title: Cryopreservation-induced nonattachment of human hepatocytes: role of adhesion molecules.
abstract: Good quality cryopreserved human hepatocytes are becoming an important source for clinical hepatocyte transplantation. However, the process of cryopreservation leads to both structural and functional impairment of hepatocytes. The aim of this study was to investigate the mechanisms of cryopreservation-induced nonattachment in human hepatocytes. Hepatocytes were cryopreserved after isolation from unused donor liver tissue. Cell attachment to collagen-coated plates was measured. A cDNA gene array system for 96 cell adhesion-related molecules was used to determine mRNA expression in fresh and cryopreserved hepatocytes. Two cell adhesion molecule proteins were investigated further: beta1-integrin, a cell-matrix adhesion molecule, and E-cadherin, a cell-cell adhesion molecule. Attachment efficiency was significantly decreased after cryopreservation of human hepatocytes. Twenty-two genes were downregulated after cryopreservation including integrins, cadherins, catenins, and matrix metalloproteinases (MMPs). Beta1-Integrin gene and protein expression were significantly decreased in cultured cryopreserved hepatocytes compared to fresh hepatocytes. There was a significant correlation between loss of beta1-integrin and attachment in cryopreserved cells. Degradation of E-cadherin was increased in cryopreserved hepatocytes. The process of cryopreservation leads to downregulation of cell adhesion molecules at the gene and the cellular level. New cryopreservation protocols are needed to prevent these effects on cell attachment.
authors.affiliation: Institute of Liver Studies, King's College London School of Medicine at King's College Hospital, London, UK.
authors.forename: Claire
authors.initials: C
authors.lastname: Terry
authors.forename: Robin D
authors.initials: RD
authors.lastname: Hughes
authors.forename: Ragai R
authors.initials: RR
authors.lastname: Mitry
authors.forename: Sharon C
authors.initials: SC
authors.lastname: Lehec
authors.forename: Anil
authors.initials: A
authors.lastname: Dhawan
mesh_terms: D002102:Cadaver; D015820:Cadherins / Q000032:analysis / Q000378:metabolism; D002448:Cell Adhesion; D015815:Cell Adhesion Molecules / Q000032:analysis* / Q000378:metabolism; D002470:Cell Survival; D017690:Cell Transplantation / Q000379:methods* / Q000639:trends; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D005109:Extracellular Matrix / Q000378:metabolism; D005615:Freezing; D006084:Graft Rejection / Q000378:metabolism; D022781:Hepatocytes / Q000737:chemistry* / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D019012:Integrin beta1 / Q000032:analysis / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000166:cytology; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods
chemical_list: D015820:Cadherins; D015815:Cell Adhesion Molecules; D003451:Cryoprotective Agents; D019012:Integrin beta1; D007770:L-Lactate Dehydrogenase