Hepatocyte transplantation for the low-density lipoprotein receptor-deficient state. A study in the Watanabe rabbit

Abstract

The Watanabe heritable hyperlipidemic (WHHL) rabbit reproduces human familial hypercholesterolemia due to a congenital low-density lipoprotein receptor deficiency and is characterized by elevated serum LDL cholesterol levels and early atherosclerosis. We attempted to transplant normal allogeneic hepatocytes into WHHL rabbits without chronic immunosuppression to cure the LDL receptor-deficient state. Livers from normal New Zealand White (NZW) rabbits were digested by intraportal perfusion of collagenase solution. Pure hepatocytes (PH) were obtained by Percoll gradient separation and nonparenchymal (NP) liver cells by pronase digestion. PH and NP were incubated with fluorescein isothiocyanate-monoclonal anti-rabbit class I, anti-class II, and anti-T cell antibodies and subjected to flow cytometry analysis. PH and NP were also used as stimulators in one way mixed lymphocyte-hepatocyte cultures (MLHC), before and after ultraviolet B light (UVB) exposure. Intraportal and intrasplenic injection of allogeneic PH were also performed in homozygous WHHL rabbits. PH were attached to collagen-coated dextran microcarriers (mc-PH) for intraperitoneal injection. Recipient control and transplanted WHHL rabbits received a single dose of cyclosporine subcutaneously (10 mg/kg/s.c.) at the time of transplantation. PH were mainly class I-positive (77.6%) and class II-negative (5.9%), while 31.5% of NP cells were class II-positive. In MLHC, PH did not stimulate proliferation, (stimulation index: 0.97 +/- 0.21), unlike NP (SI: 23.7). This latter response was abrogated by prior exposure of NP to UVB light. Intraportal injection of PH (n = 4) reduced serum LDL cholesterol to 60% of baseline, an effect lasting 2-3 weeks, and dose-dependent. Intraperitoneal mc-PH, 4 x 10(8) (n = 4), reduced serum LDL cholesterol levels to 45% of baseline more than 4 weeks posttransplant (P = 0.04). We conclude that transplantation of normal allogeneic NZW rabbit mc-PH reduces serum LDL cholesterol levels in homozygous WHHL rabbits without chronic immunosuppression. Longitudinal studies will establish if less atherosclerosis develops in mc-PH WHHL recipients than sham controls.