Elsevier

Journal of Hepatology

Volume 39, Issue 3, September 2003, Pages 365-373
Journal of Hepatology

Liver regeneration in a retrorsine/CCl4 – induced acute liver failure model: do bone marrow-derived cells contribute?

https://doi.org/10.1016/S0168-8278(03)00264-2Get rights and content

Abstract

Background/Aims: Adult bone marrow contains progenitors capable of generating hepatocytes. Here a new liver failure model is introduced to assess whether bone marrow-derived progeny contribute to liver regeneration after acute hepatotoxic liver failure.

Methods: Retrorsine was used to inhibit endogenous hepatocyte proliferation, before inducing acute liver failure by carbon tetrachloride. Bone marrow chimeras were generated before inducing liver failure to trace bone marrow-derived cells. Therefore, CD45 and major histocompatibility complex (MHC) class I dimorphic rat models were applied.

Results: Early after acute liver failure a multilineage inflammatory infiltrate was observed, mainly consisting of granulocytes. In long-term experiments small numbers of CD90+/CD45 cells of donor origin occurred in clusters associated with portal triads. Bone marrow cell infusion was not able to enhance liver regeneration. Cellular hypertrophy was the predominant way of liver mass regeneration in models applying retrorsine.

Conclusions: Retrorsine pretreatment did not affect sensitivity for carbon tetrachloride. A multilineage inflammatory infiltrate was observed in rats whether pretreated with retrorsine or not. Few donor cells co-expressing CD90 (THY 1) were present in recipient livers, which may resemble donor-derived hematopoietic progenitors or oval cells. No other donor cells within liver parenchyma were detected. This is in contrast to other cell infusion models of acute cell death.

Introduction

Adult stem cell transplantation might be a feasible treatment option for liver dysfunctions. It has recently been shown that bone marrow derived adult stem cells are capable of transdifferentiation into muscle cells [11], [12], [20], hepatocytes [1], [4], [18], [23] and even neurons [5], [19], [26], [37]. Theise et al. could show that in long-term observations male recipients of a female liver allograft had a small amount of male hepatocytes within the graft [34], [35]. As normal adult bone marrow contains a small amount of hepatic progenitor cells [3], it remains open whether these cells were transdifferentiated hematopoietic stem cells or hepatic progenitor cells seeding the liver. That transdifferentiation from hematopoietic stem cells to hepatocytes is possible was presented by Lagasse et al. [23]. In their study livers of fumarylacetoacetate hydrolase mutant mice (FAH−/− knock out), were almost completely restored with normally functioning hepatocytes after transplantation with bone marrow-derived hematopoietic stem cells. In this FAH-deficient mouse model, so-called oval cells are the first donor-derived cells to appear. Proliferation of oval cells [24], [25] is typically induced by application of the acetylaminofluorene/partial hepatectomy protocol (AAF/PH) in rodents. In contrast to occurrence of oval cells, Gordon et al. presented the hypothesis that proliferation of a novel type of hepatic progenitor can be observed when retrorsine is administered in the PH protocol instead of AAF [13], [14], [15], [16]. Cells restoring liver mass in the retrorsine/PH protocol are morphologically and immunohistochemically distinct from those in the AAF/PH protocol. Gordon's group therefore suggested to term them small hepatocyte-like progenitor cells (SHPC). Instead of surgically removing parts of the liver by PH, loss of mature hepatocytes in the AAF model can also be achieved by administration of carbon tetrachloride (CCl4).

In the present study a new rat liver failure model is introduced to assess the contribution of bone marrow-derived cells to liver regeneration. It was designed in analogy to other models of acute cell death applying a stem cell infusion at the time of artificial cell death. CD45 and MHC class I dimorphic bone marrow chimeras were used to trace bone marrow-derived cells [8].

The present study had three major goals: (1) to describe the influx of donor hematopoietic cells into the recipient liver; (2) to study whether SHPCs and/or oval cells may derive from donor hematopoietic cells in this model; and (3) to answer the question whether bone marrow-derived stem cells may be relevant to enhance liver regeneration after acute liver failure.

Section snippets

Animals

In the CD45 dimorphic rat model system LEW (RT1lll/RT7a) rats were used as recipients and LEW.7B (RT1lll/RT7b) rats as bone marrow donors [9], [38]. LEW.1A (RT1aaa/RT7a) rats were used as recipients in the MHC class I dimorphic model system, whereas LEW.1WR2 (RT1uaa/RT7a) rats were used as donors. Donor and recipient cells were distinguished by haplotype-specific antibodies (see below).

Rats were bred in the Central Animal Laboratory of Hannover Medical School and kept under standard conditions.

No functional benefit of bone marrow cell infusions following acute hepatotoxic liver failure in the retrorsine/CCl4 model

To assess whether bone marrow-derived cells can contribute to regeneration of liver function in a model of acute liver failure, LEW and LEW.1A rats were treated in accordance with protocol 2, Fig. 1. Six h after the administration of increasing doses of carbon tetrachloride animals received BMC from LEW or LEW.1WR2 donors. In additional study groups, recipient rats were pretreated with retrorsine (Table 1). No significant differences in survival could be observed between groups, which did or

Discussion

In the present study a new rat model of acute hepatic failure is introduced to analyze the contribution of bone marrow-derived cells to the regenerative process of the liver. Recently it was reported that bone marrow bound progenitors can contribute to regeneration when transplanted into damaged tissues [6], [20], [37]. It was the objective of this study to analyze whether adult BMC can significantly contribute to regeneration of liver function. For this purpose, stable hematopoietic chimeras

Acknowledgements

Ms Irina Kucuk and Ms Damaris Leemhuis is thanked for excellent technical assistance. LEW.7B rats and anti-RT1Au monoclonal antibodies were a kind gift from Prof. Dr Kurt Wonigeit, Dept. for Visceral- and Transplantation Surgery, Hannover Medical School. Rats were bred by Ms Britta Trautewig and Ms Lisa Haenisch. Gamma irradiation was carried out in collaboration with Dr Rolf Baumann, Dept. for Radiotherapy, Hannover Medical School. Technical assistance in FHBs lab was provided by Ms Aimee

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