Regulation of endothelial cell arrangements within hMSC - HUVEC co-cultured aggregates.

BACKGROUND: Micro-mass culturing or cellular aggregation is an effective method used to form mineralised bone tissue. Poor core cell viability, however, is often an impeding characteristic of large micro-mass cultures, and equally for large tissue-engineered bone grafts. Because of this, efforts are being made to enhance large graft perfusion, often through pre-vascularisation, which involves the co-culture of endothelial cells and bone cells or stem cells. METHODS: This study investigated the effects of different aggregation techniques and culture conditions on endothelial cell arrangements in mesenchymal stem cell and human umbilical vein endothelial cell co-cultured aggregates when endothelial cells constituted just 5%. Two different cellular aggregation techniques, i.e. suspension culture aggregation and pellet culture aggregation, were applied alongside two subsequent culturing techniques, i.e. hydrostatic loading and static culturing. Endothelial cell arrangements were assessed under such conditions to indicate potential pre-vascularisation. RESULTS: Our study found that the suspension culture aggregates cultured under hydrostatic loading offered the best environment for enhanced endothelial cell regional arrangements, closely followed by the pellet culture aggregates cultured under hydrostatic loading, the suspension culture aggregates cultured under static conditions, and the pellet culture aggregates cultured under static conditions. CONCLUSIONS: The combination of particular aggregation techniques with dynamic culturing conditions appeared to have a synergistic effect on the cellular arrangements within the co-cultured aggregates.

The Use of Finite Element Analyses to Design and Fabricate Three-Dimensional Scaffolds for Skeletal Tissue Engineering.

Computational modeling has been increasingly applied to the field of tissue engineering and regenerative medicine. Where in early days computational models were used to better understand the biomechanical requirements of targeted tissues to be regenerated, recently, more and more models are formulated to combine such biomechanical requirements with cell fate predictions to aid in the design of functional three-dimensional scaffolds. In this review, we highlight how computational modeling has been used to understand the mechanisms behind tissue formation and can be used for more rational and biomimetic scaffold-based tissue regeneration strategies. With a particular focus on musculoskeletal tissues, we discuss recent models attempting to predict cell activity in relation to specific mechanical and physical stimuli that can be applied to them through porous three-dimensional scaffolds. In doing so, we review the most common scaffold fabrication methods, with a critical view on those technologies that offer better properties to be more easily combined with computational modeling. Finally, we discuss how modeling, and in particular finite element analysis, can be used to optimize the design of scaffolds for skeletal tissue regeneration.

Influence of Additive Manufactured Scaffold Architecture on the Distribution of Surface Strains and Fluid Flow Shear Stresses and Expected Osteochondral Cell Differentiation.

Scaffolds for regenerative medicine applications should instruct cells with the appropriate signals, including biophysical stimuli such as stress and strain, to form the desired tissue. Apart from that, scaffolds, especially for load-bearing applications, should be capable of providing mechanical stability. Since both scaffold strength and stress-strain distributions throughout the scaffold depend on the scaffold's internal architecture, it is important to understand how changes in architecture influence these parameters. In this study, four scaffold designs with different architectures were produced using additive manufacturing. The designs varied in fiber orientation, while fiber diameter, spacing, and layer height remained constant. Based on micro-CT (µCT) scans, finite element models (FEMs) were derived for finite element analysis (FEA) and computational fluid dynamics (CFD). FEA of scaffold compression was validated using µCT scan data of compressed scaffolds. Results of the FEA and CFD showed a significant impact of scaffold architecture on fluid shear stress and mechanical strain distribution. The average fluid shear stress ranged from 3.6 mPa for a 0/90 architecture to 6.8 mPa for a 0/90 offset architecture, and the surface shear strain from 0.0096 for a 0/90 offset architecture to 0.0214 for a 0/90 architecture. This subsequently resulted in variations of the predicted cell differentiation stimulus values on the scaffold surface. Fluid shear stress was mainly influenced by pore shape and size, while mechanical strain distribution depended mainly on the presence or absence of supportive columns in the scaffold architecture. Together, these results corroborate that scaffold architecture can be exploited to design scaffolds with regions that guide specific tissue development under compression and perfusion. In conjunction with optimization of stimulation regimes during bioreactor cultures, scaffold architecture optimization can be used to improve scaffold design for tissue engineering purposes.

Mold-Based Application of Laser-Induced Periodic Surface Structures (LIPSS) on Biomaterials for Nanoscale Patterning.

Laser-induced periodic surface structures (LIPSS) are highly regular, but at the same time contain a certain level of disorder. The application of LIPSS is a promising method to functionalize biomaterials. However, the absorption of laser energy of most polymer biomaterials is insufficient for the direct application of LIPSS. Here, we report the application of LIPSS to relevant biomaterials using a two-step approach. First, LIPSS are fabricated on a stainless steel surface. Then, the structures are replicated onto biomaterials using the steel as a mold. Results show that LIPSS can be transferred successfully using this approach, and that human mesenchymal stromal cells respond to the transferred structures. With this approach, the range of biomaterials that can be supplied with LIPSS increases dramatically.

Degradable amorphous scaffolds with enhanced mechanical properties and homogeneous cell distribution produced by a three-dimensional fiber deposition method.

The mechanical properties of amorphous, degradable, and highly porous poly(lactide-co-caprolactone) structures have been improved by using a 3D fiber deposition (3DF) method. Two designs of 3DF scaffolds, with 45° and 90° layer rotation, were printed and compared with scaffolds produced by a salt-leaching method. The scaffolds had a porosity range from 64% to 82% and a high interconnectivity, measured by micro-computer tomography. The 3DF scaffolds had 8-9 times higher compressive stiffness and 3-5 times higher tensile stiffness than the salt-leached scaffolds. There was a distinct decrease in the molecular weight during printing as a consequence of the high temperature. The chain microstructure was, however, not affected; the glass transition temperature and the decomposition temperature were constant. Human OsteoBlast-like cells were cultured in vitro and the cell morphology and distribution were observed by scanning electron microscopy and fluorescence microscopy. The cell distribution on the 3DF scaffolds was more homogeneous than the salt-leached scaffolds, suggesting that 3DF scaffolds are more suitable as porous biomaterials for tissue engineering. These results show that it is possible to design and optimize the properties of amorphous polymer scaffolds. The 3DF method produce amorphous degradable poly(lactide-co-caprolactone) that are strong and particularly suitable for cell proliferation.