Cell transplantation after oxidative hepatic preconditioning with radiation and ischemia–reperfusion leads to extensive liver repopulation

Abstract

The inability of transplanted cells to proliferate in the normal liver hampers cell therapy. We considered that oxidative hepatic DNA damage would impair the survival of native cells and promote proliferation in transplanted cells. Dipeptidyl peptidase-deficient F344 rats were preconditioned with whole liver radiation and warm ischemia-reperfusion followed by intrasplenic transplantation of syngeneic F344 rat hepatocytes. The preconditioning was well tolerated, although serum aminotransferase levels rose transiently and hepatic injury was observed histologically, along with decreased catalase activity and 8-hydroxy adducts of guanine, indicating oxidative DNA damage. Transplanted cells did not proliferate in the liver over 3 months in control animals and animals preconditioned with ischemia-reperfusion alone. Animals treated with radiation alone showed some transplanted cell proliferation. In contrast, the liver of animals preconditioned with radiation plus ischemia-reperfusion was replaced virtually completely over 3 months. Transplanted cells integrated in the liver parenchyma and liver architecture were preserved normally. These findings offer a paradigm for repopulating the liver with transplanted cells. Progressive loss of cells experiencing oxidative DNA damage after radiation and ischemia-reperfusion injury could be of significance for epithelial renewal in additional organs.oxidative damage ͉ hepatocyte L iver repopulation with transplanted cells is of considerable interest for cell and gene therapy (1). Transplanted hepatocytes integrate in the liver parenchyma, function normally, and survive life-long (2-4). However, transplanted cells do not proliferate in the normal adult liver, whereas specific therapies require a significant transplanted cell mass. Proliferation in transplanted cells depends on whether native cells are at survival͞proliferation disadvantages, as suggested by animal studies using exogenous toxins or natural disease, e.g., fumaryl acetoacetate hydroxylase (FAH) mice (hereditary tyrosinemia type-1), Long-Evans Cinnamon (LEC) rats (Wilson's disease), P-glycoprotein-2 (Pgy-2) mutant mice (progressive familial intrahepatic cholestasis), etc. (5-12). Initial clinical studies in familial hypercholesterolemia (FH) or Crigler-Najjar syndrome substantiated these principles (13,14).Genotoxic liver injury is a potent stimulus for transplanted cell proliferation. Rats exposed to retrorsine, a pyrrolizidine alkaloid, or whole liver radiation (RT), which produce DNA adducts and oxidative injury, respectively, lead to extensive transplanted cell proliferation in conjunction with two-thirds partial hepatectomy (PH) (15, 16). Although PH induces hepatic DNA synthesis, its additional effects include oxidative DNA damage, senescence-type changes, including p21 expression, polyploidy, attenuated proliferation capacity, and hepatocyte apoptosis (17-19). Both retrorsine and RT increase PH-induced hepatic polyploidy and apoptosis (20,21). Moreover, the thyroid hormone T3, which regulates PH-induced polyplo...

Stem cells in liver regeneration, fibrosis and cancer: the good, the bad and the ugly

“…Many strategies have been employed to improve the proliferation of transplanted cells within the recipient liver. Preconditioning with irradiation and ischaemia reperfusion injury (IRI) markedly improved the repopulation of dipeptidyl peptidase IV (DPPIV)-negative livers by syngeneic F344 wild-type hepatocytes [34]; likewise, transient IRI performed on LDL receptor-deficient Watanabe rabbits improved the therapeutic benefit (ie lowering of LDL cholesterol) of multiple intraportal infusions of allogeneic hepatocytes [35]. In the retrorsine pretreated and partially hepatectomized DPPIV-deficient rat, superior repopulation by wild-type hepatocytes was achieved by selecting donor cells with low rather than high asialoglycoprotein receptor expression [36].…”

Liver gene therapy: advances and hurdles

“…85,89,90 In combination with a strong regenerative pressure (eg twothirds partial hepatectomy, thyroid hormone administration, ischemia-reperfusion injury or FasL gene transfer), a nearly complete repopulation of host liver by donor hepatocytes was obtained after transplantation of 0.5% of the recipient liver mass. 89,[91][92][93] Similarly, lentivirally transduced hepatocytes were massively expanded in retrorsine/partial hepatectomy-treated rats. 94 Rather than blocking hepatocyte proliferative ability, host hepatocytes were selectively depleted by Fas ligandinduced apoptosis for expanding fas-resistant donor hepatocytes.…”

Ischemic Preconditioning Affects Long-Term Cell Fate through DNA Damage–Related Molecular Signaling and Altered Proliferation

“…For DPPIV and g-glutamyltranspeptidase (GGT), sections were fixed in chloroforme acetone and were stained as described previously. 14,15 For heme oxygenase 1 (HMOX1), sections were stained with anti-rat HMOX1 antibody (Dr. Irving Listowsky, Albert Einstein College of Medicine). For apoptosis, terminal deoxynucleotidyl transferase biotin-dUTP nick end-labeling (TUNEL) was used (ApopTag peroxidase in situ apoptosis kit; EMD Millipore, Billerica, MA).…”

“…14,15,18,19 For liver repopulation, DPPIV À rats were preconditioned with 30 mg/kg retrorsine at 6 and 8 weeks of age, and two-thirds PH was performed 4 weeks later. 22 At 1 week after PH, 5 Â 10 6 hepatocytes were injected into spleen in 0.25 mL serum-free RPMI 1640 medium.…”

“…To examine these possibilities, we used rats deficient in dipeptidyl peptidase IV (DPPIV À ) as a model for studies of liver injury induced by IP, carbon tetrachloride (CCl 4 ), or oxidative stress, along with assays of hepatocyte engraftment and proliferation, 14,15,18,19 followed by dissection of relevant cell signaling pathways.…”