Design and integration of a novel bioreactor for articular cartilage tissue manufacturing with real-time feedback of chondrogenic gene expression.

Abstract

Differentiating mesenchymal stromal cells into healthy chondrocytes for articular cartilage (AC) requires a variety of biochemical and biomechanical stimuli. While the effects of such stimuli on chondrogenic markers are well-studied, no bioreactor has thus far collected real-time data from mRNA expression for use in closed-loop control. Here, we present the design of an automated bioreactor that applies perfusion, oscillating hydrostatic pressure (OHP), and varying concentrations of multiple biochemical growth factors to the cell cultures while measuring fluorescent markers of gene expression in real-time. Our system achieves a gradated hydrodynamic environment across the length of cell cultures with flow rates between 0.46 and 3.2 cm s^-1and shear stresses between 5.0 and 60 mPa, can apply OHP at up to 0.5 Hz and 4.83 MPa, and can dynamically control the concentrations of two biochemical growth factors. The integrated fluorescent fiberscope system can measure fluorescent dye concentrations as low as 3 nm within the cell chamber. Our LNA/DNA nano-biosensor is designed to track Sox9 mRNA, a gene critical to the chondrogenic process. The reactor is controlled via MATLAB Simulink and includes remote observation and control features to increase flexibility and minimize downtime. This novel bioreactor provides a platform for further AC research with the ultimate design goal of generating a transfer function that maps growth factor inputs to mRNA expression outputs for real-time control.