Cold Preservation of Human Adult Hepatocytes for Liver Cell Therapy

Abstract

Hepatocyte transplantation is a promising alternative therapy for the treatment of hepatic failure, hepatocellular deficiency, and genetic metabolic disorders. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in sufficient quantity and of the quality required for biotherapy. In this study, first we assessed how cold storage in three clinically safe preservative solutions (UW, HTS-FRS, and IGL-1) affects the viability and in vitro functionality of human hepatocytes. Then we evaluated whether such cold-preserved human hepatocytes could engraft and repopulate damaged livers in a mouse model of liver failure. Human hepatocytes showed comparable viabilities after cold preservation in the three solutions. The ability of fresh and cold-stored hepatocytes to attach to a collagen substratum and to synthesize and secrete albumin, coagulation factor VII, and urea in the medium after 3 days in culture was also equally preserved. Coldstored hepatocytes were then transplanted in the spleen of immunodeficient mice previously infected with adenoviruses containing a thymidine kinase construct and treated with a single dose of ganciclovir to induce liver injury. Engraftment and liver repopulation were monitored over time by measuring the blood level of human albumin and by assessing the expression of specific human hepatic mRNAs and proteins in the recipient livers by RT-PCR and immunohistochemistry, respectively. Our findings show that cold-stored human hepatocytes in IGL-1 and HTS-FRS preservative solutions can survive, engraft, and proliferate in a damaged mouse liver. These results demonstrate the usefulness of human hepatocyte hypothermic preservation for cell transplantation.

Ubiquitin-proteasome system and oxidative stress in liver transplantation

“…Due to its cost-effectiveness and simplicity, cold storage with conventional hypothermia remains the main method for liver conservation. In fact, major advances in the field of organ preservation have included the development of new improved preservation solutions capable of reducing cold-induced cellular damage[ 46 ]. Euro-Collins solution was first developed in the 1970s; while more recently, University of Wisconsin (UW)[ 47 ] Celsior[ 48 ], Histidine-Ketoglutarate (HTK)[ 49 ] and Institute Georges Lopez (IGL-1)[ 50 ] solutions tend to be extensively used for liver transplantation.…”

Effective Hypothermic Storage of Human Pluripotent Stem Cell-Derived Cardiomyocytes Compatible With Global Distribution of Cells for Clinical Applications and Toxicology Testing

“…A variety of preservation solutions, such as ViaSpan (University of Wisconsin solution), Celsior, Custodiol histidine‐tryptophan‐ketoglutarate solution and, more recently, HypoThermosol (HTS), were developed taking these factors into consideration [15] and are commercially available today. These solutions showed to be effective for hypothermic storage of a large diversity of cells, tissues, and organs (including neonatal rat ventricular cardiomyocytes [16], renal cells [17], hepatocytes [18, 19], liver tissue [20], synthetic human epidermis tissue [21], kidney [22], pancreas [23], liver [20], and heart [24, 25]). Importantly, some of these formulations can be directly administered into patients, avoiding additional poststorage culture manipulations/costs and washing steps.…”

Development of novel tools for thein vitroinvestigation of drug-induced liver injury

“…As a potential simple and convenient strategy, hypothermic preserved human hepatocytes offer the opportunity to repeat studies at different times using cells from the same single donor or a combination of multiple donors [30]. Although cold storage has a deleterious effect on the viability of human hepatocytes, studies have shown that the albumin secretion, coagulation factor VII, Phase II activities and ammonia metabolism of the remaining viable cells are comparable to fresh hepatocytes [37,38]. Besides, in cryopreserved hepatocytes, the activities of most drug-metabolizing enzymes, glutathione transferase and various transporters are similar to those in freshly isolated cells [35,36].…”