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Therapeutic efficacy of human hepatocyte transplantation in a SCID/uPA mouse model with inducible liver disease.

Douglas DN, Kawahara T, Sis B, Bond D, Fischer KP, Tyrrell DL, Lewis JT, Kneteman NM - PLoS ONE (2010)

Bottom Line: In vitro experiments demonstrated efficient killing of vTK expressing hepatoma cells after GCV treatment.Surprisingly, vTK/GCV-dependent apoptosis and mitochondrial aberrations were also localized to bystander vTK-negative HH.Functional support by engrafted HH may be secured by strategies aimed at limiting this bystander effect.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, University of Alberta, Edmonton, Alberta, Canada. donnad@ualberta.ca

ABSTRACT

Background: Severe Combined Immune Deficient (SCID)/Urokinase-type Plasminogen Activator (uPA) mice undergo liver failure and are useful hosts for the propagation of transplanted human hepatocytes (HH) which must compete with recipient-derived hepatocytes for replacement of the diseased liver parenchyma. While partial replacement by HH has proven useful for studies with Hepatitis C virus, complete replacement of SCID/uPA mouse liver by HH has never been achieved and limits the broader application of these mice for other areas of biomedical research. The herpes simplex virus type-1 thymidine kinase (HSVtk)/ganciclovir (GCV) system is a powerful tool for cell-specific ablation in transgenic animals. The aim of this study was to selectively eliminate murine-derived parenchymal liver cells from humanized SCID/uPA mouse liver in order to achieve mice with completely humanized liver parenchyma. Thus, we reproduced the HSVtk (vTK)/GCV system of hepatic failure in SCID/uPA mice.

Methodology/principal findings: In vitro experiments demonstrated efficient killing of vTK expressing hepatoma cells after GCV treatment. For in vivo experiments, expression of vTK was targeted to the livers of FVB/N and SCID/uPA mice. Hepatic sensitivity to GCV was first established in FVB/N mice since these mice do not undergo liver failure inherent to SCID/uPA mice. Hepatic vTK expression was found to be an integral component of GCV-induced pathologic and biochemical alterations and caused death due to liver dysfunction in vTK transgenic FVB/N and non-transplanted SCID/uPA mice. In SCID/uPA mice with humanized liver, vTK/GCV caused death despite extensive replacement of the mouse liver parenchyma with HH (ranging from 32-87%). Surprisingly, vTK/GCV-dependent apoptosis and mitochondrial aberrations were also localized to bystander vTK-negative HH.

Conclusions/significance: Extensive replacement of mouse liver parenchyma by HH does not provide a secure therapeutic advantage against vTK/GCV-induced cytotoxicity targeted to residual mouse hepatocytes. Functional support by engrafted HH may be secured by strategies aimed at limiting this bystander effect.

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Impact of vTK/GCV on non transplanted SCID/uPA mouse liver.Gross Appearance (upper panels) and H&E staining (lower panels) of livers from non-transplanted and age-matched vTK+SCID/uPA mice (3 months old) treated with 0, 25, 50 and 100 mg/kg GCV (i.p. every 48 h for 10 days). RN, regenerative nodules; PL, pale liver.
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pone-0009209-g005: Impact of vTK/GCV on non transplanted SCID/uPA mouse liver.Gross Appearance (upper panels) and H&E staining (lower panels) of livers from non-transplanted and age-matched vTK+SCID/uPA mice (3 months old) treated with 0, 25, 50 and 100 mg/kg GCV (i.p. every 48 h for 10 days). RN, regenerative nodules; PL, pale liver.

Mentions: Like (vTK−)SCID/uPA mouse liver, the majority of the liver parenchyma in vTK+SCID/uPA mice was occupied by intensely red colored RN surrounded by pale liver parenchyma (Fig. 5, upper panels). RN appeared to be particularly sensitive to GCV since their redness could be diminished by GCV treatment; the livers of vTK+ mice that received the highest dose of GCV were universally pale and the discrimination between RN and surrounding pale liver parenchyma was less evident. Like their FVB/N counterparts, the livers from vTK+ SCID/uPA mice that received GCV treatment displayed hepatocytes with dffuse cytoplasmic and nuclear enlargemenet with mononuclear inflammatory cell infiltration (Fig. 5, lower panels). Areas of necrosis, acidophilic bodies, necrosis and apoptotic bodies were also featured in these livers but were absent from vTK− SCID/uPA mice and vTK+ SCID/uPA mice that did not receive GCV (not shown). Having confirmed GCV sensitivity in non-transplanted mice, GCV was administered to transplanted vTK+ and vTK-SCID/uPA mice that had established levels of human chimerism (summarized in Table 2). The average pre-dose ALT/AST values for all vTK− and vTK+ chimeric mice were within the range of the non-transplanted vTK-SCID/uPA mice. These were 135 +/− 40 IU/L and 313 +/− 72 IU/L, respectively. The experiment had to be terminated after 14 days of GCV administration due to health crisis seen only in vTK+ chimeric mice (with the exception of mouse v241 which had to be euthanized after 10 days of GCV administration). These mice had significantly elevated aminotransferases relative to experimental vTK− chimeric mice (Fig. 6). Serum creatinine concentrations remained in the normal range for all chimeric experimental mice (not shown).


Therapeutic efficacy of human hepatocyte transplantation in a SCID/uPA mouse model with inducible liver disease.

Douglas DN, Kawahara T, Sis B, Bond D, Fischer KP, Tyrrell DL, Lewis JT, Kneteman NM - PLoS ONE (2010)

Impact of vTK/GCV on non transplanted SCID/uPA mouse liver.Gross Appearance (upper panels) and H&E staining (lower panels) of livers from non-transplanted and age-matched vTK+SCID/uPA mice (3 months old) treated with 0, 25, 50 and 100 mg/kg GCV (i.p. every 48 h for 10 days). RN, regenerative nodules; PL, pale liver.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823785&req=5

pone-0009209-g005: Impact of vTK/GCV on non transplanted SCID/uPA mouse liver.Gross Appearance (upper panels) and H&E staining (lower panels) of livers from non-transplanted and age-matched vTK+SCID/uPA mice (3 months old) treated with 0, 25, 50 and 100 mg/kg GCV (i.p. every 48 h for 10 days). RN, regenerative nodules; PL, pale liver.
Mentions: Like (vTK−)SCID/uPA mouse liver, the majority of the liver parenchyma in vTK+SCID/uPA mice was occupied by intensely red colored RN surrounded by pale liver parenchyma (Fig. 5, upper panels). RN appeared to be particularly sensitive to GCV since their redness could be diminished by GCV treatment; the livers of vTK+ mice that received the highest dose of GCV were universally pale and the discrimination between RN and surrounding pale liver parenchyma was less evident. Like their FVB/N counterparts, the livers from vTK+ SCID/uPA mice that received GCV treatment displayed hepatocytes with dffuse cytoplasmic and nuclear enlargemenet with mononuclear inflammatory cell infiltration (Fig. 5, lower panels). Areas of necrosis, acidophilic bodies, necrosis and apoptotic bodies were also featured in these livers but were absent from vTK− SCID/uPA mice and vTK+ SCID/uPA mice that did not receive GCV (not shown). Having confirmed GCV sensitivity in non-transplanted mice, GCV was administered to transplanted vTK+ and vTK-SCID/uPA mice that had established levels of human chimerism (summarized in Table 2). The average pre-dose ALT/AST values for all vTK− and vTK+ chimeric mice were within the range of the non-transplanted vTK-SCID/uPA mice. These were 135 +/− 40 IU/L and 313 +/− 72 IU/L, respectively. The experiment had to be terminated after 14 days of GCV administration due to health crisis seen only in vTK+ chimeric mice (with the exception of mouse v241 which had to be euthanized after 10 days of GCV administration). These mice had significantly elevated aminotransferases relative to experimental vTK− chimeric mice (Fig. 6). Serum creatinine concentrations remained in the normal range for all chimeric experimental mice (not shown).

Bottom Line: In vitro experiments demonstrated efficient killing of vTK expressing hepatoma cells after GCV treatment.Surprisingly, vTK/GCV-dependent apoptosis and mitochondrial aberrations were also localized to bystander vTK-negative HH.Functional support by engrafted HH may be secured by strategies aimed at limiting this bystander effect.

View Article: PubMed Central - PubMed

Affiliation: Department of Surgery, University of Alberta, Edmonton, Alberta, Canada. donnad@ualberta.ca

ABSTRACT

Background: Severe Combined Immune Deficient (SCID)/Urokinase-type Plasminogen Activator (uPA) mice undergo liver failure and are useful hosts for the propagation of transplanted human hepatocytes (HH) which must compete with recipient-derived hepatocytes for replacement of the diseased liver parenchyma. While partial replacement by HH has proven useful for studies with Hepatitis C virus, complete replacement of SCID/uPA mouse liver by HH has never been achieved and limits the broader application of these mice for other areas of biomedical research. The herpes simplex virus type-1 thymidine kinase (HSVtk)/ganciclovir (GCV) system is a powerful tool for cell-specific ablation in transgenic animals. The aim of this study was to selectively eliminate murine-derived parenchymal liver cells from humanized SCID/uPA mouse liver in order to achieve mice with completely humanized liver parenchyma. Thus, we reproduced the HSVtk (vTK)/GCV system of hepatic failure in SCID/uPA mice.

Methodology/principal findings: In vitro experiments demonstrated efficient killing of vTK expressing hepatoma cells after GCV treatment. For in vivo experiments, expression of vTK was targeted to the livers of FVB/N and SCID/uPA mice. Hepatic sensitivity to GCV was first established in FVB/N mice since these mice do not undergo liver failure inherent to SCID/uPA mice. Hepatic vTK expression was found to be an integral component of GCV-induced pathologic and biochemical alterations and caused death due to liver dysfunction in vTK transgenic FVB/N and non-transplanted SCID/uPA mice. In SCID/uPA mice with humanized liver, vTK/GCV caused death despite extensive replacement of the mouse liver parenchyma with HH (ranging from 32-87%). Surprisingly, vTK/GCV-dependent apoptosis and mitochondrial aberrations were also localized to bystander vTK-negative HH.

Conclusions/significance: Extensive replacement of mouse liver parenchyma by HH does not provide a secure therapeutic advantage against vTK/GCV-induced cytotoxicity targeted to residual mouse hepatocytes. Functional support by engrafted HH may be secured by strategies aimed at limiting this bystander effect.

Show MeSH
Related in: MedlinePlus