Optimizing Normoxic Conditions in Liver Devices

The authors of this paper experimented with the possibility of improving oxygenation to hepatocytes in a liver replacement device by adjusting the porosity of the extracellular matrix that said hepatocytes were embedded in. The investigators cultured Sprague Dawley rat hepatocytes on ECM gels constituted from collagen gel, with the addition of sterilized hollow polystyrene microspheres in varying degrees with different cellular densities as well as heterogeneous densities . The gels were also constructed in two different forms, a sandwich configuration as well as homogeneous dispersal. Oxygen transport visualization was accomplished with the addition of dichlorotris ruthenium hydrate, as the dye is sensitive to the presence of molecular oxygen. The cultures were then exposed to an atmosphere of 95% oxygen/5% carbon dioxide. Evaluation of cell viability was done through optical examination as well as Flouview and Metamorph Imaging software. Metabolic status was monitored by NADH autofluorescence and ROS generation. Results showed that the highest oxygen transport distances were achieved with the highest concentration of microspheres (40uL/mL) with an accompanying relative higher oxygenation at distances further away from the origin of oxygen. This is regardless of the configuration of the tissue culture. Examination of cell viability also revealed a definable distance limit at which the majority of cells beyond the limit died. The 40uL/mL enhanced ECM resulted in a 200% increase of this boundary in comparison to normal collagen ECM, from 150 um to 420 um or higher. NADH fluorescence analysis again showed that the oxygen concentration as a function of distance from oxygen origin decreased at a much slower rate in the 40uL/mL eECM. ROS generation was measured to decrease just as much as in lower or no enhancement; however, absolute values were measured to be less per cell.

The successful development of a bioartificial liver assistive or replacement device is crucial due to susceptibility of the liver to damage. With its various roles in metabolism, detoxification, circulation, and biochemical regulation, the liver is exposed to almost all harmful substances that can pass through the body. In addition, the spread of viral hepatitis has results in the same magnitude of annual deaths as HIV/AIDS, with a large portion of the infected population unresponsive to current treatments. At the same time, one of the difficulties in keep artificial organ tissue alive is proper oxygenation. Hepatocytes are particularly demanding in terms of oxygenation; the liver as a whole is one of the largest consumers of oxygen in the body. This paper shows that it is possible to modify the hepatic scaffolding in a conceptually simple manner to minimize not only hypoxia but also hyperoxia, which can also cause tissue damage in the form of free radicals. As a result, the lifespan of BLADs can be extended, accompanied by a noticeable increase in hepatocyte metabolic performance. This ECM modification is applicable to improving the engineering of other complex, multi-unit structures such as the kidney.