www.elsevier.com/locate/jhep

Journal of Hepatology 49 (2008) 6–8

Editorial

A symphony of techniques for liver cell therapy,only applicable to rats? q

Michael Ott

Clinic for Gastroenterology, Hepatology, Endocrinology and Twincore Center for Infectious Diseases,

Hannover Medical School, Feodor-Lynen Str. 7–9, 30625 Hannover, Germany

See Article, pages 99–106

The present issue of the Journal includes a study byKawashita et al., that investigates the potential ofimmortalized hepatocytes to repopulate the liver ofGunn rats after preparative irradiation and partial hep-atectomy [1]. Primary hepatocytes from Gunn rats weretransduced with a thermolabile mutant simian virus 40T-antigen as well as the human UGTA1a1 gene. Thetransduced cells were transplanted into recipient Gunnrats via intrasplenic injection. After various time points,serum bilirubin concentrations, human UGT1A1 levelsand other parameters were measured. Three treatmentgroups and one control group receiving the parental cellline without transduction with the human UGTA1a1gene were followed over a time period of 140 days.The results demonstrate that the in vitro amplified andtransduced hepatocytes engraft, function and proliferatein the recipient liver. Radiation therapy of the recipientliver in combination with partial hepatectomy was mosteffective in reducing serum bilirubin levels.

All single components of this study are well knownand have been published before. It was one of the manyachievements of Jayanta and Namita Roy-Chowdhury’slaboratory to have thoroughly characterized the Gunnrat as a model for cell and gene therapy [2,3]. Their lab-oratory was also among the first to create and character-ize immortalized hepatocytes, to study ex vivo genetherapy for metabolic liver diseases and, in co-operation

0168-8278/$34.00 � 2008 European Association for the Study of the Liver.

doi:10.1016/j.jhep.2008.04.009

Associate Editor: M. Traunerq The author declares that he does not have anything to disclose

regarding funding from industries or conflict of interest with respect tothis manuscript.

E-mail address: OTT-MHH@gmx.de

with other groups at the Albert Einstein College, toapply preparative hepatic irradiation in combinationwith hepatectomy to induce liver repopulation by trans-planted cells. The present paper now orchestrates all thepieces into one therapeutic approach and shows that abilirubin glucuronidation deficiency state can be cor-rected in rats.

The authors provide evidence that immortalizedhepatocytes can be expanded in culture and trans-planted into a recipient liver. This is both, a surprisingand a promising result, since primary adult hepatocytestrypsinized from collagen coated cell culture plateshardly engraft in a recipient liver after transplantation.Of course, favourable conditions were created for thecells. Preparative irradiation of the liver combined withpartial hepatectomy has been shown before to provide afavourable environment for cell engraftment and repop-ulation and probably was the key for the generation ofsufficient liver mass derived from UGT1A1 gene cor-rected Gunn rat hepatocytes [4,5]. Additional help forreducing the bilirubin levels after transplantation wasprovided by transducing the immortalized hepatocyteswith a retrovirus expressing the UGT1A1 gene. Com-pared to freshly isolated ‘‘wild-type” hepatocytes, thetransduced immortalized Gunn rat hepatocytes werepreviously shown to over-express the UGT1A1 geneby twofold [6]. In contrast, endogenously produced glu-curonidation activity is likely to be down-regulated witha gradual loss of the differentiated hepatic phenotype inimmortalized and expanded ‘‘wild-type” hepatocytes. Itwould be interesting to know whether immortalized‘‘wild-type” hepatocytes would be as efficient for thecorrection of hyperbilirubinemia as isolated primary

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M. Ott / Journal of Hepatology 49 (2008) 6–8 7

adult hepatocytes or gene corrected immortalized Gunnrat-derived hepatocytes [7].

Will this strategy lead us to new horizons in the celltherapy of metabolic liver diseases? Yes and no. Oneof the major limitations for the development of liver celltherapy programs is the shortage of human hepatocytes.Many laboratories have focussed on stem cells as arenewable source of hepatocytes, but overall, extensiveliver tissue formation has not been achieved with eitheradult or embryonic stem cells. It is one of the main mes-sages of this report that immortalized hepatocytes canengraft, proliferate and correct a disease phenotype ofa recipient liver. Safety of the cells remains a concern.Activation of the SV40 T-antigen is known to triggermolecular events in the target cell, which are not fullyreversible by inactivation of the gene. Although tumourshave not been detected in the animals, activation ofalternative pathways inducing proliferation and newtechnologies such as flanking transferred genes withloxP targets for Cre recombinase mediated excisionmay be needed for future clinical application [8].

It is obvious from the published data that hepatocytetransplantation provides a therapeutic benefit in patientswith hereditary metabolic liver disease, but cannot com-pletely and stably correct a disease phenotype [9,10].Most patients, which are considered as targets for celltherapy do not have signs of a liver pathology and thuswould not provide an intrinsic growth advantage fortransplanted cells. Although well established in surgicaldepartments, partial hepatectomy of the recipient liveras part of a cell therapy protocol will probably not beaccepted for the often very young patients with meta-bolic liver disease. Mild transient ischemia/reperfusiondamage of the liver may provide an alternative methodin order to increase engraftment and proliferation oftransplanted hepatocytes. In a conclusive study Malhiand co-workers have demonstrated that �90 min ofischemia and reperfusion in combination with prepara-tive hepatic irradiation can result in massive repopula-tion by transplanted cells [11]. The inherent risks ofthe procedure may be significantly reduced by restrictionto single liver lobes. Cell transplantation combined withregional ischemia/reperfusion of less than half of theliver mass partially corrected hypercholesterolemia evenin the absence of radiation theraphy [12]. The radiationdosage required for inhibition of hepatocyte prolifera-tion would not necessarily impair liver function, butwould definitely cause DNA damage and carry the riskfor tumour formation. Stephenne et al. have recentlyreviewed the application of radiation therapy in childrenwith liver organ transplantation and did not find anincreased risk for tumour formation or other patholo-gies in the follow-up [13].

Would any of these procedures be too risky forpatients with hereditary metabolic liver disease? Lessonsmay be learned from the hematologists. For years bone

marrow transplantation has been a common practice forthe treatment of congenital immune disorders and otherdefects affecting bone marrow-derived cells. Completeor partial myeloablation with chemotherapeutic drugsis often part of the protocol to facilitate the engraftmentof donor bone marrow stem cells [14].

Advocacy for more aggressive cell therapy protocolswould have to consider that liver organ transplantationis available for most of the children in the western worldand provides long-term correction of the metabolicdefect. Nevertheless, cell therapy concepts as an alterna-tive or an additional approach to organ transplantationmay offer potential benefits for selected patients withmetabolic liver disease. It should be the objective ofthe international scientific community to develop thebest protocols and techniques for the benefit of patients.The fascinating experiments presented by Kawashitaet al. should help us to move closer to the routine appli-cation of cell therapies for liver diseases.

References

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[2] Chowdhury JR, Kondapalli R, Chowdhury NR. Gunn rat: amodel for inherited deficiency of bilirubin glucuronidation. AdvVet Sci Comp Med 1993;37:149–173.

[3] Kim BH, Takahashi M, Tada K, Bosma PJ, Roy-Chowdhury J,Roy-Chowdhury N. Cell and gene therapy for inherited deficiencyof bilirubin glucuronidation. Proc Natl Acad Sci USA1999;96:10349–10354.

[4] Takahashi M, Deb NJ, Kawashita Y, Lee SW, Furgueil J,Okuyama T, et al. A novel strategy for in vivo expansion oftransplanted hepatocytes using preparative hepatic irradiationand FasL-induced hepatocellular apoptosis. Gene Ther2003;10:304–313.

[5] Christiansen H, Koenig S, Krause P, Hermann RM, Rave-FrankM, Proehl T, et al. External-beam radiotherapy as preparativeregimen for hepatocyte transplantation after partial hepatectomy.Int J Radiat Oncol Biol Phys 2006;65:509–516.

[6] Tada K, Roy-Chowdhury N, Prasad V, Kim BH, Manchikala-pudi P, Fox IJ, et al. Long-term amelioration of bilirubinglucuronidation defect in Gunn rats by transplanting geneticallymodified immortalized autologous hepatocytes. Cell Transplant1998;7:607–616.

[7] Guha C, Parashar B, Deb NJ, Garg M, Gorla GR, Singh A, et al.Normal hepatocytes correct serum bilirubin after repopulation ofGunn rat liver subjected to irradiation/partial resection. Hepatol-ogy 2002;36:354–362.

[8] Kobayashi N, Fujiwara T, Westerman KA, Inoue Y, Sakaguchi M,Noguchi H, et al. Prevention of acute liver failure in rats withreversibly immortalized human hepatocytes. Science 2000;287:1258–1262.

[9] Nussler A, Konig S, Ott M, Sokal E, Christ B, Thasler W, et al.Present status and perspectives of cell-based therapies for liverdiseases. J Hepatol 2006;45:144–159.

[10] Fisher RA, Strom SC. Human hepatocyte transplantation:worldwide results. Transplantation 2006;82:441–449.

[11] Malhi H, Gorla GR, Irani AN, Annamaneni P, Gupta S. Celltransplantation after oxidative hepatic preconditioning with

8 M. Ott / Journal of Hepatology 49 (2008) 6–8

radiation and ischemia-reperfusion leads to extensive liver repop-ulation. Proc Natl Acad Sci USA 2002;99:13114–13119.

[12] Attaran M, Schneider A, Grote C, Zwiens C, Flemming P, GratzKF, et al. Regional and transient ischemia/reperfusion injury inthe liver improves therapeutic efficacy of allogeneic intraportalhepatocyte transplantation in low-density lipoprotein receptordeficient Watanabe rabbits. J Hepatol 2004;41:837–844.

[13] Stephenne X, Najimi M, Janssen M, Reding R, de Ville de GoyetJ, Sokal EM. Liver allograft radiotherapy to treat rejection inchildren: efficacy in orthotopic liver transplantation and long-termsafety. Liver Int 2005;25:1108–1113.

[14] Notarangelo LD, Forino C, Mazzolari E. Stem cell transplanta-tion in primary immunodeficiencies. Curr Opin Allergy ClinImmunol 2006;6:443–448.