A perfusion bioreactor for engineering bone constructs: an in vitro and in vivo study.

A perfusion bioreactor, which was designed based on fluidized bed concepts, was validated for the culture of bone constructs of clinically relevant size. For this study, natural coral has been used as three-dimensional scaffolds. This biomaterial is a microporous, biocompatible, osteoconductive, and absorbable scaffold. This perfusion bioreactor provided a stable environment in terms of osmolarity, pH, and, most importantly, oxidative stress. Bone constructs engineered in this system resulted in significantly higher cell proliferation and homogenous cell distribution than those cultured under static conditions. Particularly relevant to the production of bioengineered bone in a clinical setting, custom-made bone constructs (each one with volume up to 30 cm(3)) could be produced using a such perfusion bioreactor. Last, but not least, the bone constructs of clinically relevant volume thus produced were shown to be osteogenic when transplanted subcutaneously in sheep.

Theoretical analysis of engineered cartilage oxygenation: influence of construct thickness and media flow rate.

A novel parallel-plate bioreactor has been shown to modulate the mechanical and biochemical properties of engineered cartilage by the application of fluid-induced shear stress. Flow or perfusion bioreactors may improve tissue development via enhanced transport of nutrients or gases as well as the application of mechanical stimuli, or a combination of these factors. The goal of this study was to complement observed experimental responses to flow by simulating oxygen transport within cartilage constructs of different thicknesses (250 microm or 1 mm). Using numerical computation of convection-diffusion equations, the evaluation of the tissue oxygenation is performed. Four culture conditions are defined based on tissue thickness and flow rates ranging from 0 to approximately 25 mL min(-1). Under these experimental conditions results show a mean oxygen concentration within the tissue varying from 0.01 to 0.19 mol m(-3) as a function of the tissue thickness and the magnitude of the applied shear stress. More generally, the influence of shear stress varying (via flow rate modification) from 10(-3) to 10 dynes cm(-2) on the tissue oxygenation is studied. The influence on the results of important physical parameters such as the maximal oxygen consumption rate of cells is discussed. Lastly, the importance of oxygen concentration in the lower chamber and its relevance to tissue oxygenation are highlighted by the model results.

In vitro acoustic waves propagation in human and bovine cancellous bone.

UNLABELLED: The acoustic behavior of cancellous bone with regard to its complex poroelastic nature has been investigated. The existence of two longitudinal modes of propagation is demonstrated in both bovine and human cancellous bone. Failure to take into account the presence of these two waves may result in inaccurate material characterization. INTRODUCTION: Acoustic wave propagation is now a commonly used nondestructive method for cancellous bone characterization. However, wave propagation in this material may be affected by fluid-solid interactions inherent to its poroelastic nature, resulting in two different longitudinal waves. This phenomenon has been demonstrated in previous studies and is in agreement with Biot's theory. The purpose of this paper is to extend these findings to human trabecular bone and to thoroughly investigate these two waves. MATERIALS AND METHODS: Sixty human and 14 bovine cancellous bone cubic specimens were tested in vitro in three different directions using an immersion acoustic transmission method. Original procedures were developed to quantify both velocity and attenuation characteristics of each wave. In term of attenuation, a modified broadband ultrasound attenuation (BUA), describing the rate of change of the frequency-dependent attenuation, was defined for each wave (FDUA). RESULTS: Both waves were identified in most of the specimens. The fast wave velocities demonstrated a negative linear correlation with porosity (1500-2300 m/s, R2 = 0.44, p < 10(-3)), whereas the slow wave velocities exhibited two different behaviors: (1) a first set of data clearly dependent on porosity showing a positive linear correlation (1150-1450 m/s, R2 = 0.26, p < 10(-3)) and (2) a second group independent on porosity. The fast wave FDUA (20-140 dB/cmMHz) showed a parabolic behavior and reached a maximum for 75% porosity (second degree relationship R2 = 0.41,p < 10(-3)), whereas a positive linear behavior was observed for the slow wave FDUA (15-40 dB/cmMHz; R2 = 0.15, p < 10(-2)). CONCLUSIONS: Existence of two wave propagation modes were demonstrated in human cancellous bone. Our data suggest that, in some cases, the amplitude of the slow wave is much larger than the amplitude of the fast wave. For this reason, care should be taken when using measurement systems that incorporate simple threshold detection because the fast wave could remain undetected. Moreover, failure to consider the presence of these two waves could result in an inaccurate quantification of cancellous bone physical properties.

A cellular tensegrity model to analyse the structural viscoelasticity of the cytoskeleton.

This study describes the viscoelastic properties of a refined cellular-tensegrity model composed of six rigid bars connected to a continuous network of 24 viscoelastic pre-stretched cables (Voigt bodies) in order to analyse the role of the cytoskeleton spatial rearrangement on the viscoelastic response of living adherent cells. This structural contribution was determined from the relationships between the global viscoelastic properties of the tensegrity model, i.e., normalized viscosity modulus (eta(*)), normalized elasticity modulus (E(*)), and the physical properties of the constitutive elements, i.e., their normalized length (L(*)) and normalized initial internal tension (T(*)). We used a numerical method to simulate the deformation of the structure in response to different types of loading, while varying by several orders of magnitude L(*) and T(*). The numerical results obtained reveal that eta(*) remains almost independent of changes in T(*) (eta(*) proportional, variant T(*+0.1)), whereas E(*) increases with approximately the square root of the internal tension T(*) (from E(*) proportional, variant T(*+0.3) to E(*) proportional, variant T(*+0.7)). Moreover, structural viscosity eta(*) and elasticity E(*) are both inversely proportional to the square of the size of the structure (eta(*) proportional, variant L(*-2) and E(*) proportional, variant L(*-2)). These structural properties appear consistent with cytoskeleton (CSK) mechanical properties measured experimentally by various methods which are specific to the CSK micromanipulation in living adherent cells. Present results suggest, for the first time, that the effect of structural rearrangement of CSK elements on global CSK behavior is characterized by a faster cellular mechanical response relatively to the CSK element response, which thus contributes to the solidification process observed in adherent cells. In extending to the viscoelastic properties the analysis of the mechanical response of the cellular 30-element tensegrity model, the present study contributes to the understanding of recent results on the cellular-dynamic response and allows to reunify the scattered data reported for the viscoelastic properties of living adherent cells.

Interrelations between elastic energy and strain in a tensegrity model: contribution to the analysis of the mechanical response in living cells.

Interactions between the physical and physiological properties of cellular sub-units result in changes in the shape and mechanical behaviour of living tissues. To understand the mechanotransmission processes, models are needed to describe the complex interrelations between the elements and the cytoskeletal structure. In this study, we used a 30-element tensegrity structure to analyse the influence of the type of loading on the mechanical response and shape changes of the cell. Our numerical results, expressed in terms of strain energy as a function of the overall deformation of the tensegrity structure, suggest that changes in cell functions during mechanical stimuli for a given potential energy are correlated to the type of loading applied, which determines the resultant changes in cell shape. The analysis of these cellular deformations may explain the large variability in the response of bone cells submitted to different types of mechanical loading.