Massive Repopulation of Rat Liver by Transplantation of Hepatocytes Into Specific Lobes of the Liver and Ligation of Portal Vein Branches to Other Lobes1

Abstract

An important consideration in application of hepatocyte transplantation is whether the number of engrafted hepatocytes is sufficient to achieve the desired effect. Here we have evaluated the proliferative potential of transplanted primary hepatocytes during regeneration of hepatic lobes. Two million hepatocytes isolated from congeneic normal Wistar-RHA rats were injected into the main portal vein of deficient, jaundiced Gunn rats. The right branch of the portal vein was ligated 24 hr before hepatocyte transplantation (group A) or transiently clamped during hepatocyte injection (group B) or 24 hr after hepatocyte injection (group C). In these groups, the three lobes supplied by the right branch of the portal vein rapidly atrophied and disappeared in 4 days, whereas the remaining lobes proliferated, as shown by size increase and 5-bromo-2-deoxy-uridine uptake. Two control groups received 2 million (group D) or 20 million hepatocytes (group E) without ligation. Hepatocyte engraftment occurred in all groups. The greatest hypobilirubinemic effect was observed in group A, in which serum bilirubin concentrations were reduced to 1.7+/-0.45 mg/dl from pretransplantation levels of 6.9+/-1.2 mg/dl. This effect was even greater than that observed after transplantation of 20 times more hepatocytes without ligation (group E). Specific endonuclease digestion of a polymerase chain reaction-amplified segment of the ugt1 gene from hepatic DNA showed that up to 25% of the DNA was of donor origin. This paralleled the hepatic bilirubin-UDP-glucuronosyltransferase activity, which was above 50% of normal. The results indicate that the transplanted hepatocytes proliferate preferentially within the regenerating lobes, replacing more than 20% of the liver mass with the progeny of the transplanted phenotypically normal hepatocytes.

Correction of the UDP-glucuronosyltransferase gene defect in the Gunn rat model of Crigler–Najjar syndrome type I with a chimeric oligonucleotide

“…These cells then were transfected with unlabeled UGT1A1 ONs, which were either complexed to PEI or encapsulated in the anionic liposomes. The frequency of G nt insertion at position 1206 was determined by hybridization of duplicate colony lifts of the PCR-amplified and cloned 379-nt stretch of exon 4 of the rat UGT1A1 gene (28).…”

“…Aliquots of 100 g of total or microsomal proteins were separated by using 7.5% SDS͞PAGE. After electrophoretic transfer onto nitrocellulose membranes, immunoblots were incubated sequentially with H 2 O 2 , 5% milk blocking solution, primary antibody (1:5,000) to rat UGT1A1 (28), and horseradish peroxidaseconjugated goat anti-rabbit IgG secondary antibody. UGT1A1 protein was detected by using the Ultra chemiluminescent system (Pierce).…”

Feasibility of Hepatocyte Transplantation-Based Therapies for Primary Hyperoxalurias

“…Potentially, this could be accomplished by nonsurgical means. We have shown previously that ligation of a branch of a portal vein of the rat liver, keeping the arterial flow unperturbed, leads to apoptosis of cells in the lobe supplied by the ligated vessel [23], resulting in disappearance of that lobe in approximately 1 week, without evidence of necrosis. Radiological equivalents of this method could be developed to ablate the nonrepopulated portions of the liver, at a second step.…”

Salivary gland

“…Most such studies have aimed to restore the function of a specific cell type or cell groups in an organ. The possibility of regenerating an entire organ by cell transplantation has not been demonstrated except for the liver, which possesses enormous regenerative capacity [17]. Recent studies have shown that transplanted bone marrow cells might replace tissue in many more organs than was previously thought possible.…”