A Novel 3D Cultured Model for Studying Early Changes in Age-Related Macular Degeneration.

Various in vitro culture systems have been used to investigate the pathogenesis of age-related macular degeneration (AMD). However, many still rely on oversimplified monolayer culture models. AMD is a complex disease, associated with the pathological changes to multiple structural components such as the Bruch's membrane, retinal pigment epithelium (RPE), and choroidal endothelial cells. This study aims to construct a novel 3D coculture model using the polycaprolactone (PCL)-gelatin electrospun scaffold, with human RPE cells (hRPE) and primate choroidal cells (RF-6A). Results from this study show that PCL-gelatin scaffolds have a highly porous ultrastructure that supports the attachment, proliferation, differentiation, and migration of the hRPEs and choroidal endothelial cells. It is also demonstrated that the PCL-gelatin 3D coculture model may be useful in exploring the molecular interplay between the hPRE and the choroidal endothelial cells, and their effects on growth factor modulation, which may be important in the pathogenesis of AMD.

Reinforcement of hydrogels using three-dimensionally printed microfibres.

Despite intensive research, hydrogels currently available for tissue repair in the musculoskeletal system are unable to meet the mechanical, as well as the biological, requirements for successful outcomes. Here we reinforce soft hydrogels with highly organized, high-porosity microfibre networks that are 3D-printed with a technique termed as melt electrospinning writing. We show that the stiffness of the gel/scaffold composites increases synergistically (up to 54-fold), compared with hydrogels or microfibre scaffolds alone. Modelling affirms that reinforcement with defined microscale structures is applicable to numerous hydrogels. The stiffness and elasticity of the composites approach that of articular cartilage tissue. Human chondrocytes embedded in the composites are viable, retain their round morphology and are responsive to an in vitro physiological loading regime in terms of gene expression and matrix production. The current approach of reinforcing hydrogels with 3D-printed microfibres offers a fundament for producing tissue constructs with biological and mechanical compatibility.

Multiphasic construct studied in an ectopic osteochondral defect model.

In vivo osteochondral defect models predominantly consist of small animals, such as rabbits. Although they have an advantage of low cost and manageability, their joints are smaller and more easily healed compared with larger animals or humans. We hypothesized that osteochondral cores from large animals can be implanted subcutaneously in rats to create an ectopic osteochondral defect model for routine and high-throughput screening of multiphasic scaffold designs and/or tissue-engineered constructs (TECs). Bovine osteochondral plugs with 4 mm diameter osteochondral defect were fitted with novel multiphasic osteochondral grafts composed of chondrocyte-seeded alginate gels and osteoblast-seeded polycaprolactone scaffolds, prior to being implanted in rats subcutaneously with bone morphogenic protein-7. After 12 weeks of in vivo implantation, histological and micro-computed tomography analyses demonstrated that TECs are susceptible to mineralization. Additionally, there was limited bone formation in the scaffold. These results suggest that the current model requires optimization to facilitate robust bone regeneration and vascular infiltration into the defect site. Taken together, this study provides a proof-of-concept for a high-throughput osteochondral defect model. With further optimization, the presented hybrid in vivo model may address the growing need for a cost-effective way to screen osteochondral repair strategies before moving to large animal preclinical trials.

Perspectives in multiphasic osteochondral tissue engineering.

Critical-sized osteochondral defects are clinically challenging, with limited treatment options available. By engineering osteochondral grafts using a patient's own cells and osteochondral scaffolds designed to facilitate cartilage and bone regeneration, osteochondral defects may be treated with less complications and better long-term clinical outcomes. Scaffolds can influence the development and structure of the engineered tissue, and there is an increased awareness that osteochondral tissue engineering concepts need to take the in vivo complexities into account in order to increase the likelihood of successful osteochondral tissue repair. The developing trend in osteochondral tissue engineering is the utilization of multiphasic scaffolds to recapitulate the multiphasic nature of the native tissue. Cartilage and bone have different structural, mechanical, and biochemical microenvironments. By designing osteochondral scaffolds with tissue-specific architecture, it may be possible to enhance osteochondral repair within shorter timeframe. While there are promising in vivo outcomes using multiphasic approaches, functional regeneration of osteochondral constructs still remains a challenge. In this review, we provide an overview of in vivo osteochondral repair studies that have taken place in the past three years, and define areas which needs improvement in future studies.

Effect of preculture and loading on expression of matrix molecules, matrix metalloproteinases, and cytokines by expanded osteoarthritic chondrocytes.

OBJECTIVE: One of the pathologic changes that occurs during osteoarthritis (OA) is the degeneration of the pericellular matrix (PCM). Since the PCM is likely to be involved in mechanotransduction, this study was undertaken to investigate the effects of PCM-like matrix accumulation in zonal OA chondrocytes and their influence on chondrocyte response to compression. METHODS: Superficial and middle/deep zone chondrocytes from macroscopically normal cartilage of OA knees were expanded and encapsulated in alginate gels. The effects of compression (short-term or long-term) and preculture on chondrocyte expression of various matrix molecules, cytokines, and matrix metalloproteinases (MMPs) were assessed. Additionally, nonexpanded chondrocytes were encapsulated in alginate and cultured in the presence or absence of transforming growth factor ß1 (TGFß1) and dexamethasone and analyzed following short-term compression experiments. RESULTS: Expanded OA chondrocytes (superficial and middle/deep zone) that were precultured for 2 weeks under free-swelling conditions prior to dynamic compression responded more sensitively to loading and had increased matrix accumulation, increased interleukin-1ß (IL-1ß) and IL-4 levels, and decreased levels of MMP-2 (in the middle/deep zone) compared to the nonloaded controls. Compression also decreased MMP-3 and MMP-13 levels even without preculture. Nonexpanded chondrocytes did not respond to compression, but differences in gene expression were found depending on the zone of harvest, time in culture, and medium composition. CONCLUSION: Our findings demonstrate that with predeposited PCM-like matrix, compressive stimulation can enhance matrix protein accumulation in expanded OA chondrocytes. Investigations into how PCM or other matrix components differentially affect this balance under mechanical loading may provide invaluable insight into OA pathogenesis and the use of expanded cells in tissue engineering and regenerative medicine-based applications.