Aging processes in dental thermoplastics - Thermoanalytical investigations and effects on Vickers as well as Martens hardness.

OBJECTIVE: The influence of various aging protocols, representing and accelerating influences present in the dental context, on possible changes in the microstructure and mechanical properties of thermoplastics was investigated. In order to minimize the complexity of the systems, first pure polymers and then later the equivalent dental polymeric materials were analyzed. MATERIALS AND METHODS: Pure polymers (Poly(methyl methacrylate) - PMMA, Polyoxymethylene homopolymer - POM-H, Polyether ether ketone - PEEK, Nylon 12 - PA12, Polypropylene - PP) were analyzed before as well as after applying different aging protocols relevant to the oral environment (ethanol, thermocycling, alkaline and acidic setting) by differential scanning calorimetry (DSC) and thermogravimetric analysis (TGA). The thermoanalytical parameters used were glass transition temperature (Tg), melting peak and crystallization peak temperature (Tpm, Tpc) and decomposition behavior. In a second step selected commercially available dental products (Telio CAD - PMMAD, Zirlux Acetal - POMD, Juvora Natural Dental Disc - PEEKD) aged by the protocol that previously showed strong effects were examined and additionally tested for changes in their Vickers and Martens hardness by Mann-Whitney-U test. RESULTS: The combinations of pure polymers and viable aging protocols analyzed within this study were identified via TGA or DSC as PA12 & thermocycling, POM-H & denture cleanser/lactic acid/ethanol, PP & lactic acid. The dental polymeric materials PMMAD and POMD due to aging in lactic acid showed slight but significantly (p < 0.01) reduced Vickers and partly Martens hardness. PEEK showed the greatest material resistance within this study.

Modern Photodynamic Glioblastoma Therapy Using Curcumin- or Parietin-Loaded Lipid Nanoparticles in a CAM Model Study.

Natural photosensitizers, such as curcumin or parietin, play a vital role in photodynamic therapy (PDT), causing a light-mediated reaction that kills cancer cells. PDT is a promising treatment option for glioblastoma, especially when combined with nanoscale drug delivery systems. The curcumin- or parietin-loaded lipid nanoparticles were prepared via dual asymmetric centrifugation and subsequently characterized through physicochemical analyses including dynamic light scattering, laser Doppler velocimetry, and atomic force microscopy. The combination of PDT and lipid nanoparticles has been evaluated in vitro regarding uptake, safety, and efficacy. The extensive and well-vascularized chorioallantois membrane (CAM) of fertilized hen's eggs offers an optimal platform for three-dimensional cell culture, which has been used in this study to evaluate the photodynamic efficacy of lipid nanoparticles against glioblastoma cells. In contrast to other animal models, the CAM model lacks a mature immune system in an early stage, facilitating the growth of xenografts without rejection. Treatment of xenografted U87 glioblastoma cells on CAM was performed to assess the effects on tumor viability, growth, and angiogenesis. The xenografts and the surrounding blood vessels were targeted through topical application, and the effects of photodynamic therapy have been confirmed microscopically and via positron emission tomography and X-ray computed tomography. Finally, the excised xenografts embedded in the CAM were analyzed histologically by hematoxylin and eosin and KI67 staining.

Fabrication of 3D Printed, Core-and-Shell Implants as Controlled Release Systems for Local siRNA Delivery.

The development and clinical translation of small interfering RNA (siRNA) therapies remains challenging owing to their poor pharmacokinetics. 3D printing technology presents a great opportunity to fabricate personalized implants for local and sustained delivery of siRNA. Hydrogels can mimic the mechanical properties of tissues, avoiding the problems associated with rigid implants. Herein, a thermoresponsive composite hydrogel suitable for extrusion 3D-printing is formulated to fabricate controlled-release implants loaded with siRNA-Lipofectamine RNAiMAX complexes. A hydrogel matrix mainly composed of uncharged agarose to protect siRNA from decomplexation is selected. Additionally, pluronic F127 and gelatin are added to improve the printability, degradation, and cell adhesion to the implants. To avoid exposing siRNA to thermal stress during the printing process, a core-and-shell design is set up for the implants in which a core of siRNA-complexes loaded-pluronic F127 is printed without heat and enclosed with a shell comprising the thermoresponsive composite hydrogel. The release profile of siRNA-complexes is envisioned to be controlled by varying the printing patterns. The results reveal that the implants sustain siRNA release for one month. The intactness of the released siRNA-complexes is proven until the eighth day. Furthermore, by changing the printing patterns, the release profiles can be tailored.

Radiofluorination of an Anionic, Azide-Functionalized Teroligomer by Copper-Catalyzed Azide-Alkyne Cycloaddition.

This study describes the synthesis, radiofluorination and purification of an anionic amphiphilic teroligomer developed as a stabilizer for siRNA-loaded calcium phosphate nanoparticles (CaP-NPs). As the stabilizing amphiphile accumulates on nanoparticle surfaces, the fluorine-18-labeled polymer should enable to track the distribution of the CaP-NPs in brain tumors by positron emission tomography after application by convection-enhanced delivery. At first, an unmodified teroligomer was synthesized with a number average molecular weight of 4550 ± 20 Da by free radical polymerization of a defined composition of methoxy-PEG-monomethacrylate, tetradecyl acrylate and maleic anhydride. Subsequent derivatization of anhydrides with azido-TEG-amine provided an azido-functionalized polymer precursor (o14PEGMA-N3) for radiofluorination. The 18F-labeling was accomplished through the copper-catalyzed cycloaddition of o14PEGMA-N3 with diethylene glycol-alkyne-substituted heteroaromatic prosthetic group [18F]2, which was synthesized with a radiochemical yield (RCY) of about 38% within 60 min using a radiosynthesis module. The 18F-labeled polymer [18F]fluoro-o14PEGMA was obtained after a short reaction time of 2-3 min by using CuSO4/sodium ascorbate at 90 °C. Purification was performed by solid-phase extraction on an anion-exchange cartridge followed by size-exclusion chromatography to obtain [18F]fluoro-o14PEGMA with a high radiochemical purity and an RCY of about 15%.

Standardization Approaches for Extracellular Vesicle Loading with Oligonucleotides and Biologics.

Extracellular vesicles (EVs) are widely recognized for their potential as drug delivery systems. EVs are membranous nanoparticles shed from cells. Among their natural features are their ability to shield cargo molecules against degradation and enable their functional internalization into target cells. Especially biological or bio-inspired large molecules (LMs), like nucleic acids, proteins, peptides, and others, may profit from encapsulation in EVs for drug delivery purposes. In the last years, a variety of loading protocols are explored for different LMs. The lack of standardization in the EV drug delivery field has impeded their comparability so far. Currently, the first reporting frameworks and workflows for EV drug loading are proposed. The aim of this review is to summarize these evolving standardization approaches and set recently developed methods into context. This will allow for enhanced comparability of future work on EV drug loading with LMs.

Retention force, translucency, and microstructural properties of translucent temporary luting cements: An in vitro study.

The aim of the study was to investigate the retention behaviour (pull-off force include adhesive remnant index = ARI) as well as translucency of various temporary luting cements and use microstructure elucidation methods to formulate explanatory approaches to their mode of action. The retention force of the temporary luting cements Provicol QM Plus (P+), Provicol QM Aesthetic (Pae), Bifix Temp (BiT), and as a reference a glass ionomer cement Meron (M) with a direct (Structur 3/S3) or an indirect (Structure CAD/SCAD) resin-based composite restauration was investigated after accelerated aging (thermocycling). Additional investigation of the physical properties was performed regarding to translucency and surface free energy. The microstructure was evaluated by X-ray diffraction, thermogravimetric analysis, differential scanning calorimetry and micro X-ray computed tomography. All tested temporary luting cements showed different pull-off forces in the range between 3.0 and 16.8 N in combination with S3 or SCAD after thermocycling. Only BiT with S3 showed pull-off forces of 129.2 N and complete retention on the restoration (ARI = 0), which was significant (p < .001) to all other samples. High translucency (BiT > Pae > M > P+) was observed for materials with lower crystalline content and low residual mass (usally resulting from higher organic content). M showed the highest surface free energy with a predominantly polar fraction, while BiT had a predominantly dispersive fraction. The highest porosity was observed in the coronal region of the restoration. The results suggest that translucency of temporary luting cements can be increased with higher organic and lower cryristall content. All combinations of cements and temporary restorations (direct/indirect; with the exception of BiT/S3) showed pull-off forces below 17 N (equivalent to a weight force of ∼1.7 kg), which allows manual detachment of the restoration by the dentist.

Utilizing 4D Printing to Design Smart Gastroretentive, Esophageal, and Intravesical Drug Delivery Systems.

The breakthrough of 3D printing in biomedical research has paved the way for the next evolutionary step referred to as four dimensional (4D) printing. This new concept utilizes the time as the fourth dimension in addition to the x, y, and z axes with the idea to change the configuration of a printed construct with time usually in response to an external stimulus. This can be attained through the incorporation of smart materials or through a preset smart design. The 4D printed constructs may be designed to exhibit expandability, flexibility, self-folding, self-repair or deformability. This review focuses on 4D printed devices for gastroretentive, esophageal, and intravesical delivery. The currently unmet needs and challenges for these application sites are tried to be defined and reported on published solution concepts involving 4D printing. In addition, other promising application sites that may similarly benefit from 4D printing approaches such as tracheal and intrauterine drug delivery are proposed.

Aging Processes and Their Influence on the Mechanical Properties of Printable Occlusal Splint Materials.

Three-dimensional (3D)-printed occlusal splints are becoming more prevalent in the treatment of tooth substance loss due to their fast and cost-effective production. The purpose of this in vitro study was to investigate whether the mechanical properties (tensile strength-TS, modulus of elasticity in tension-ME, and Vickers hardness-HV) vary between the materials (printed dimethacrylate-based resins: Keyprint KeySplint soft-KEY, Luxaprint Ortho Plus-LUX, V-Print splint-VPR, printed methacrylate-based resins Freeprint splint 2.0-FRE, and milled methacrylate-based material, CLEAR splint-CLE), and the influence of aging processes (extraoral storage conditions and nightly or daily use) was examined. The printed methacrylate-based resins (FRE, LUX, and VPR) had much higher TS (43.7-48.5 MPa compared to 12.3-13.3 MPa), higher ME (2.01-2.37 GPa compared to 0.43-0.72 GPa), and higher HV (11.8-15.0 HV compared to 3.3-3.5 HV) than both of the methacrylate-based resins (KEY and CLE) after the production process. Although the TS, ME, and HV of the printed dimethacrylate resins (FRE, LUX, and VPR) decreased significantly under humid conditions with possibly elevated temperatures (thermocycling as well as 37 °C), these mechanical properties were significantly higher than both methacrylate-based resins (KEY and CLE). Therefore, printed dimethacrylate resins should be used rather than methacrylate-based resins for high expected masticatory forces, low wall thicknesses, or very long wearing times (≥6 months).

Synergistic siRNA Loading of Extracellular Vesicles Enables Functional Delivery into Cells.

RNA interference opened new approaches for disease treatment but safe and efficient cell delivery remains a bottleneck. Extracellular vesicles (EVs) are known to naturally shuttle RNA. Due to their potent cell internalization and low-cost scalability, milk-derived EVs in particular are considered promising RNA delivery systems. However, low drug loading currently impedes their use. Here, innovative exogenous loading strategies for small interfering RNA (siRNA) are explored and systematically compared regarding encapsulation efficiency, loading capacity, and loading concentration. Firstly, siRNA is pre-accumulated in liposomes or stabilized calcium phosphate nanoparticles (CaP-NP). The selected systems, which exhibited neutral or negative zeta potentials, are then applied for EV loading. Secondly, EVs are concentrated and applied to protocols known for liposome loading: dehydration-rehydration of vesicles, based on freeze-drying, and mixing by dual asymmetric centrifugation (DAC) after ultracentrifugation. Additionally, DAC after EV ultracentrifugation is combined with CaP-NP leading to a synergistic loading performance. The balance between energy input for siRNA loading and EV integrity is evaluated by monitoring the EV size, marker proteins, and morphology. For the EV-based siRNA formulation via DAC plus CaP-NP, EV properties are sufficiently maintained to protect the siRNA from degradation and deliver cell-death siRNA dose-dependently in Caco-2 cells.

Investigating 3R In Vivo Approaches for Bio-Distribution and Efficacy Evaluation of Nucleic Acid Nanocarriers: Studies on Peptide-Mimicking Ionizable Lipid.

Formulations based on ionizable amino-lipids have been put into focus as nucleic acid delivery systems. Recently, the in vitro efficacy of the lipid formulation OH4:DOPE has been explored. However, in vitro performance of nanomedicines cannot correctly predict in vivo efficacy, thereby considerably limiting pre-clinical translation. This is further exacerbated by limited access to mammalian models. The present work proposes to close this gap by investigating in vivo nucleic acid delivery within simpler models, but which still offers physiologically complex environments and also adheres to the 3R guidelines (replace/reduce/refine) to improve animal experiments. The efficacy of OH4:DOPE as a delivery system for nucleic acids is demonstrated using in vivo approaches. It is shown that the formulation is able to transfect complex tissues using the chicken chorioallantoic membrane model. The efficacy of DNA and mRNA lipoplexes is tested extensively in the zebra fish (Danio rerio) embryo which allows the screening of biodistribution and transfection efficiency. Effective transfection of blood vessel endothelial cells is seen, especially in the endocardium. Both model systems allow an efficacy screening according to the 3R guidelines bypassing the in vitro-in vivo gap. Pilot studies in mice are performed to correlate the efficacy of in vivo transfection.

Mineralizing Gelatin Microparticles as Cell Carrier and Drug Delivery System for siRNA for Bone Tissue Engineering.

The local release of complexed siRNA from biomaterials opens precisely targeted therapeutic options. In this study, complexed siRNA was loaded to gelatin microparticles cross-linked (cGM) with an anhydride-containing oligomer (oPNMA). We aggregated these siRNA-loaded cGM with human mesenchymal stem cells (hMSC) to microtissues and stimulated them with osteogenic supplements. An efficient knockdown of chordin, a BMP-2 antagonist, caused a remarkably increased alkaline phosphatase (ALP) activity in the microtissues. cGM, as a component of microtissues, mineralized in a differentiation medium within 8-9 days, both in the presence and in the absence of cells. In order to investigate the effects of our pre-differentiated and chordin-silenced microtissues on bone homeostasis, we simulated in vivo conditions in an unstimulated co-culture system of hMSC and human peripheral blood mononuclear cells (hPBMC). We found enhanced ALP activity and osteoprotegerin (OPG) secretion in the model system compared to control microtissues. Our results suggest osteoanabolic effects of pre-differentiated and chordin-silenced microtissues.

Amphiphilic Anionic Oligomer-Stabilized Calcium Phosphate Nanoparticles with Prospects in siRNA Delivery via Convection-Enhanced Delivery.

Convection-enhanced delivery (CED) has been introduced as a concept in cancer treatment to generate high local concentrations of anticancer therapeutics and overcome the limited diffusional distribution, e.g., in the brain. RNA interference provides interesting therapeutic options to fight cancer cells but requires nanoparticulate (NP) carriers with a size below 100 nm as well as a low zeta potential for CED application. In this study, we investigated calcium phosphate NPs (CaP-NPs) as siRNA carriers for CED application. Since CaP-NPs tend to aggregate, we introduced a new terpolymer (o14PEGMA(1:1:2.5) NH3) for stabilization of CaP-NPs intended for delivery of siRNA to brain cancer cells. This small terpolymer provides PEG chains for steric stabilization, and a fat alcohol to improve interfacial activity, as well as maleic anhydrides that allow for both labeling and high affinity to Ca(II) in the hydrolyzed state. In a systematic approach, we varied the Ca/P ratio as well as the terpolymer concentration and successfully stabilized NPs with the desired properties. Labeling of the terpolymer with the fluorescent dye Cy5 revealed the terpolymer's high affinity to CaP. Importantly, we also determined a high efficiency of siRNA binding to the NPs that caused very effective survivin siRNA silencing in F98 rat brain cancer cells. Cytotoxicity investigations with a standard cell line resulted in minor and transient effects; no adverse effects were observed in organotypic brain slice cultures. However, more specific cytotoxicity investigations are required. This study provides a systematic and mechanistic analysis characterizing the effects of the first oligomer of a new class of stabilizers for siRNA-loaded CaP-NPs.

Microtissues from mesenchymal stem cells and siRNA-loaded cross-linked gelatin microparticles for bone regeneration.

The aim of this study was the evaluation of cross-linked gelatin microparticles (cGM) as substrates for osteogenic cell culture to assemble 3D microtissues and their use as delivery system for siRNA to cells in these assemblies. In a 2D transwell cultivation system, we found that cGM are capable to accumulate calcium ions from the surrounding medium. Such a separation of cGM and SaOS-2 â€‹cells consequently led to a suppressed matrix mineral formation in the SaOS-2 culture on the well bottom of the transwell system. Thus, we decided to use cGM as component in 3D microtissues and get a close contact between calcium ion accumulating microparticles and cells to improve matrix mineralization. Gelatin microparticles were cross-linked with a N,N-diethylethylenediamine-derivatized (DEED) maleic anhydride (MA) containing oligo (pentaerythritol diacrylate monostearate-co-N-isopropylacrylamide-co-MA) (oPNMA) and aggregated with SaOS-2 or human mesenchymal stem cells (hMSC) to microtissue spheroids. We systematically varied the content of cGM in microtissues and observed cell differentiation and tissue formation. Microtissues were characterized by gene expression, ALP activity and matrix mineralization. Mineralization was detectable in microtissues with SaOS-2 â€‹cells after 7 days and with hMSC after 24-28 days in osteogenic culture. When we transfected hMSC via cGM loaded with Lipofectamine complexed chordin siRNA, we found increased ALP activity and accelerated mineral formation in microtissues in presence of BMP-2. As a model for positive paracrine effects that indicate promising in vivo effects of these microtissues, we incubated pre-differentiated microtissues with freshly seeded hMSC monolayers and found improved mineral formation all over the well in the co-culture model. These findings may support the concept of microtissues from hMSC and siRNA-loaded cGM for bone regeneration.

Adjustable Thermo-Responsive, Cell-Adhesive Tissue Engineering Scaffolds for Cell Stimulation through Periodic Changes in Culture Temperature.

A three-dimensional (3D) scaffold ideally provides hierarchical complexity and imitates the chemistry and mechanical properties of the natural cell environment. Here, we report on a stimuli-responsive photo-cross-linkable resin formulation for the fabrication of scaffolds by continuous digital light processing (cDLP), which allows for the mechano-stimulation of adherent cells. The resin comprises a network-forming trifunctional acrylate ester monomer (trimethylolpropane triacrylate, or TMPTA), N-isopropyl acrylamide (NiPAAm), cationic dimethylaminoethyl acrylate (DMAEA) for enhanced cell interaction, and 4-acryloyl morpholine (AMO) to adjust the phase transition temperature (Ttrans) of the equilibrium swollen cross-polymerized scaffold. With glycofurol as a biocompatible solvent, controlled three-dimensional structures were fabricated and the transition temperatures were adjusted by resin composition. The effects of the thermally induced mechano-stimulation were investigated with mouse fibroblasts (L929) and myoblasts (C2C12) on printed constructs. Periodic changes in the culture temperature stimulated the myoblast proliferation.

Biodegradable macromers for implant bulk and surface engineering.

Macromers, polymeric molecules with at least two functional groups for cross-polymerization, are interesting materials to tailor mechanical, biochemical and degradative bulk and surface properties of implants for tissue regeneration. In this review we focus on macromers with at least one biodegradable building block. Manifold design options, such as choice of polymeric block(s), optional core molecule and reactive groups, as well as cross-co-polymerization with suitable anchor or linker molecules, allow the adaptation of macromer-based biomaterials towards specific application requirements in both hard and soft tissue regeneration. Implants can be manufactured from macromers using additive manufacturing as well as molding and templating approaches. This review summarizes and discusses the overall concept of biodegradable macromers and recent approaches for macromer processing into implants as well as techniques for surface modification directed towards bone regeneration. These aspects are reviewed including a focus on the authors' contributions to the field through research within the collaborative research project Transregio 67.

Osteogenic Potential of Mesenchymal Stem Cells from Adipose Tissue, Bone Marrow and Hair Follicle Outer Root Sheath in a 3D Crosslinked Gelatin-Based Hydrogel.

Bone transplantation is regarded as the preferred therapy to treat a variety of bone defects. Autologous bone tissue is often lacking at the source, and the mesenchymal stem cells (MSCs) responsible for bone repair mechanisms are extracted by invasive procedures. This study explores the potential of autologous mesenchymal stem cells derived from the hair follicle outer root sheath (MSCORS). We demonstrated that MSCORS have a remarkable capacity to differentiate in vitro towards the osteogenic lineage. Indeed, when combined with a novel gelatin-based hydrogel called Osteogel, they provided additional osteoinductive cues in vitro that may pave the way for future application in bone regeneration. MSCORS were also compared to MSCs from adipose tissue (ADMSC) and bone marrow (BMMSC) in a 3D Osteogel model. We analyzed gel plasticity, cell phenotype, cell viability, and differentiation capacity towards the osteogenic lineage by measuring alkaline phosphatase (ALP) activity, calcium deposition, and specific gene expression. The novel injectable hydrogel filled an irregularly shaped lesion in a porcine wound model displaying high plasticity. MSCORS in Osteogel showed a higher osteo-commitment in terms of calcium deposition and expression dynamics of OCN, BMP2, and PPARG when compared to ADMSC and BMMSC, whilst displaying comparable cell viability and ALP activity. In conclusion, autologous MSCORS combined with our novel gelatin-based hydrogel displayed a high capacity for differentiation towards the osteogenic lineage and are acquired by non-invasive procedures, therefore qualifying as a suitable and expandable novel approach in the field of bone regeneration therapy.

A focus on critical aspects of uptake and transport of milk-derived extracellular vesicles across the Caco-2 intestinal barrier model.

Bovine milk-derived extracellular vesicles (EVs) hold promises as oral drug delivery systems. Since EV bioavailability studies are difficult to compare, key factors regarding EV uptake and intestinal permeability remain little understood. This work aims to critically study uptake and transport properties of milk-derived EVs across the intestinal barrier in vitro by standardization approaches. Therefore, uptake properties were directly compared to liposomes in intestinal Caco-2 cells. Reliable staining results were obtained by the choice of three distinct EV labeling sites, while non-specific dye transfer and excess dye removal were carefully controlled. A novel fluorescence correction factor was implemented to account for different labeling efficiencies. Both EV and liposome uptake occurred mainly energy dependent with the neonatal Fc receptor (FcRn) providing an exclusive active pathway for EVs. Confocal microscopy revealed higher internalization of EVs whereas liposomes rather remained attached to the cell surface. Internalization could be improved when changing the liposomal formulation to resemble the EV lipid composition. In a Caco-2/HT29-MTX co-culture liposomes and EVs showed partial mucus penetration. For transport studies across Caco-2 monolayers we further established a standardized protocol considering the distinct requirements for EVs. Especially insert pore sizes were systematically compared with 3 µm inserts found obligatory. Obtained apparent permeability coefficients (Papp) reflecting the transport rate will allow for better comparison of future bioavailability testing.

Is Micro X-ray Computer Tomography a Suitable Non-Destructive Method for the Characterisation of Dental Materials?

The aim of the study was to investigate the effect of X-rays used in micro X-ray computer tomography (µXCT) on the mechanical performance and microstructure of a variety of dental materials. Standardised bending beams (2 × 2 × 25 mm3) were forwarded to irradiation with an industrial tomograph. Using three-dimensional datasets, the porosity of the materials was quantified and flexural strength was investigated prior to and after irradiation. The thermal properties of irradiated and unirradiated materials were analysed and compared by means of differential scanning calorimetry (DSC). Single µXCT measurements led to a significant decrease in flexural strength of polycarbonate with acrylnitril-butadien-styrol (PC-ABS). No significant influence in flexural strength was identified for resin-based composites (RBCs), poly(methyl methacrylate) (PMMA), and zinc phosphate cement (HAR) after a single irradiation by measurement. However, DSC results suggest that changes in the microstructure of PMMA are possible with increasing radiation doses (multiple measurements, longer measurements, higher output power from the X-ray tube). In summary, it must be assumed that X-ray radiation during µXCT measurement at high doses can lead to changes in the structure and properties of certain polymers.

Characterisation of the Filler Fraction in CAD/CAM Resin-Based Composites.

The performance of dental resin-based composites (RBCs) heavily depends on the characteristic properties of the individual filler fraction. As specific information regarding the properties of the filler fraction is often missing, the current study aims to characterize the filler fractions of several contemporary computer-aided design/computer-aided manufacturing (CAD/CAM) RBCs from a material science point of view. The filler fractions of seven commercially available CAD/CAM RBCs featuring different translucency variants were analysed using Scanning Electron Microscopy (SEM) with Energy Dispersive X-ray Spectroscopy (EDS), Micro-X-ray Computed Tomography (µXCT), Thermogravimetric Analysis (TG) and X-ray Diffractometry (XRD). All CAD/CAM RBCs investigated included midifill hybrid type filler fractions, and the size of the individual particles was clearly larger than the individual specifications of the manufacturer. The fillers in Shofu Block HC featured a sphericity of ≈0.8, while it was <0.7 in all other RBCs. All RBCs featured only X-ray amorphous phases. However, in Lava Ultimate, zircon crystals with low crystallinity were detected. In some CAD/CAM RBCs, inhomogeneities (X-ray opaque fillers or pores) with a size <80 µm were identified, but the effects were minor in relation to the total volume (<0.01 vol.%). The characteristic parameters of the filler fraction in RBCs are essential for the interpretation of the individual material's mechanical and optical properties.

Extrusion-Printing of Multi-Channeled Two-Component Hydrogel Constructs from Gelatinous Peptides and Anhydride-Containing Oligomers.

The performance of artificial nerve guidance conduits (NGC) in peripheral nerve regeneration can be improved by providing structures with multiple small channels instead of a single wide lumen. 3D-printing is a strategy to access such multi-channeled structures in a defined and reproducible way. This study explores extrusion-based 3D-printing of two-component hydrogels from a single cartridge printhead into multi-channeled structures under aseptic conditions. The gels are based on a platform of synthetic, anhydride-containing oligomers for cross-linking of gelatinous peptides. Stable constructs with continuous small channels and a variety of footprints and sizes were successfully generated from formulations containing either an organic or inorganic gelation base. The adjustability of the system was investigated by varying the cross-linking oligomer and substituting the gelation bases controlling the cross-linking kinetics. Formulations with organic N­methyl-piperidin-3-ol and inorganic K2HPO4 yielded hydrogels with comparable properties after manual processing and extrusion-based 3D-printing. The slower reaction kinetics of formulations with K2HPO4 can be beneficial for extending the time frame for printing. The two-component hydrogels displayed both slow hydrolytic and activity-dependent enzymatic degradability. Together with satisfying in vitro cell proliferation data, these results indicate the suitability of our cross-linked hydrogels as multi-channeled NGC for enhanced peripheral nerve regeneration.