An optically accessible two-chamber perfusion bioreactor for label-free metabolic imaging of three-dimensional tissue models under dynamic flow conditions

Bioreactors play an important role in tissue engineering as they tackle challenges that arise when developing three-dimensional (3D) tissue models. Bioreactor design is a complex task due to the combination of technology-specific and tissue-specific requirements, with general requirements like biocompatibility and sterility. Here we present a novel design for a two-chamber perfusion bioreactor that provides optical accessibility on both the apical and basal surfaces of 3D tissue models based on small intestine submucosa (SIS) scaffolds, while also enabling tissue culture under dynamic flow conditions. The flow of cell culture medium inside of the bioreactor was simulated in order to obtain uniform mechanical stimulation by means of shear stress on the apical surface of the tissue model (Surface uniformity index > 0.9). The shear stress magnitude lies in physiologically relevant ranges (Average shear stress ~ 1.4 mPa). As a proof-of-concept experiment a 3D tissue model consisting of spheroids from FaDu cells growing on the surface of the acellular scaffold SISser was cultured under dynamic flow conditions during six days and characterized using different optical imaging techniques. Attachment and growth of the spheroids over the scaffold was monitored using brightfield microscopy, after six days the area covered by the spheroids increased 3.45 times. Additionally, label-free metabolic imaging was performed by using nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) as reporter. Fluorescence lifetime imaging microscopy (FLIM) of the coenzymes revealed differences in both the fluorescence intensity and lifetime of the spheroids and the scaffold. The bioreactor should serve as a tool for future research by providing an environment that supports tissue growth, while enabling the integration of non-invasive, non-destructive functional imaging techniques.