Comparative Phenotype and Immunogenicity of Freshly Isolated and Immortalized Rat Hepatocytes
Abstract:Immortalized hepatocytes are an attractive cell source for hepatocyte transplantation and gene transfer. We compared the phenotype and immunogenicity of freshly isolated (FIH) and immortalized (IMH) rat hepatocytes. Effect of culture and proinflammatory cytokines (TNF-α, IFN-γ) was studied on phenotype. FIH were isolated by collagenase digestion. Two SV40 immortalized hepatocyte cell lines were tested (RH1 and P9). Immunophenotyping was performed by FACS analysis using anti-rat-specific antibodies. Immunogenicity was evaluated by a mixed lymphocyte hepatocyte reaction (MLHR). FIH suspension was an almost homogeneous parenchymal cell population with few (1–2%) CD8+ cells. FIH showed a positive staining for ICAM-1 (20–35%) and for Class I (RT1A, 30–60%) but no staining for Class II (RT1B). After 48 h of culture, the already ICAM-1-positive cells were more strongly stained and additionally 3.6% of the cells (possibly endothelial cells) were Class II positive. IMH showed a consistent expression of Class I (93–97%) and ICAM-1 (95–97%) but no expression of Class II. Culture of IMH for 48 h had no effect on Class II expression but increased ICAM-1 expression. Addition of TNF-α at 1000 UI/ml to cultures of FIH or IMH increased Class I and ICAM-1 expression whereas IFN-γ (50 or 1000 UI/ml) had no evident effect. Hepatocyte immunogenicity, assessed in MLHR and appreciated by the stimulation index (SI) test/SI syngeneic control, was similar for IMH (RH1: 2.68 ± 0.89; P9: 2.37 ± 0.78) and FIH (2.52 ± 0.18). In conclusion, despite some quantitative immunophenotypic differences, FIH and IMH induced the same proliferation rate of allogeneic T lymphocytes. Thus, immortalized hepatocytes may constitute an appropriate cellular model to study the prevention of hepatocyte rejection by gene transfer.
Document Type: Research Article
Affiliations: 1: †Laboratoire de Thérapeutique Immuno-moléculaire, EFS/EA 2284/Inserm E-0119, 1 Bd A Fleming, BP 1937, 25020 Besançon cedex, France 2: *Laboratoire de Chirurgie Expérimentale, Fondation Transplantation, 5 Avenue Molière, 67200 Strasbourg, France 3: ‡University of Strathclyde, Bioengineering Unit, Wolfson Center, 106 Rottenrow, Glasgow 64 ONW, Scotland 4: §Faculté de Médecine et Pharmacie, Laboratoire de Biologie Cellulaire, Place Saint Jacques, 25030 Besançon, France
Publication date: August 1, 2001
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