Subnormothermic Preservation Maintains Viability and Function in a Porcine Hepatocyte Culture Model Simulating Bioreactor Transport

Abstract

Bioartificial liver (BAL) systems have been developed to bridge patients with acute liver failure (ALF) to liver transplantation or liver regeneration. Clinical application of BAL systems is dependent on the supportive quality of cells used and direct availability of the whole system. Reliable transport of BAL systems from the laboratory to remote treatment centers is therefore inevitable. Subsequently, preservation conditions play a crucial role during transport of a BAL, with temperature being one of the most determining factors. In this study, we assessed the effect of subnormothermic preservation on freshly isolated porcine hepatocytes cultured in monolayer under oxygenation. Additionally, the effect of the University of Wisconsin (UW) preservation solution was compared with Williams' E (WE) culture medium at 4°C. The control group was cultured for 3 days at 37°C, whereas the transport groups were cultured at 4°C, 15°C, 21°C, or 28°C for 24 h at day 2. All groups were tested each day for cell damage and hepatic functions. Subnormothermic culture (i.e., 15°C to 28°C) for a period of 24 h did not reduce any hepatic function and did not increase cellular damage. In contrast, culture of hepatocytes in WE medium and preservation in UW solution at 4°C significantly reduced hepatic function. In conclusion, freshly isolated porcine hepatocytes can be preserved for 24 h at subnormothermic temperatures as low as 15°C. Future research will focus on the implementation of the AMC-BAL in an oxygenated culture medium perfusion system for transport between the laboratory and the hospital.

Primary throughput screening of protectants for hypothermic preservation of bioartificial liver in gel entrapped hepatocytes

“…As commonly stated, the rewarming phase aggravated hypothermia-induced cell injury [11,36]. This was supported by our observation that cell viability decreased more significantly in rewarming than in hypothermic preservation (Fig.…”

“…These protectants also have many different protective mechanism according to different injuries during hypothermia and rewarming treatment, including antioxidants [5,6], iron chelators [7], MPT inhibitors [10], cell membrane and cytoskeleton stabilizers [8], and some could decrease metabolic activity [9]. Besides, several culture systems of hepatocytes in monolayer [11], sandwich [12], gel entrapment [5] and spheroids culture [13] have been employed for attenuating hypothermiainduced cell damage. Moreover, due to the severe cell damage after preservation at 4 • C, mild hypothermic temperatures (such as 15, 21 and 28 • C) had been tried for hypothermic preservation of hepatocytes that did improve the preservation [11].…”

“…Besides, several culture systems of hepatocytes in monolayer [11], sandwich [12], gel entrapment [5] and spheroids culture [13] have been employed for attenuating hypothermiainduced cell damage. Moreover, due to the severe cell damage after preservation at 4 • C, mild hypothermic temperatures (such as 15, 21 and 28 • C) had been tried for hypothermic preservation of hepatocytes that did improve the preservation [11]. However, chemical compounds could not completely inhibit cell injury by hypothermia/rewarming treatment while the required circulation of medium during preservation at mild temperatures was inconvenient for its storage and transportation [14].…”

Characterization of Placental Mesenchymal Stem Cells Spheroids after Generation, Hypothermic and Subnormothermic Storage

“…In our view, subnormothermic (20 С) and hypothermic (4 С) regimes are promising for the spheroids short-term storage. Their effectiveness is demonstrated on other objects [7,8].…”

New concepts in liver preservation: How the pendulum sways back

“…Decreasing the liver temperature to 28°C resulted in a significant reduction of ischemia/reperfusion injury in pig livers in the setting of in situ warm ischemia 20. Also, the metabolic functions of isolated porcine hepatocytes preserved at 15 or 28°C did not differ from the metabolic functions of those preserved at 37°C and were far better than the metabolic functions of those preserved at 4°C 21, 22. In the article by Vairetti et al,2 subnormothermic perfusion preservation was chosen at 20°C for preservation of steatotic livers in order to maintain mitochondrial function and yet avoid the rigidifying effect of a low temperature on the cell membranes of the hepatocytes.…”