Silk granular hydrogels self-reinforced with regenerated silk fibroin fibers.

Granular hydrogels with high stability, strength, and toughness are laborious to develop. Post-curing is often employed to bind microgels chemically and enhance mechanical properties. Here a unique strategy was investigated to maintain microgels together with a novel self-reinforced silk granular hydrogel composed of 10 wt% 20 kDa poly(ethylene glycol) dimethacrylate microgels and regenerated silk fibroin fibers. The principle is to use the swelling of microgels to concentrate the surrounding solution and regenerate silk fibroin in situ. Self-reinforcement is subsequently one of the added functions. We showed that silk fibroin in most compositions was homogeneously distributed and had successfully regenerated in situ around microgels, holding them together in a network-like structure. FTIR analysis revealed the presence of amorphous and crystalline silk fibroin, where 50% of the secondary structures could be assigned to strong ß-sheets. Swelling ratios, i.e. 10-45 vol%, increased proportionally with the microgel content, suggesting that mainly microgels governed swelling. In contrast, the elastic modulus, i.e. 58-296 kPa, increased almost linearly with silk fibroin content. Moreover, we showed that the precursor could be injected and cast into a given shape. Viscous precursors of various compositions were also placed side by side to create mechanical gradients. Finally, it was demonstrated that silk granular hydrogel could successfully be synthesized with other microgels like gelatin methacryloyl. Silk granular hydrogels represent, therefore, a novel class of self-reinforced hydrogel structures with tunable swelling and elastic properties.

An Intrinsically-Adhesive Family of Injectable and Photo-Curable Hydrogels with Functional Physicochemical Performance for Regenerative Medicine.

Attaching hydrogels to soft internal tissues is crucial for the development of various biomedical devices. Tough sticky hydrogel patches present high adhesion, yet with lack of injectability and the need for treatment of contacting surface. On the contrary, injectable and photo-curable hydrogels are highly attractive owing to their ease of use, flexibility of filling any shape, and their minimally invasive character, compared to their conventional preformed counterparts. Despite recent advances in material developments, a hydrogel that exhibits both proper injectability and sufficient intrinsic adhesion is yet to be demonstrated. Herein, a paradigm shift is proposed toward the design of intrinsically adhesive networks for injectable and photo-curable hydrogels. The bioinspired design strategy not only provides strong adhesive contact, but also results in a wide window of physicochemical properties. The adhesive networks are based on a family of polymeric backbones where chains are modified to be intrinsically adhesive to host tissue and simultaneously form a hydrogel network via a hybrid cross-linking mechanism. With this strategy, adhesion is achieved through a controlled synergy between the interfacial chemistry and bulk mechanical properties. The functionalities of the bioadhesives are demonstrated for various applications, such as tissue adhesives, surgical sealants, or injectable scaffolds.

Hybrid granular hydrogels: combining composites and microgels for extended ranges of material properties.

Developing hydrogels with optimal properties for specific applications is challenging as most of these properties, such as toughness, stiffness, swelling or deformability, are interrelated. The improvement of one property usually comes at the cost of another. In order to decouple the interdependence between these properties and to extend the range of material properties for hydrogels, we propose a strategy that combines composite and microgel approaches. The study focuses first on tailoring the swelling performance of hydrogels while minimally affecting other properties. The underlying principle is to partially substitute some of the hydrogels with pre-swollen microgels composed of the same materials. Swelling reductions up to 45% were obtained. Those granular hydrogels were then reinforced with nano-fibrillated cellulose fibres obtaining hybrid granular materials to improve their toughness and to further reduce their initial swelling. Four different structures of neat, granular and composite hydrogels including 63 different hydrogel compositions based on 20 kDa poly(ethylene glycol)dimethacrylate showed that the swelling ratio could be tailored without significantly affecting elastic modulus and deformation performance. The results explain the role of the PEGDM precursors on the swelling of the microgels as well as the influence of the microgel and fibre contents on the final properties. Moreover, the precursors of hydrogels with similar mechanical or swelling performance were injectable with a wide range of complex viscosities from 0.1 Pa s to over 1000 Pa s offering new opportunities for applications in confined as well as in unconfined environments.

Composite Double-Network Hydrogels To Improve Adhesion on Biological Surfaces.

Despite the development of hydrogels with high mechanical properties, insufficient adhesion between these materials and biological surfaces significantly limits their use in the biomedical field. By controlling toughening processes, we designed a composite double-network hydrogel with ∼90% water content, which creates a dissipative interface and robustly adheres to soft tissues such as cartilage and meniscus. A double-network matrix composed of covalently cross-linked poly(ethylene glycol) dimethacrylate and ionically cross-linked alginate was reinforced with nanofibrillated cellulose. No tissue surface modification was needed to obtain high adhesion properties of the developed hydrogel. Instead, mechanistic principles were used to control interfacial crack propagation. Comparing to commercial tissue adhesives, the integration of the dissipative polymeric network on the soft tissue surfaces allowed a significant increase in the adhesion strength, such as ∼130 kPa for articular cartilage. Our findings highlight the significant role of controlling hydrogel structure and dissipation processes for toughening the interface. This research provides a promising path to the development of highly adhesive hydrogels for tissues repair.

A photopolymerized composite hydrogel and surgical implanting tool for a nucleus pulposus replacement.

Nucleus pulposus replacements have been subjected to highly controversial discussions over the last 40 years. Their use has not yet resulted in a positive outcome to treat herniated disc or degenerated disc disease. The main reason is that not a single implant or tissue replacement was able to withstand the loads within an intervertebral disc. Here, we report on the development of a photo-polymerizable poly(ethylene glycol)dimethacrylate nano-fibrillated cellulose composite hydrogel which was tuned according to native tissue properties. Using a customized minimally-invasive medical device to inject and photopolymerize the hydrogel insitu, samples were implanted through an incision of 1 mm into an intervertebral disc of a bovine organ model to evaluate their long-term performance. When implanted into the bovine disc model, the composite hydrogel implant was able to significantly re-establish disc height after surgery (p < 0.0025). The height was maintained after 0.5 million loading cycles (p < 0.025). The mechanical resistance of the novel composite hydrogel material combined with the minimally invasive implantation procedure into a bovine disc resulted in a promising functional orthopedic implant for the replacement of the nucleus pulposus.

Miniature probe for the delivery and monitoring of a photopolymerizable material.

Photopolymerization is a common method to cure materials initially in a liquid state, such as dental implants or bone or tissue fillers. Recent advances in the development of biocompatible gel- and cement-systems open up an avenue for in situ photopolymerization. For minimally invasive surgery, such procedures require miniaturized surgical endoscopic probes to activate and control photopolymerization in situ. We present a miniaturized light probe in which a photoactive material can be (1) mixed, pressurized, and injected, (2) photopolymerized/photoactivated, and (3) monitored during the chemical reaction. The device is used to implant and cure poly(ethylene glycol) dimethacrylate-hydrogel-precursor in situ with ultraviolet A (UVA) light (365 nm) while the polymerization reaction is monitored in real time by collecting the fluorescence and Raman signals generated by the 532-nm excitation light source. Hydrogels could be delivered, photopolymerized, and monitored by the probe up to a curing depth of 4 cm. The size of the photopolymerized samples could be correlated to the fluorescent signal collected by the probe, and the reproducibility of the procedure could be demonstrated. The position of the probe tip inside a bovine caudal intervertebral disc could be estimated in vitro based on the collected fluorescence and Raman signal.

Photopolymerizable hydrogels for implants: Monte-Carlo modeling and experimental in vitro validation.

Photopolymerization is commonly used in a broad range of bioapplications, such as drug delivery, tissue engineering, and surgical implants, where liquid materials are injected and then hardened by means of illumination to create a solid polymer network. However, photopolymerization using a probe, e.g., needle guiding both the liquid and the curing illumination, has not been thoroughly investigated. We present a Monte Carlo model that takes into account the dynamic absorption and scattering parameters as well as solid-liquid boundaries of the photopolymer to yield the shape and volume of minimally invasively injected, photopolymerized hydrogels. In the first part of the article, our model is validated using a set of well-known poly(ethylene glycol) dimethacrylate hydrogels showing an excellent agreement between simulated and experimental volume-growth-rates. In the second part, in situ experimental results and simulations for photopolymerization in tissue cavities are presented. It was found that a cavity with a volume of 152 mm3 can be photopolymerized from the output of a 0.28-mm2 fiber by adding scattering lipid particles while only a volume of 38 mm3 (25%) was achieved without particles. The proposed model provides a simple and robust method to solve complex photopolymerization problems, where the dimension of the light source is much smaller than the volume of the photopolymerizable hydrogel.

Biodegradable polylactide/hydroxyapatite nanocomposite foam scaffolds for bone tissue engineering applications.

Supercritical carbon dioxide processing of poly-L-lactide (PLLA)/hydroxyapatite (nHA) nanocomposites was investigated as a means to prepare foams suitable as scaffolds in bone tissue engineering applications. For given foaming parameters, addition of nHA to the PLLA gave reduced cell sizes and improved homogeneity in the size distribution, but did not significantly affect the degree of crystallinity, which remained of the order of 50 wt% in all the foams. The compressive modulus and strength were primarily influenced by the porosity and there was no significant reinforcement of the matrix by the nHA. The mechanical properties of the foams were nevertheless comparable with those of trabecular bone, and by adjusting the saturation pressure and depressurization rate it was possible to generate porosities of about 85 %, an interconnected morphology and cell diameters in the range 200-400 µm from PLLA containing 4.17 vol% nHA, satisfying established geometrical requirements for bone replacement scaffolds.

Nanofibrillated cellulose composite hydrogel for the replacement of the nucleus pulposus.

The swelling and compressive mechanical behavior as well as the morphology and biocompatibility of composite hydrogels based on Tween® 20 trimethacrylate (T3), N-vinyl-2-pyrrolidone (NVP) and nanofibrillated cellulose (NFC) were assessed in the present study. The chemical structure of T3 was verified by Fourier transform infrared spectroscopy and proton nuclear magnetic resonance, and the degree of substitution was found to be around 3. Swelling ratios of neat hydrogels composed of different concentrations of T3 and NVP were found to range from 1.5 to 5.7 with decreasing concentration of T3. Various concentrations of cellulose nanofibrils (0.2-1.6wt.%) were then used to produce composite hydrogels that showed lower swelling ratios than neat ones for a given T3 concentration. Neat and composite hydrogels exhibited a typical nonlinear response under compression. All composite hydrogels showed an increase in elastic modulus compared to neat hydrogel of about 3- to 8-fold, reaching 18kPa at 0% strain and 62kPa at 20% strain for the hydrogel with the highest NFC content. All hydrogels presented a porous and homogeneous structure, with interconnected pore cells of around 100nm in diameter. The hydrogels are biocompatible. The results of this study demonstrate that composite hydrogels reinforced with NFC may be viable as nucleus pulposus implants due to their adequate swelling ratio, which may restore the annulus fibrosus loading, and their increased mechanical properties, which could possibly restore the height of the intervertebral discs.

In vivo cyclic loading as a potent stimulatory signal for bone formation inside tissue engineering scaffold.

In clinical situations, bone defects are often located at load bearing sites. Tissue engineering scaffolds are future bone substitutes and hence they will be subjected to mechanical stimulation. The goal of this study was to test if cyclic loading can be used as stimulatory signal for bone formation in a bone scaffold. Poly(L-lactic acid) (PLA)/ 5% beta-tricalcium phosphate (beta-TCP) scaffolds were implanted in both distal femoral epiphyses of eight rats. Right knees were stimulated (10N, 4Hz, 5 min) five times, every two days, starting from the third day after surgery while left knees served as control. Finite element study of the in vivo model showed that the strain applied to the scaffold is similar to physiological strains. Using micro-computed tomography (CT), all knees were scanned five times after the surgery and the related bone parameters of the newly formed bone were quantified. Statistical modeling was used to estimate the evolution of these parameters as a function of time and loading. The results showed that mechanical stimulation had two effects on bone volume (BV): an initial decrease in BV at week 2, and a long-term increase in the rate of bone formation by 28%. At week 13, the BV was then significantly higher in the loaded scaffolds.

Human fetal bone cells associated with ceramic reinforced PLA scaffolds for tissue engineering.

Fetal bone cells were shown to have an interesting potential for therapeutic use in bone tissue engineering due to their rapid growth rate and their ability to differentiate into mature osteoblasts in vitro. We describe hereafter their capability to promote bone repair in vivo when combined with porous scaffolds based on poly(l-lactic acid) (PLA) obtained by supercritical gas foaming and reinforced with 5 wt.% beta-tricalcium phosphate (TCP). Bone regeneration was assessed by radiography and histology after implantation of PLA/TCP scaffolds alone, seeded with primary fetal bone cells, or coated with demineralized bone matrix. Craniotomy critical size defects and drill defects in the femoral condyle in rats were employed. In the cranial defects, polymer degradation and cortical bone regeneration were studied up to 12 months postoperatively. Complete bone ingrowth was observed after implantation of PLA/TCP constructs seeded with human fetal bone cells. Further tests were conducted in the trabecular neighborhood of femoral condyles, where scaffolds seeded with fetal bone cells also promoted bone repair. We present here a promising approach for bone tissue engineering using human primary fetal bone cells in combination with porous PLA/TCP structures. Fetal bone cells could be selected regarding osteogenic and immune-related properties, along with their rapid growth, ease of cell banking and associated safety.

What static and dynamic properties should slalom skis possess? Judgements by advanced and expert skiers.

Flexural and torsional rigidity are important properties of skis. However, the flexural and torsional rigidity that lead to optimal performance remain to be established. In the present study, four pairs of slalom skis that differed in flexural and torsional rigidity were tested by advanced and expert skiers. Using a 10-item questionnaire, different aspects of the skis' performance were rated on a 9-point scale. For each pair of skis, physical measurements were compared with the ratings of the two groups of skiers. Correlations (Spearman) were then determined between (i) different mechanical properties of the skis (static and dynamic), (ii) subjective assessments of the participants, and (iii) properties of the skis and the participants' assessments. The latter showed that expert skiers rate the aspects of the skis more accurately than advanced skiers. Most importantly, expert skiers are particularly sensitive to torsion of the skis. These results suggest that such highly rated elements should be addressed in future ski designs.

Repair of critical size defects in the rat cranium using ceramic-reinforced PLA scaffolds obtained by supercritical gas foaming.

Bioresorbable scaffolds made of poly(L-lactic acid) (PLA) obtained by supercritical gas foaming were recently described as suitable for tissue engineering, portraying biocompatibility with primary osteoblasts in vitro and interesting mechanical properties when reinforced with ceramics. The behavior of such constructs remained to be evaluated in vivo and therefore the present study was undertaken to compare different PLA/ceramic composite scaffolds obtained by supercritical gas foaming in a critical size defect craniotomy model in Sprague-Dawley rats. The host-tissue reaction to the implants was evaluated semiquantitatively and similar tendencies were noted for all graft substitutes: initially highly reactive but decreasing with time implanted. Complete bone-bridging was observed 18 weeks after implantation with PLA/ 5 wt % beta-TCP (PLA/TCP) and PLA/5 wt % HA (PLA/HA) scaffolds as assessed by histology and radiography. We show here for the first time that this solvent-free technique provides a promising approach in tissue engineering demonstrating both the biocompatibility and osteoconductivity of the processed structures in vivo.

Architecture and properties of anisotropic polymer composite scaffolds for bone tissue engineering.

Bone is a complex porous composite structure with specific characteristics such as viscoelasticity and anisotropy, both in morphology and mechanical properties. Bone defects are regularly filled with artificial tissue grafts, which should ideally have properties similar to those of natural bone. Open cell composite foams made of bioresorbable poly(L-lactic acid) (PLA) and ceramic fillers, hydroxyapatite (HA) or beta-tricalcium phosphate (beta-TCP), were processed by supercritical CO2 foaming. Their internal 3D-structure was then analysed by micro-computed tomography (microCT), which evidenced anisotropy in morphology with pores oriented in the foaming direction. Furthermore compressive tests demonstrated anisotropy in mechanical behaviour, with an axial modulus up to 1.5 times greater than the transverse modulus. Composite scaffolds also showed viscoelastic behaviour with increased modulus for higher strain rates. Such scaffolds prepared by gas foaming of polymer composite materials therefore possess suitable architecture and properties for bone tissue engineering applications.

Biocompatibility of bioresorbable poly(L-lactic acid) composite scaffolds obtained by supercritical gas foaming with human fetal bone cells.

The aim of this investigation was to test the biocompatibility of three-dimensional bioresorbable foams made of poly(L-lactic acid) (PLA), alone or filled with hydroxyapatite (HA) or beta-tricalcium phosphate (beta-TCP), with human primary osteoblasts, using a direct contact method. Porous constructs were processed by supercritical gas foaming, after a melt-extrusion of ceramic/polymer mixture. Three neat polymer foams, with pore sizes of 170, 310, and 600 microm, and two composite foams, PLA/5 wt% HA and PLA/5 wt% beta-TCP, were examined over a 4-week culture period. The targeted application is the bone tissue-engineering field. For this purpose, human fetal and adult bone cells were chosen because of their highly osteogenic potential. The association of fetal bone cells and composite scaffold should lead to in vitro bone formation. The polymer and composite foams supported adhesion and intense proliferation of seeded cells, as revealed by scanning electron microscopy. Cell differentiation toward osteoblasts was demonstrated by alkaline phosphatase (ALP) enzymatic activity, gamma-carboxylated Gla-osteocalcin production, and the onset of mineralization. The addition of HA or beta-TCP resulted in higher ALP enzymatic activity for fetal bone cells and a stronger production of Gla-osteocalcin for adult bone cells.