Optimizing the medium perfusion rate in bone tissue engineering bioreactors

Abstract

There is a critical need to increase the size of bone grafts that can be cultured in vitro for use in regenerative medicine. Perfusion bioreactors have been used to improve the nutrient and gas transfer capabilities and reduce the size limitations inherent to static culture, as well as to modulate cellular responses by hydrodynamic shear. Our aim was to understand the effects of medium flow velocity on cellular phenotype and the formation of bone-like tissues in three-dimensional engineered constructs. We utilized custom-designed perfusion bioreactors to culture bone constructs for five weeks using a wide range of superficial flow velocities (80, 400, 800, 1200 and 1800 μm/s), corresponding to estimated initial shear stresses ranging from 0.6 – 20 mPa. Increasing the flow velocity significantly affected cell morphology, cell-cell interactions, matrix production and composition, and the expression of osteogenic genes. Within the range studied, the flow velocities ranging from 400 to 800 μm/s yielded the best overall osteogenic responses. Using mathematical models, we determined that even at the lowest flow-velocity (80 μm/s) the oxygen provided was sufficient to maintain viability of the cells within the construct. Yet it was clear that this flow-velocity did not adequately support the development of bone-like tissue. The complexity of the cellular responses found at different flow-velocities underscores the need to use a range of evaluation parameters to determine the quality of engineered-bone.

Three-Dimensional Characterization of Tissue-Engineered Constructs by Contrast-Enhanced Nanofocus Computed Tomography

“…These dynamics may be attributed to fluid flow and convective nutrient and oxygen transport throughout the TE construct volume during culture. [40][41][42]46 In this study, CEnano-CT revealed a dramatic increase of ECM content for bioreactor cultured TE constructs (up to 75% of void volume) with respect to the statically cultured ones (less than 4% of void volume). The low ECM quantity in the static culture case was below the CE-nano-CT threshold for ECM quantification, which in this study was determined to be an ECM volume lower than 4% of the total Ti scaffold void volume (Fig.…”

Engineering bone tissue substitutes from human induced pluripotent stem cells

“…We previously reported that perfusion culture is critical for engineering large compact bone grafts from adult and hESC-derived mesenchymal progenitors (2,9,25). In the present study we extended our osteoconductive scaffold-perfusion bioreactor culture model to test the bone-forming potential of the mesenchymal progenitors derived from the hiPSC lines BC1 and 1013A, and hESC line H9, which exhibited excellent osteogenic potential in monolayer assays ( Fig.…”

Bioreactor Systems for Human Bone Tissue Engineering

“…Using a biomimetic approach of bone engineering, Grayson et al demonstrated the effect of perfusion rate on BM-derived hMSCs seeded onto decellularized bone plugs (4 mm diameter and 4 mm height) during a five-week perfusion culture [67]. The authors found that increasing the flow velocity of perfusion medium from 80 to 1800 μm/s (corresponding to estimated initial shear stresses ranging from 0.0006 to 0.02 Pa) significantly affected cell morphology, cell-cell interactions, matrix production and composition, and the expression of osteogenic genes and that intermediate flow velocities (400 to 800 μm/s) yielded the best osteogenic outcomes.…”