title: 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. Cold-stored 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.
authors.affiliation: INSERM, U1040, Institut de Recherche en Biothérapie, F-34295 Montpellier, France.
authors.forename: Cedric
authors.initials: C
authors.lastname: Duret
authors.forename: Daniel
authors.initials: D
authors.lastname: Moreno
authors.forename: Anangi
authors.initials: A
authors.lastname: Balasiddaiah
authors.forename: Solene
authors.initials: S
authors.lastname: Roux
authors.forename: Phillipe
authors.initials: P
authors.lastname: Briolotti
authors.forename: Edith
authors.initials: E
authors.lastname: Raulet
authors.forename: Astrid
authors.initials: A
authors.lastname: Herrero
authors.forename: Helene
authors.initials: H
authors.lastname: Ramet
authors.forename: Christine
authors.initials: C
authors.lastname: Biron-Andreani
authors.forename: Sabine
authors.initials: S
authors.lastname: Gerbal-Chaloin
authors.forename: Jeanne
authors.initials: J
authors.lastname: Ramos
authors.forename: Francis
authors.initials: F
authors.lastname: Navarro
authors.forename: Jean
authors.initials: J
authors.lastname: Hardwigsen
authors.forename: Patrick
authors.initials: P
authors.lastname: Maurel
authors.forename: Rafael
authors.initials: R
authors.lastname: Aldabe
authors.forename: Martine
authors.initials: M
authors.lastname: Daujat-Chavanieu
mesh_terms: D000328:Adult; D000368:Aged; D000418:Albumins / Q000096:biosynthesis; D000818:Animals; D049109:Cell Proliferation; D002470:Cell Survival; D064987:Cell- and Tissue-Based Therapy / Q000379:methods*; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology*; D005167:Factor VII / Q000096:biosynthesis; D005260:Female; D015774:Ganciclovir / Q000009:adverse effects; D022781:Hepatocytes / Q000166:cytology / Q000637:transplantation*; D006801:Humans; D008099:Liver / Q000166:cytology; D008107:Liver Diseases / Q000628:therapy*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008875:Middle Aged; D019188:Organ Preservation Solutions / Q000494:pharmacology*; D012709:Serum Albumin / Q000032:analysis; D013154:Spleen / Q000166:cytology; D014183:Transplantation, Heterologous; D014508:Urea / Q000378:metabolism
chemical_list: D000418:Albumins; D003451:Cryoprotective Agents; C504757:IGL-1 solution; D019188:Organ Preservation Solutions; D012709:Serum Albumin; D014508:Urea; D005167:Factor VII; D015774:Ganciclovir