Performance of rat liver microorgnas (LMOs) cold preserved in BG35 solution as the biological component of a new flat-plate model of bioartificial liver (BAL)

PIZARRO D., BERARDI F., SCANDIZZI A., MEDIAVILLA M. G., RODRÍGUEZ J. V. AND MAMPRIN M.

Rosario

Congreso; 49th Annual Meeting of the Society for Cryobiology; 2012

CAIC- Society for Cryobiology

BAL devices constitute an innovative therapeutic approach currently accepted as a bridge to liver transplantation for patients with liver failure. Usually, the patient?s blood or plasma is circulated extra-corporeally through a bioreactor that houses a metabolically active component which performs the functions of the damaged liver. Among these, ammonia detoxification is crucial because the accumulation of this metabolite is highly toxic to the central nervous system. Our first BAL prototype consisted of a hollow fiber-based cartridge with blood flowing through the fiber?s lumen. This device showed an effective ammonia depuration rate using isolated hepatocytes as cellular component. But, since the ?ideal? biological component should contain all liver cell types for a maximum response, we became interested in evaluating LMOs: tissue slices that retain the basic micro architecture of the liver lobe and its physiological characteristics. Therefore, we built up a flat-based BAL model, suitable to use fresh LMOs. Nonetheless, in order for a BAL to become a useful therapeutic option, its biological component should be viable, functional and ready to use when is needed. Accordingly, we reported the development of a novel preservation solution (BG35) that showed the same efficacy as ViaSpan® to protect LMOs against cold preservation injury for 48 hs. So, the aim of the present work was to evaluate the performance of cold preserved rat LMOs in our new BAL model. LMOs were manually cut from livers (338±27 ìm thickness, n=25) and stored in BG35 and ViaSpan® solutions at 0°C. Freshly cut LMOs were used as controls. After 48 hs of preservation, 1.5 g of LMOs were loaded into the BAL and an ammonia overload (1.06 ± 0.12 mM, n=9) was added to the blood before initiating the system perfusion at 9 mL/min (blood volume=35 mL). Samples of blood and extra-fiber fluid were taken after 60 and 120 min of perfusion. We determined LDH release (%) and ammonia detoxification capacity (% of the initial dose detoxified at different times). After 120 min of perfusion, LMOs cold preserved in BG35 and ViaSpan® were able to detoxify 57.4 ± 12.1 % and 51.9 ± 6.0 %, respectively, of the initial ammonia overload (n=3). No significant differences were found with Controls (49.3 ± 8.8%, n=3). All groups showed a good maintenance of their viability: LDH release after 120 min was 10.5 ± 5.4 % and 9.2 ± 3.4% for BG35 and ViaSpan® preserved LMOs, respectively, similar to Controls (6.1 ± 2.2 %, n=3). In conclusion, LMOs cold preserved in both solutions showed a good ammonia detoxification capacity that enables their use as biological component of a BAL device. The in vitro results encourage us to further test the system in an animal model of acute liver failure. Also, these experiments show that the prototype designed constitutes a simple and inexpensive tool to investigate the effects of preservation, cryopreservation, or culture on the LMOs functions.