Experimental Models of Acute and Chronic Liver Failure in Nude Mice to Study Hepatocyte Transplantation

Abstract

Although hepatocyte transplantation is a promising therapy for acute liver failure in human, there is still a lack of animal models suffering from hepatic injury in which the benefits of hepatocyte transplantation could be evaluated solely, without the bias caused by immunosuppression. As a consequence, the aim of the study was first to develop reproducible models of partial hepatectomy and of thioacetamide (TA)-or Jo2-induced acute liver failure in nude mice. Chronic liver disease was also investigated by repeated injections of sublethal doses of thioacetamide. Survival rates, routine histologic observations, alanin aminotransferase sera content, Ki67, and caspase 3 immunodetection were investigated both after 40% partial hepatectomy and after toxic-induced damages. Liver injuries were more severe and/or precocious in nude mice than in Balb/ c mice for a given treatment with a maximum of acute injury obtained 24 h after single toxic injection, and were found to be transitory and reversible within 10 days. Toxics induced apoptosis followed by necrosis, confirming recent published data. Onset of fibrosis leading to reproducible chronic cirrhosis in nude mice correlated with increasing number of Ki67-positive cells, indicating that high levels of cell proliferation occurred. Chronic cirrhosis progressively reversed to fibrosis when the treatment ceased. Preliminary results demonstrated that engrafted xenogeneic hepatocytes could be detected in the host liver by anti-MHC class I immunohistochemistry. Fractions enriched in 2n or 4n hepatocytes by cell sorting using a flow cytometer were equivalent to the unpurified fraction in terms of engraftment in control nude mice or in nude mice subjected to PH. However, in mice suffering from liver injury 24 h after Jo2 or TA treatment, the engraftment of 2n hepatocytes was about twice that of an unpurified hepatocyte population or of a population enriched in 4n hepatocytes.

The Nude Mouse as Model for Liver Deficiency Study and Treatment and Xenotransplantation

“…Histological features of liver regeneration, Ki67 and caspase 3 expression, proved that both mechanisms of apoptosis and cell proliferation coexist without any necrotic stage during liver regeneration [62]. This confirms literature data [65].…”

“…In our experience [62], and in contrast to the more resistant wild-type Balb/c mice, Nude mice did not survive 70% partial hepatectomy performed according to the technique of Higgins and Anderson [63] and died within 24 h following the surgery. Our finding is not in accordance with the report of Strom et al [64] referring to a high survival rate of Nude mice submitted to two third partial hepatectomy.…”

“…Forty percent partial hepatectomy was successful and consisted, after median laparotomy, in the resection of the left lateral liver lobe of Nude mice, after ligation using 3 to 5 titanium 3-mm clips for stapling (LIGACLIPrm, Ethicon, France). Hemostasis was performed by dabbing the bloody cut surface with caution using a hemostatic cellulose gauze (SURGICELrm, Johnson & Johnson, France), with removal of this gauze, to decrease postoperative mortality [62]. We recommend the surgery not to exceed 5 min.…”

Protection against Acute Hepatocellular Injury Afforded by Liver Fibrosis Is Independent of T Lymphocytes

“…This is of prime importance in order to achieve adequate experimental observations in immunodeficient animals with liver fibrosis, for example in hepatocellular carcinoma models. Indeed, high mortality rate was consistently observed in a dimethylnitrosamine (DMN) model using NOD-SCID mice and also in a Foxn1 nu/nu mice model using IP injections of TAA [27, 33]. In the present study, using step-wise increased dosage of TAA and a more continuous delivery through drinking water, toxicity of TAA was reduced without undermining the extent of its fibrogenic impact.…”

“…Foxn1 nu/nu mice (nude mice) are athymic and therefore lack mature T-cells [26]. Immunodeficient mice are frequently used in hepatocellular carcinoma (HCC) xenograft models but evaluating the impact of fibrosis in these models is difficult since currently used methods to induce fibrosis (IP injections of TAA at 200 μg/g bw) lead to high mortality rates in these animals [27]. Therefore the aims of this study were 1) to develop a method for the induction of fibrosis in athymic mice without significant mortality; 2) asses and characterize the process of liver fibrosis induced in these mice compared to immunocompetent models of fibrosis and 3) evaluate the role, if any, of T lymphocytes on the protection afforded by liver fibrosis against acute hepatocellular injury [25].…”