsive Poloxamer 407 (P407), and protein-based Matrigel/collagen hydrogels. All hydrogels enabled release of LNPs over days, with kinetics strongly depending on hydrogel composition and polymer concentration. LNPs were quantitatively recovered from all hydrogel types, except from Matrigel/collagen where incomplete matrix dissolution was the limiting step. Lower polymer concentrations generally accelerated nanoparticle release. PEG offered greatest tunability of release kinetics; at the same time the recovery of the LNP-incorporated fluorescent dye DiI was reduced to about 80 %, indicating partial dye leakage. Alginate hydrogels exhibited recovery of DiI below 50 % and broader particle size distributions after release, while P407 hydrogels largely preserved LNP characteristics. Although quantitative recovery from Matrigel/collagen hydrogels was limited, released LNPs remained readily available for cellular uptake. Notably, LNPs released from low- and intermediate-concentration Matrigel/collagen hydrogels achieved approximately 80-90 % of the eGFP expression compared to mRNA-LNP that were not embedded into a hydrogel. Importantly, cellular uptake and transfection experiments demonstrated that all investigated hydrogels released biologically active mRNA-LNPs capable of mediating protein expression. Moreover, our findings show that hydrogel composition is a critical determinant of mRNA-LNP release, stability, and functional delivery. This work provides design principles for the development of hydrogel-based mRNA delivery systems aimed at sustained antigen availability and prolonged vaccine responses. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/741169v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@138d9eforg.highwire.dtl.DTLVardef@16c0edaorg.highwire.dtl.DTLVardef@1432dd1org.highwire.dtl.DTLVardef@17511b5_HPS_FORMAT_FIGEXP M_FIG C_FIG

Osteoarthritis, characterized by cartilage degradation and synovial inflammation, has spurred interest in mechano-piezoelectric bio-hydrogel therapies that can both relieve symptoms and reverse progression. However, current approaches lack sufficient piezoelectric output and dual cartilage/inflammation targeting. To address this, we demonstrated a mechano-piezoelectric peptide hydrogel composed of nanofibers integrating piezoelectric cues with mesenchymal stromal cells (MSCs) recruitment and PIEZO2 mechanosignaling. Molecularly, the hydrogel's seed peptide incorporated four functions: COL2A1 targeting, MMP-13 responsiveness, MSCs homing, and self-assembly. Overexpressed MMP-13 in the osteoarthritis niche triggers gelation, promoting MSCs recruitment and drug retention. Fluorination modulates hierarchical nanofiber assembly, enhancing mechanical and piezoelectric properties, as confirmed by morphological, biophysical, and computational analysis. The trifluoromethyl-modified, 4-octyl itaconate (4-OI) loaded formulation reverses osteoarthritis via PI3K/AKT activation and Wnt/{beta}-catenin suppression, as shown by improved Osteoarthritis Research Society International (OARSI) scores, bone microarchitecture, and cartilage matrix. This synergy of mechano-piezoelectric cues and 4-OI offers a clinically promising strategy for osteoarthritis.

Macrophages are highly dynamic cells that maintain tissue homeostasis by regulating both initiation and resolution of inflammation. During efferocytosis, macrophages recognize the eat me signal, phosphatidylserine (PS), exposed at the surface of apoptotic cells, leading to the resolution of inflammation and acquisition of a pro-resolving phenotype. Inspired by this endogenous mechanism, PS-based biomaterials have demonstrated immunomodulatory potential. However, the molecular mechanisms underlying PS-mediated macrophage reprogramming remain poorly understood. Here, submicron PS-exposing polymeric particles (PSPs; [~]300 nm) were developed to improve the suitability of PSP formulations for future systemic administration while preserving their immunomodulatory activity. PSPs were efficiently internalized by macrophages through both actin- and dynamin-dependent pathways. PSP treatment significantly reduced IL-6 and IL-12p70 production in LPS-stimulated macrophages, whereas induction of the classical anti-inflammatory M2 marker CD206 was limited. Transcriptomic analysis revealed coordinated attenuation of inflammatory signaling pathways, including downregulation of Myd88, Nfkb1, Rel, and Irf8, together with activation of NRF2-associated antioxidant pathways characterized by increased expression of Nfe2l2, Hmox1, Prdx1, Gclm, and Gclc. Activation of antioxidant-associated genes together with reduced Irf8 expression suggests that PSP promotes inflammatory resolution through coordinated redox adaptation and selective attenuation of inflammatory signaling. Collectively, these findings provide mechanistic insight into PS-mediated macrophage reprogramming and support the future development of systemically administered therapies for chronic inflammatory diseases, including vascular inflammatory disorders. HighlightsO_LISubmicron PSPs retain immunomodulatory activity of apoptotic cell-mimicking biomaterials. C_LIO_LIPSPs are rapidly internalized through actin- and dynamin-dependent pathways. C_LIO_LIPSPs attenuate inflammatory signaling and selectively suppress IL-6 and IL-12p70 production. C_LIO_LIPSPs induce NRF2-associated antioxidant and glutathione responses. C_LIO_LITranscriptomics reveals an early redox-adaptive macrophage program. C_LI

Edible plant derived extracellular vesicles (PDEVs) are emerging as biocompatible, orally deliverable nanocarriers with therapeutic potential; however, their phytometabolite cargo, gastrointestinal stability, and source specific biological functions remain poorly characterized. Here, PDEVs were isolated from four phytochemically distinct plant based foods (black carrot, ginger, garlic, and turmeric), selected for their diverse bioactivity, and characterized by transmission electron microscopy, nanoparticle tracking analysis, and zeta potential. Gastrointestinal stability was evaluated in simulated digestion model. Source specific phytometabolites were profiled by untargeted LC MS MS metabolomics. Functionally validated in ammonia stressed epithelial cells and steatotic hepatocytes. PDEVs exhibited characteristic cup shaped morphology with particle sizes ranging from 60 to 214 nm and zeta potentials of -6.0 to -49.0 mV. PDEVs retained colloidal stability, supporting their suitability for oral delivery. We identified 572 phytometabolites with distinct source specific signatures, including lignin and quercetin in carrot EVs, [6] gingerol and silymarin in ginger EVs, diosgenin in garlic EVs, and curcumin in turmeric EVs. These metabolites found associated to antioxidant, anti inflammatory, epithelial barrier, lipid metabolic, and apoptotic pathways. Functional validation demonstrated carrot EVs significantly enhanced epithelial barrier integrity by increasing claudin (>8-fold, p<0.05), occludin (>2-fold, p<0.05). Ginger EVs restored ZO 1 while suppressing cyclin D1 and MMP9(p<0.05). Garlic and turmeric EVs attenuated inflammatory signaling by reducing STAT3, AKT1, and TNF , whereas turmeric EVs additionally decreased caspase 3 and PTGS2(p<0.01). In steatotic hepatocytes, garlic EVs significantly reduced PNPLA3 (p<0.001) and SREBP 1c while increasing PPAR- (p=0.002). Hence, our results indicate that edible PDEVs are gastrointestinally stable, phytometabolite enriched nanocarriers with distinct source specific functional properties, supporting their potential as orally deliverable nutraceuticals for improving gut liver functions.

Tympanic membrane perforations remain a common clinical problem, and although surgical intervention through tympanoplasty achieves high success rates, it is associated with donor-site morbidity, surgical complexity and limited restoration of the native radial and circumferential collagen architecture. In this study, 3D extrusion printing was utilized to create an active scaffold and attempt to promote collagen organization through shear-mediated structural alignment. An alginate-carboxymethyl cellulose (CMC) hydrogel with bovine SIS-dECM was prepared and investigated for its suitability as a bioink alternative to tympanoplasty grafts. The physiochemical, rheological and printability characteristics of the hydrogel were assessed. Successful decellularization was confirmed by histological analysis. The incorporation of the SIS-dECM into the hydrogel led to increased swelling, lower apparent viscosity, yield stress and flow stress while maintaining favourable printability and filament stability. Polarized optical microscopy was also used to study the influence of printing speed on the alignment of collagen to mimic the native tympanic membrane radial collagen architecture. Compared with the cast controls, the printed samples presented stronger birefringence signals. Biological evaluation demonstrated that the 15% dECM hydrogel exhibited the highest live cell area percentage and live/dead ratio after 48 h. In addition, the chick chorioallantoic membrane assay demonstrated that the dECM-containing hydrogels improved vascular density. The findings establish a printable, biologically active dECM bioink capable of generating bulk collagen organization through extrusion printing as a platform for tympanic membrane regeneration.

Sensory and motor neurons differ significantly in their morphology, structural organization, functional properties, and mode of action. However, despite this heterogeneity, many in vitro studies focus only on a single neuronal subtype, mainly being sensory neurons, limiting the translational potential and relevance of these studies for the evaluation of therapeutic options for spinal cord injury. In this study, we investigate the differentiation, maturation, and neuronal outgrowth of human induced pluripotent stem cell (iPSC)-derived motor and sensory neurospheres using polyethylene glycol (PEG)-microgels with various bioactive coatings. Our results show subtype-specific responses to the PEG-microgel scaffolds, with respect to motor and sensory neurosphere morphology and size. Furthermore, we compare the formation of the PEG-microgel/scaffolds when starting from iPSCs-derived precursor neuron spheres versus undifferentiated iPSCs. We observe notable differences in structural organization, maturity, and neuronal outgrowth between the two approaches, as well as between motor and sensory neurospheres. Together, these results underline the importance of studying motor and sensory neurons separately and highlight the need for a controlled, tunable culture platform to assess the impact of the microenvironment and to improve the physiological relevance of in vitro platforms for neuron-based research.

Precise control of growth factor delivery remains a challenge for directing stem cell differentiation in three-dimensional (3D) engineered tissues. In this study, engineered cell-like vesicles are used as programmable microenvironments to enable sustained and localized delivery of growth factors within visible-light-crosslinked GelMA hydrogels. Giant unilamellar vesicles (GUV) loaded with FGF-2 and TGF-{beta}3 were incorporated into bioinks with BM-MSC to drive keratocyte differentiation without repeated soluble growth factor supplementation. ELISA measurements confirmed the removal of non-encapsulated growth factors and the release of the vesicle cargo following induced vesicle rupture. Fluorescence monitoring showed a progressive reduction in detectable FGF-2- and TGF-{beta}3-loaded GUV during culture, while droplet-scale analysis demonstrated the co-deposition of cells and vesicles after printing. After 14 days of differentiation, differentiated cells expressed ALDH1A1, ALDH3A1, lumican, keratocan, and collagen I without induction of -SMA. Interestingly, keratocyte-associated differentiation was retained after drop-on-demand bioprinting, confirmed by qPCR analysis. These findings establish growth factor-loaded vesicles as bioprintable instructive niches capable of supporting localized keratocyte differentiation within 3D corneal constructs.

Hydrogels are widely used in sensing, delivery, and tissue engineering because their transport properties can be tuned through material design. However, while hydrogel permeability is often characterized using small molecules, many practical applications depend on the uptake and retention of much larger species, including protein conjugates and nanoparticles. Here, we systematically investigate how polyethylene glycol (PEG)-acrylate hydrogel microparticle formulation influences accumulation of signal-generating probes spanning a broad size range. We fabricated particles across a 36-condition design space varying nominal PEG-acrylate molecular weight, polymer weight percent, and UV crosslinking dose, and related formulation-dependent probe accumulation to particle swelling behavior. Increasing nominal PEG-acrylate molecular weight and decreasing polymer weight percent produced more highly swollen particles and strongly enhanced accumulation of fluorescent streptavidin conjugates, with the largest effects observed for bulky labels such as allophycocyanin and phycoerythrin. Gold nanoparticle accumulation was even more formulation-restricted, with detectable colorimetric signal observed primarily in the most permissive formulations. These findings establish design rules linking PEG hydrogel formulation to size-dependent accumulation and show that formulations suitable for small probes may be inadequate for larger reporters. More broadly, this framework may inform the design of hydrogels for particle-based assays as well as other applications where transport of macromolecules or nanoscale materials is important.

Three-dimensional (3D) spheroid culture provides a physiologically relevant alternative to conventional two-dimensional culture, but reliable spheroid formation in microwells depends on limiting cell-substrate adhesion. Pluronic F127 is an amphiphilic triblock copolymer that forms a hydrated surface layer, reducing protein adsorption. Here, we evaluated whether this intrinsic anti-fouling property could restore an anti-adhesive surface in used microwell plates to promote spheroid formation. Using chondrogenic ATDC5 and pancreatic {beta}-cell INS-1 cells, we characterized spheroid assembly kinetics, F127 cytotoxicity, surface hydrophilicity, protein adsorption, and spheroid morphology including size and shape factor. Both cell types formed compact spheroids within 24 hours on commercial anti-adhesive microwells. F127 coating markedly reduced water contact angle and protein adsorption, confirming increased surface hydrophilicity and reduced protein fouling. In microwells stripped of their original surface coating, F127 coating amounts of approximately 0.011-0.045 mg/cm2 consistently promoted spheroid formation across both cell types. Soluble F127 concentrations were confirmed to be non-cytotoxic up to 0.625% (w/v), while even complete dissolution of the highest tested coating amount would correspond to only 0.025% (w/v) F127. This simple, reproducible, and low-cost surface-modification strategy may provide an accessible approach for re-functionalizing microwell platforms for 3D cell culture.

Lung-targeted delivery of mRNA with lipid nanoparticles (LNPs) demonstrates high potential for therapeutic applications in pulmonary disorders. However, progress in pulmonary mRNA therapeutics is constrained by the challenges of engineering lipids that are both safe and highly effective at targeting the lungs. To meet these critical needs, we designed triazine-based (TZ) ionizable lipids with cyanuric chloride as the linker between the cationic head and the lipophilic tail, which allows for easy derivatization capable of systemic mRNA delivery. Three TZ-based lipids were synthesized using the same ionizable headgroups while differing in the carbon tail length and evaluated for their in vitro and in vivo protein expression. Notably, all three lipids result in pulmonary expression after intravenous administration, but the TZ lipid containing a C14 tail does so without any indication of thrombosis, both in vitro and in vivo as compared to other formulations. Our findings highlight the effect of minor chemical modifications driving altered in vivo activity, thus enabling new opportunities for safe pulmonary delivery of mRNA for lung-related diseases.

Musculoskeletal symptoms are frequently reported following silicone breast implantation. However, the biological mechanisms linking implant-derived silicone exposure to skeletal muscle alterations remain poorly understood, partly because the biological effects of silicone have long been debated in the context of its biocompatibility. Here, we chemically characterized the low-molecular-weight fraction of the breast implant silicone exposome, readily released from implant gel through gel bleed, and investigated its potential biological consequences using an integrated approach combining analytical chemistry, clinical transcriptomics and histology, and controlled in vitro muscle experiments. Transcriptomic analyses of periprosthetic tissues associated with silicone implant rupture revealed unexpected myogenic and neuromuscular signatures in tissue conventionally regarded as predominantly fibrous, together with alterations in lipid metabolism and transport. These findings were supported by histological evidence of close interactions between periprosthetic tissue and skeletal muscle. Chemical analysis of the implant-gel extract detected linear siloxane L2 and cyclic siloxanes D3-D8, with tentative assignment of D9. In vitro, C2C12 cells exposed to the implant-gel extract showed up to 30% reduced viability and decreased expression of key neuromyogenic genes. Together, these findings provide convergent chemical, clinical, and experimental evidence that low-molecular-weight constituents of the breast implant silicone exposome may constitute a biologically active exposure capable of affecting skeletal muscle. The associated alterations in lipid metabolism and transport further provide a mechanistic framework for investigating the cellular handling and potential tissue distribution of hydrophobic silicone-derived species. These findings position silicone gel bleed as a biologically relevant source of chemical exposure rather than solely a material-integrity phenomenon.

Damage to the skin by trauma, burns, or surgical procedures often results in uncontrolled bleeding, which remains a leading cause of preventable death following injury, yet most conventional hemostatic materials are engineered solely to arrest bleeding and often adhere strongly to the wound bed, causing pain, rebleeding, and disruption of newly formed tissue upon removal. Here, we report a DNA hydrogel that structurally mimics neutrophil extracellular traps (NETs) and is crosslinked using a bioactive small molecule with potent autophagy-inducing, cardioprotective, anti-inflammatory, antioxidant, and mitochondria-protective properties, integrating rapid hemostasis with active support for tissue regeneration in a single biomaterial. The DNA network provides an intrinsically biocompatible, biodegradable scaffold capable of recruiting platelets and erythrocytes to achieve rapid clot formation, while the bioactive crosslinker is released as the network degrades, delivering a sustained cytoprotective and anti-inflammatory stimulus directly at the wound site. The hydrogel was characterised physiochemically and evaluated for cytocompatibility, hemolytic potential, hemostatic efficacy, and wound-healing performance in a murine model. Results demonstrate that the bioactive-crosslinked DNA hydrogel achieves rapid, effective hemostasis, while accelerating wound closure and supporting regenerative tissue remodelling. This dual-function platform offers a promising strategy for next-generation wound-care biomaterials that unite immediate bleeding control with accelerated, natural tissue healing.

The recent development of Volumetric Imaging via Photochemical Sectioning (VIPS) has enabled nanoscale imaging of whole-mount tissue samples of virtually any size by embedding intact tissue in a photocleavable, superabsorbent hydrogel. However, the efficacy of sample embedding, imaging, and photochemical sectioning is fundamentally governed by the mechanical stiffness, structural stability, and photodegradation kinetics of the photocleavable hydrogel (PC-gel) polymer network. To elucidate the effect of the photosensitive crosslinker design on these critical properties, we synthesized a set of photocleavable crosslinkers (PCs) with varying polyethylene glycol (PEG) backbone lengths and prepared the corresponding PC-gels under a fixed monomer formulation and polymerization condition. We quantified and compared the viscoelastic properties of the formed PC-gels at their swollen states, and found that the crosslinker length markedly reshaped the PC-gel mechanics. In addition, we evaluated the light-triggered degradation of the PC-gels using both wide-field and spatially-controlled illumination. We found that PC-1000, PC-1500, and PC-2000 gels remained comparably photodegradable, all enabling on-demand, spatially confined decrosslinking under such illuminations. These results provide practical guidelines for modulating the crosslinker architecture of PC-gel polymer networks to achieve optimal physicochemical properties for whole-mount tissue imaging using VIPS.

Disruption of the wound healing cascade can result in pathological outcomes, including fibrosis due to myofibroblast-mediated contraction and collagen deposition. Despite the clinical significance, effective treatments for fibrosis remain limited as current therapies often show inconsistent efficacy, adverse effects, and patient discomfort. Combinatorial therapeutic strategies integrating biomaterial scaffolds with gene delivery have shown promise in regenerative healing. MicroRNAs (miRNAs) are key regulators of fibrotic signalling in cells, including fibroblasts and myofibroblasts. Specifically, miRNA-29b is notable for downregulating pro-fibrotic genes, including collagen type I, reducing ECM accumulation, and limiting fibroblast/myofibroblast overactivation. In this context, the present work develops a collagen-GAG (CG) scaffold platform for delivery of miRNA-29b complexed GET nanoparticles to inhibit fibrosis. Initially, bioinformatic analysis of miRNA-29b validated its involvement in ECM-associated pathways and processes, followed by successful nanoparticle internationalisation in primary dermal fibroblasts. The anti-fibrotic efficacy of the optimised miRNA-29b nanoparticles was subsequently demonstrated by significant reductions in collagen deposition and -SMA expression, both key indicators of myofibroblast differentiation and fibrosis. The optimised miRNA-29b formulation was then incorporated into 3D porous collagen-GAG (CG) scaffolds, which modulated fibrotic gene expression while preserving scaffold structure conducive to fibroblast/myofibroblast infiltration and proliferation. Finally, functional outcomes of seeded TGF-{beta}-stimulated fibroblasts, including reduced matrix contraction, -SMA expression, and ECM deposition, were comparable to those observed in non-fibrotic conditions, thereby confirming the therapeutic potential of scaffold-mediated miRNA-29b delivery. Together, these findings demonstrate that scaffold-mediated miRNA-29b delivery represents a promising anti-fibrotic strategy for wound healing by mitigating myofibroblast activation, limiting matrix contraction, and preventing pathological ECM accumulation.

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.

Introduction: Advanced biomaterials for regenerative medicine are increasingly expected to combine multifunctionality and compatibility with tissue-specific cellular processes. In this context, hydroxyapatite-based platforms modified through ionic substitution represent promising candidates, as they may integrate structural similarity to bone mineral with additional biological functionality and luminescent properties, enabling diagnostic applications and real-time monitoring. In this study, we evaluated whether silicate-phosphate substituted calcium hydroxyapatite Ca10(PO4)6-x(SiO4)x(OH)2 (where x = 1.5) co-doped with lithium(I), europium(III), and gadolinium(III) ions (Si-HAp-LEG) affects the osteogenic, chondrogenic, and adipogenic differentiation potential of human bone marrow stromal/stem cells (BMSCs). Methods: Human BMSCs were cultured under lineage-specific differentiation conditions in the presence of undoped silicate-substituted phosphate hydroxyapatite (abbr. as Si-HAp), which served as a control, and two distinct Si-HAp-LEG formulations differing in gadolinium(III) (Gd3+) as well as lithium (Li+) and europium(III) (Eu3+) ion concentrations: Si-HAp-LEG-221 (1 mol% Gd3+ ion) and Si-HAp-LEG-222 (2 mol% Gd3+ ion). Differentiation-associated phenotypic outcomes, including extracellular matrix formation and lipid accumulation, were evaluated using Safranin O, Alizarin Red, and Oil Red O staining. In parallel, biomaterial-induced molecular responses were characterized at the transcriptomic and protein levels using RT-qPCR for selected coding and non-coding RNAs and Western blot analysis for representative lineage-associated proteins. Results: Histochemical evaluation confirmed that, across all tested biomaterial groups, BMSCs retained the ability to form mineralized calcium deposits, proteoglycan-rich extracellular matrix, and intracellular lipid accumulation under osteogenic, chondrogenic, and adipogenic conditions, respectively. Quantitative staining analysis revealed no significant Si-HAp-LEG-dependent enhancement of terminal differentiation outcomes compared with undoped Si-HAp. In turn, the molecular response differed between biomaterials modifications. Si-HAp-LEG-222 induced the most prominent changes in transcriptional and post-transcriptional regulators, particularly within BMP/SMAD-associated pathways under osteogenic and chondrogenic conditions, underlying a potential link between gadolinium concentration and osteogenic lineage commitment. However, these transcriptomic responses were not mirrored by consistent changes at the protein level. The results suggest that silicate-phosphate substituted hydroxyapatite co-doped with Li+, Eu3+, and Gd3+ ions primarily affects the early regulatory pathways associated with BMSCs differentiation rather than enhancing their terminal maturation. Discussion: In conclusion, the collective data indicate that Li+, Eu3+, and Gd3+ ions LEG co-doping broadens the multifunctional potential of Si-HAp by introducing imaging-related properties while preserving its underlying pro-regenerative character. Li+, Eu3+, and Gd3+ ions co-doped LEG-substituted Si-HAp may therefore be considered a compatible biomaterial platform that maintains BMSC cellular plasticity and supports balanced, differentiation-dependent modulation of lineage-associated molecular responses.

Vascularization, which is critical for most engineered tissue constructs, is dependent on interactions between endothelial cells and spatiotemporally presented biochemical and biophysical cues. Yet, most hydrogels define these cues at the time of fabrication, thus precluding adjustments to actively drive vascular formation. We developed acoustically responsive scaffolds (ARSs) that use focused ultrasound to trigger growth factor release and localized matrix remodeling within fibrin hydrogels. ARSs were formed by incorporating phase-shift emulsions containing basic fibroblast growth factor (bFGF) along with perfluorohexane (C6) or perfluorooctane (C8). Upon ultrasound exposure, stable bubbles were generated in C6-ARSs that locally compacted the matrix and increased macroscale stiffness. Comparatively, in C8-ARSs, ultrasound generated macropores without impacting viscoelastic properties. Ultrasound increased bFGF release from both ARS types, which enhanced in vitro and in vivo vasculogenic assembly in ARSs with co-encapsulated endothelial cells and fibroblasts. Our data also show that ultrasound-driven matrix remodeling without bFGF release increased endothelial sprouting. In C6-ARSs, elevated levels of F-actin were observed in both cell types adjacent to bubbles as well as increased YAP intensity and nuclear asymmetry. Together, these results establish ARSs as reconfigurable hydrogels that couple on-demand release of biochemical cues with programmable matrix restructuring to direct three-dimensional microvascular assembly.

Understanding the physicochemical factors that govern siRNA nanocarrier assembly is essential for the rational design of effective delivery systems. By optimizing various lipid compositions, cholesterol content and PEG length we created a peptide-functionalized cationic liposomal platform made of DOPE/TAP lipids with cholesterol-anchored nona-arginine (R9-Chol) for siRNA complexation, intracellular transport and effective silencing of the target EGFR gene. Analysis of {zeta}-potential and dynamic light scattering allowed to rationally design formulation of stable, monodisperse nanoscale lipoplexes with a positive surface charge. With fluorescence polarization, circular dichroism and agarose gel electrophoresis we found an optimal siRNA:liposome complexation ratio of 1:77, which protected siRNA from ribonuclease-mediated degradation. Morphological imaging confirmed a shift from discrete vesicular structures to organized multilamellar lipoplexes, consistent with electrostatically driven self-assembly. In cellular studies, the optimized nanocarrier promoted efficient uptake of fluorescent siRNA in MDA-MB-231 cells and achieved functional delivery of anti-EGFR, leading to substantially reduced expression of the target gene at both transcript and protein levels. This work offers mechanistic understanding of peptide-assisted lipid:siRNA assembly and positions R9-functionalized DOPE/TAP liposomes as a promising platform for siRNA delivery.

Collagen, a major component of the extracellular matrix, regulates cellular behaviors, such as adhesion, differentiation, and angiogenesis. These functions are mediated by interactions between specific amino acid motifs within the collagen triple-helical structure and collagen-binding biomolecules. These include cell-surface receptors, such as integrins, discoidin domain receptors (DDRs), and syndecans, a family of transmembrane heparan sulfate proteoglycans (HSPGs). Signals mediated by these receptors are integrated to regulate cell fate. However, native collagen simultaneously presents multiple receptor-binding motifs, making it difficult to isolate receptor-specific functions and to evaluate receptor crosstalk. Here, we introduce a composition-controlled artificial collagen matrix platform that enables independent tuning of multiple receptor-binding motifs within a constant triple-helical scaffold. This material was produced by disulfide crosslinking of chemically synthesized collagen-like triple-helical peptides, each bearing a single defined receptor-binding sequence. By varying the mixing ratios of these peptides before crosslinking, we systematically controlled the composition of receptor-binding motifs within the matrices. We applied this platform to nerve growth factor-dependent neuronal differentiation of PC12 cells, a process supported by collagen. Matrices containing only integrin-binding sequences were sufficient to support this differentiation. Incorporation of an HSPG-binding sequence had little additional effect, whereas incorporation of a DDR-binding sequence suppressed integrin-mediated differentiation and coincided with DDR phosphorylation. These results reveal opposing roles of collagen-binding integrins and DDRs in regulating PC12 cell differentiation. Composition-controlled artificial collagen provides a versatile matrix platform for dissecting functional crosstalk among collagen receptors.

1.Surface functionality and pH play a decisive role in governing the structural dynamics, hydration, and drug-binding behaviour of dendrimers. Here, all-atom molecular dynamics (MD) simulations were performed on five generations of PAMAM (G1-G5) and PETIM (G2-G6) dendrimers with O-core and N-core architectures, functionalized with amine, carboxylic acid, or sugar terminal groups under different protonation states. Protonation of the tertiary branch-point amines expands the dendrimer structure, increases internal porosity and hydration, and enhances structural fluctuations across both families. In contrast, non-protonated amine -NH2 (NP) and carboxylic acid -COOH (NP) terminated dendrimers, together with deprotonated carboxylate-COO- (DeP) systems, retain comparatively compact conformations. Sugar-functionalized dendrimers ({beta}-galactose-terminated PETIM and D-glucose-terminated PAMAM) are most hydrated and structurally rigid, whereas amine-terminated dendrimers exhibit the greatest conformational dynamics. PAMAM dendrimers with -NH2, -NH3+, and -COO- terminal groups are generally more hydrated than their PETIM counterparts. However, {beta}-galactose-terminated PETIM dendrimers are more hydrophilic than D-glucose-terminated PAMAM dendrimers. N-core PETIM dendrimers also adopt more compact and spherical conformations than equivalent O-core PETIM dendrimers. Drug-binding MD simulations show that curcumin binding is dominated by van der Waals (vdW) interactions, whereas doxorubicin complexation is primarily driven by electrostatic interactions. Among the investigated surface functionalities, -NH2 (NP), -NH3+ (P), -COOH (NP), and -COO- (DeP) terminations exhibit the most favourable drug-binding characteristics. Except for deprotonated carboxylate systems, curcumin binds more strongly than doxorubicin. Overall, these findings establish molecular-level relationships between surface functionality, protonation state, dendrimer architecture, and drug-binding behaviour, providing design principles for pH-responsive dendrimer nanocarriers with enhanced drug-loading and controlled-release performance. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=75 SRC="FIGDIR/small/742721v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@119bf29org.highwire.dtl.DTLVardef@1554d86org.highwire.dtl.DTLVardef@154a254org.highwire.dtl.DTLVardef@16d5c5b_HPS_FORMAT_FIGEXP M_FIG C_FIG