3D bioprinting of methacrylated hyaluronic acid (MeHA) hydrogel with intrinsic osteogenicity.

In bone regenerative medicine there is a need for suitable bone substitutes. Hydrogels have excellent biocompatible and biodegradable characteristics, but their visco-elastic properties limit their applicability, especially with respect to 3D bioprinting. In this study, we modified the naturally occurring extracellular matrix glycosaminoglycan hyaluronic acid (HA), in order to yield photo-crosslinkable hydrogels with increased mechanical stiffness and long-term stability, and with minimal decrease in cytocompatibility. Application of these tailor-made methacrylated hyaluronic acid (MeHA) gels for bone tissue engineering and 3D bioprinting was the subject of investigation. Visco-elastic properties of MeHA gels, measured by rheology and dynamic mechanical analysis, showed that irradiation of the hydrogels with UV light led to increased storage moduli and elastic moduli, indicating increasing gel rigidity. Subsequently, human bone marrow derived mesenchymal stromal cells (MSCs) were incorporated into MeHA hydrogels, and cell viability remained 64.4% after 21 days of culture. Osteogenic differentiation of MSCs occurred spontaneously in hydrogels with high concentrations of MeHA polymer, in absence of additional osteogenic stimuli. Addition of bone morphogenetic protein-2 (BMP-2) to the culture medium further increased osteogenic differentiation, as evidenced by increased matrix mineralisation. MeHA hydrogels demonstrated to be suitable for 3D bioprinting, and were printed into porous and anatomically shaped scaffolds. Taken together, photosensitive MeHA-based hydrogels fulfilled our criteria for cellular bioprinted bone constructs within a narrow window of concentration.

Prolonged presence of VEGF promotes vascularization in 3D bioprinted scaffolds with defined architecture.

Timely vascularization is essential for optimal performance of bone regenerative constructs. Vascularization is efficiently stimulated by vascular endothelial growth factor (VEGF), a substance with a short half-life time. This study investigates the controlled release of VEGF from gelatin microparticles (GMPs) as a means to prolong VEGF activity at the preferred location within 3D bioprinted scaffolds, and the effects on subsequent vascularization. The release of VEGF from GMPs was continuous for 3 weeks during in vitro studies, and bioactivity was confirmed using human endothelial progenitor cells (EPCs) in migration assays. Traditional and real-time migration assays showed immediate and efficient EPC migration in the presence of GMP-released VEGF, indistinguishable from VEGF-solution that was added to the medium. Matrigel scaffolds containing EPCs and VEGF, which was released either in a fast or sustained fashion by application of GMPs, were investigated for their in vivo vasculogenic capacity. Implantation in subcutaneous pockets in nude mice for one week demonstrated that vessel formation was significantly higher in the VEGF sustained-release group compared to the fast release group. In addition, regional differences with respect to VEGF release were introduced in 3D bioprinted EPC-laden scaffolds and their influence on vasculogenesis was investigated in vivo. The different regions were retained and vessel formation occurred analogous with the results seen in the Matrigel plugs. We conclude that GMPs are suitable to generate sustained release profiles of bioactive VEGF, and that they can be used to create defined differentiation regions in 3D bioprinted heterogeneous constructs, allowing a new generation of smart scaffold design. The prolonged presence of VEGF led to a significant increase in scaffold vascularization when applied in vivo.

Sustained release of BMP-2 in bioprinted alginate for osteogenicity in mice and rats.

The design of bioactive three-dimensional (3D) scaffolds is a major focus in bone tissue engineering. Incorporation of growth factors into bioprinted scaffolds offers many new possibilities regarding both biological and architectural properties of the scaffolds. This study investigates whether the sustained release of bone morphogenetic protein 2 (BMP-2) influences osteogenicity of tissue engineered bioprinted constructs. BMP-2 loaded on gelatin microparticles (GMPs) was used as a sustained release system, which was dispersed in hydrogel-based constructs and compared to direct inclusion of BMP-2 in alginate or control GMPs. The constructs were supplemented with goat multipotent stromal cells (gMSCs) and biphasic calcium phosphate to study osteogenic differentiation and bone formation respectively. BMP-2 release kinetics and bioactivity showed continuous release for three weeks coinciding with osteogenicity. Osteogenic differentiation and bone formation of bioprinted GMP containing constructs were investigated after subcutaneous implantation in mice or rats. BMP-2 significantly increased bone formation, which was not influenced by the release timing. We showed that 3D printing of controlled release particles is feasible and that the released BMP-2 directs osteogenic differentiation in vitro and in vivo.

Guideline on anterior cruciate ligament injury.

The Dutch Orthopaedic Association has a long tradition of development of practical clinical guidelines. Here we present the recommendations from the multidisciplinary clinical guideline working group for anterior cruciate ligament injury. The following 8 clinical questions were formulated by a steering group of the Dutch Orthopaedic Association. What is the role of physical examination and additional diagnostic tools? Which patient-related outcome measures should be used? What are the relevant parameters that influence the indication for an ACL reconstruction? Which findings or complaints are predictive of a bad result of an ACL injury treatment? What is the optimal timing for surgery for an ACL injury? What is the outcome of different conservative treatment modalities? Which kind of graft gives the best result in an ACL reconstruction? What is the optimal postoperative treatment concerning rehabilitation, resumption of sports, and physiotherapy? These 8 questions were answered and recommendations were made, using the "Appraisal of Guidelines for Research and Evaluation" instrument. This instrument seeks to improve the quality and effectiveness of clinical practical guidelines by establishing a shared framework to develop, report, and assess. The steering group has also developed 7 internal indicators to aid in measuring and enhancing the quality of the treatment of patients with an ACL injury, for use in a hospital or practice.

Treatment of lymphatic malformations with OK-432 (Picibanil): review of the literature.

INTRODUCTION: Lymphatic malformations (LM) are benign structural defects that can cause serious complications because of their size and location. Traditionally, surgical removal was the first treatment modality, but this could be associated with many complications and risks. Since Ogita introduced OK-432 (picibanil) in 1987 as a treatment method, this sclerosant has become popular. This paper is a review of the trials published so far on this topic. PATIENTS AND METHODS: A literature search of English trials with 5 or more patients in it with LM who had never been treated before was done. The paper had to use the microcystic-macrocystic classification and have a mean follow-up of more than a year to be included in this review. Results were classified as "excellent" when the lesions show a regression of more than 90%, "good" when regression is more than 50%, and "poor" when shrinkage is less than 50% (this also includes no response at all). RESULTS: Twenty-seven percent of microcystic LMs show an excellent result; 33%, a good result; and 40%, a poor result. Of the macrocystic LMs, 88% have excellent results. Recurrence rates vary from 5% to 8%. The adverse effects are mostly mild. DISCUSSION: Most trials have a short follow-up; therefore, there are uncertainties when it comes to cure and regression. Mostly, the adverse effects of OK-432 are trivial and disappear after a week, but the need for a temporary tracheostomy has been described. Screening for allergic reactions to penicilline is needed, with the risk of anaphylactic shock in mind. It is difficult to compare the different techniques used by the authors, and none of the trials included in this study are randomized controlled trials; most are retrospective and were so-called level 4 studies. CONCLUSIONS: This review demonstrates that OK-432 is an effective way to treat LM. Because of a possible risk of airway obstruction, treatment should always take place in specialized treatment facilities. Macrocystic lesions show a better response to OK-432 treatment than microcystic lesions. Serious complications with OK-432 are infrequent, and this type of sclerotherapy seems to have no influence on future surgery. We therefore suggest the use of OK-432 as an effective first-line treatment of LMs.