Development of a dextrin-based hydrogel for bone regeneration

[Excerpt] Bone tissue engineering is a very challenging and promising field, which handles with the limitations of bone regenerative capacity and the failure of current orthopedic implants [1]. This work describes the preparation and characterization of an injectable dextrin-based hydrogel (oDex) able to incorporate nanoparticles, cells, biomolecules or Bonelike~ granules [2]. (...

Development and characterization of an injectable dextrin-based hydrogel for bone regeneration

Bone is a dynamic, highly vascularized tissue that remodels itself continuously over an individual ́s lifetime. It plays several important roles in maintaining homeostasis of the body systems [ 1 , 2 ] . However, this regenerative capac ity is limited and, as in the case of large bone defects, where the template for an orchestrated regeneration is absent, surgical proce dures are needed [ 2 ] . In this respect , bone tissue engineering is a very challe nging and promising field given the need to mimic bone mechanical and biological functions and also due to the failure of current orthoped ic implants. The general concept consists in the development of three - dimensional scaffolds, from biocompatible materials (natu ral or synthetic), which confer temporary support for the regeneration of bone tissue, while the scaffold itself will be resorbed and replaced by new ly formed tissue [ 2 , 3 ] . Hydrogels are cross - linked networks made of natural or synthetic polymers, which are able to support high water contents [ 4 ] . These materials are usua lly biocompatible, have the ability to mimic physiological conditions, promote an environment that can protect cells or unstable drugs, their physical characteristics can be controlled to some extent and some can be injected in vivo . These features make th em attractive materials in the biomedical field for cell encapsulation, drug or gene delivery or to act as an interfa ce between tissue and materials [ 4 - 7 ] . Natural polymers are advantageous for this kind of applications since they are cheap raw materials, bear a great biocompatibility and are usually biodegradable [ 8 ] . Dextrin is low molecular weight carbohydrate, generally regarded as safe (GRAS), obtained from partial hydrolysis of starch or glycogen [ 9 ] . It is a glucose polymer linked by α - 1,4 glycosidic linkages with some degree of branching due to the presence of α - 1,6 bonds [ 10 ] . I t is biocompatible and non - immunogenic, degradable by α - amylases and can undergo renal clearance avoiding tissue accumulation [ 11 , 12 ] . This work describes the preparation and characterization of an injectable dextrin - bas ed hydrogel (oDex) able to incorporate nanoparticles , cells, biomolecules or Bonelike ® granules [ 13 ] . Bonelike ® is a Biosckin - molecular and cell therapies S.A. proprietary synthetic bone graft, and the outcome of the project will result in a novel injectable presentation of this product. The hydrogel was produced by dextrin oxidation with sodium periodate followed by cross - linking with a dihydrazide [ 14 ] . In vitro characterization of oDex hydrogel has shown acceptable m echanical properties, overall good biocompatibility and the ability to be combined with other materials such as a nanogel and urinary bladder matrix, without affecting its structure. The cytotoxicity of the free dihydrazide was evaluated and only a mild in hibitory effect on cell proliferation was observed for the concentration used in the hydrogel crosslinking. The biocompatibilit y of oDex hydrogels was confirmed through the encapsulation of cells, which were able to endure the gelation process. Subcutaneou s implants were performed in Sasco Sprague Dawley rats in order to evaluate the inflammatory response and systemic effects of oDex hydrogels and their combination with Bonelike ® and human mesenchymal stem cells isolated from umbilical cord’s Wharton jelly. After 3 and 15 days post - implantation, a quantitative evaluation of both responses was performed according to ISO 10993 by a scoring system leading to a classification of the implanted material as s light irritant even when associated to Bonelike ® or to the cellular system. The performance of oDex hydrogel combined with Bonelike granules and/or UBM in bone defects was investigated in New Zealand rabbits. Bone defects in several anatomical locations (t ibiae and cranium) of non - critical and critical size were filled with those materials. Histological analysis showed that oDex does not constitute a barrier for cellular colonization and proliferation since the defects that were filled with these materials presented a higher degree of regeneration and a higher amount of collagen fibers with higher organization degrees, when compared with the empty defects. Even though oDex hydrogels purpose is to act as an injectable carrier for osteoconductive materials, li ke Bonelike ® granules, the hydrogel itself seems to assists the regenerative pro cess when compared with the empty defects. This is due to the 3D supp ort conferred by hydrogels that facilitates cell migration to the defect site. Moreover, the presence of UB M strongly stimulates the bone regeneration, for levels comparable with the Bonelike ® conditions, since an increase in cellular colonization and organization in the defect site can be denoted. A sterilization protocol for oDex hydrogels by gamma and beta r adiation was investigated through irradiation of oxidized dextrin solutions. Despite b oth kinds of radiation induced slight differences in the storage modulus of the hydrogels, indicating the occurrence of chain scission/cross - linking effects on the dextri n cha in, all materials were gelable after the irradiation treatments . These effects seem to not be dose or temperature dependent and the irradiation process in liquid or solid state also does not induce major differences in the rheology of the final hydrog els. Due to its known advantages, gamma radiation seems to be a suitable sterilization method for oxidized dextrin solutions. The stability of gamma irradiated dextrin solutions was evaluated up to 8 months. Despite the increase of storage modulus of the h ydrogels over the time, this effect does not constitute a disadvantage since it improves elastic behavior of the hydrogels. oDex hydrogels provides a system that can carry and stabilize cells, nanogels, Bonelike ® granules and other biomolecules. It is a pr omising biomaterial due to its biocompatibility, and potential to promote an adequate environment for bone regeneration. Its injectability allows a minimal invasive surgical procedure with decreased patient morbidity, lower risk of infection and reduced sc ar formation. This work has been developed in the scope of an European project that allowed collaborations with research groups, which have complementary expertise. The tight collaboration between University of Minho and Bioskin S.A. company, envisioning t echnology transfer and product valorization, has resulted in a published international patent of the product ( WO2011070529A2 ) [ 15 ] . Currently, a new set of pre - clinical trials in sheep model s are being planned as well as the submission of a request for the authorization for the clinical trialsGrant SFRH/BD/64571/2009 from Fundação para a Ciência e Tecnologia (FCT), Portugal. We thank FCT funding through EuroNanoMed ENMED/0002/2

Potential of injectable dextrin-based hydrogel for biomedical applications

Bone tissue engineering is a very challenging and promising field, which handles with the limitations of bone regenerative capacity and the failure of current orthopedic implants [1]. This work describes the preparation and characterization of an injectable dextrinbased hydrogel (oDex) through dextrin oxidation followed by cross-linking with a dihydrazide [2]. In vitro and in vivo experiments allowed to conclude that this system can carry and stabilize cells, nanogels, Bonelike® granules [3] and other biomolecules. This is a promising biomaterial due to its biocompatibility, and potential to promote an adequate environment for bone regeneration, which was increased by the combined bioactive molecules. Its injectability allows a minimal invasive surgical procedure with decreased patient morbidity, lower infection risk and reduced scar formation. Furthermore, an adequate sterilization protocol for this kind of material was established. The tight collaboration between University of Minho and Bioskin S.A. company, envisioning technology transfer and product valorization, has resulted in a published international patent of the product (WO2011070529A2) [4]. Currently, the submission of a request for the authorization for the clinical trials is being planned

Intracellular T lymphocyte cytokine profiles in the aqueous humour of patients with uveitis and correlation with clinical phenotype

This study aimed to investigate whether T cells in aqueous humour are different in different types of uveitis and correlate with clinical phenotype. Patients with clinically different types of uveitis, but all displaying active anterior uveitis, were phenotyped and samples of aqueous humour (AH) and peripheral blood (PB) collected. Cells from AH and PB were separated by centrifugation and by density gradient centrifugation (to obtain mononuclear cells PBMC), respectively. Cells were activated with PMA and ionomycin in the presence of Brefeldin A, stained for surface markers and intracellular cytokines, and analysed by flow cytometry. The cytokine profile was correlated with the clinical phenotype. Increased percentages of interleukin (IL)-10(+)-, but not interferon (IFN)-γ(+) T lymphocytes were found in AH compared with PB in patients with acute anterior uveitis (AAU), FHC or chronic panuveitis (PU). There was a trend towards elevated levels of IL-10(+) T cells in AH from patients with FHC compared with AH from acute uveitis and panuveitis patients. Increased levels of IL-10(+) T cells in AH compared with PB were also found in samples from patients with isolated uveitis, but not those with associated systemic disease. Levels of cytokine-positive T cells were not associated with the use of topical steroids or to the severity of the anterior uveitis. While type I cytokine-producing T lymphocytes are present in AH during AU, the presence of increased proportions of IL-10(+) T lymphocytes in AH from patients with uveitis may be indicative of an anti-inflammatory mechanism that may influence the type and course of ocular inflammation in these patients

Infliximab treatment for ocular and extraocular manifestations of Behçet's disease

Purpose: To assess the efficacy and safety of infliximab in the treatment of sight-threatening uveitis and extraocular manifestations in patients with Behçet's disease. Methods: Twelve patients with Behçet's disease and uveitis were treated with infliximab after unsuccessful therapy with other immunosuppressive drugs. The main outcome measures were as follows: the number of uveitis relapses, the number of Behçet's disease-related extraocular lesions, and the amount of corticosteroids administered during the treatment as well as during an equal prior period of time while the patients were on other immunosuppressive agents. Visual acuity was recorded at the beginning of infliximab therapy and at the end of follow-up, and was defined as stable if it did not change from baseline, increased if it showed at least one line of improvement from baseline, and decreased if it showed at least a one line decrease from baseline. Results: During an average follow-up of 16.67 ± 7.63 months (median, 15 months), 11 patients (91.6%) showed a reduction in the number of ocular relapses (relapse/month, from 0.35 ± 0.17 to 0.12 ± 0.17, P < 0.001). All of the patients (n = 11) who were taking corticosteroids before infliximab were able to reduce the amount of corticosteroids taken daily during infliximab treatment (from 24.33 ± 10.84∈mg/prednisone per day to 8.97 ± 6.81∈mg/prednisone per day, P < 0.001), and all presented with a reduced onset of extraocular manifestations of Behçet's disease (mean total number, from 2.83 ± 3.61 to 1.51 ± 2.35, P = 0.039). One patient, who had to stop treatment 2 months after starting because of the onset of pulmonary tuberculosis, showed the same number of relapses during infliximab treatment but was able to reduce the mean daily corticosteroid dose. Visual acuity increased by one or more lines in three eyes (12.5%) and remained unchanged in 87.5% of the eyes. Infliximab-related side effects appeared in four patients (33.3%). Conclusions: Infliximab was effective in the treatment of uveitis in these Behçet's disease patients, significantly reducing the number of ocular relapses and making possible a significant reduction in the daily dose of corticosteroids administered. Extraocular manifestations of Behçet's disease were also controlled by infliximab. Nevertheless, side effects were not uncommon, and an extensive study of systemic conditions before infliximab administration had to be carried out to exclude systemic infection, particularly prior tuberculosis. © Japanese Ophthalmological Society 2007