RESUMEN
BACKGROUND/AIMS: To optimise vaccination strategies for immunotherapy in the liver, we have generated a line of transgenic mice expressing beta-Galactosidase downstream of the alpha-fetoprotein promoter (AFP/betaGal). METHODS: betaGal expression was documented by qRT-PCR, enzyme activity and immunohistochemistry. betaGal-specific CD8+ T-cell activation in mice immunised with various vectors was measured by interferon-gamma ELISpot. RESULTS: Like AFP, betaGal expression was detected in fetal hepatocytes and disappeared around birth. In adult mice, a CD8+ T-cell response to betaGal was observed after immunisation with betaGal adenovirus or plasmid DNA but not with betaGal protein or after retroviral infection. When betaGal was re-expressed in adult hepatocytes, immunisation with betaGal adenovirus triggered T-cell mediated elimination of betaGal-expressing hepatocytes. However, the response was weaker than in AFP/betaGal animals in which betaGal was only present around birth. CONCLUSIONS: In AFP/betaGal mice, betaGal is a fetal liver self-antigen. Interestingly, the basal tolerance to betaGal displayed by these animals is increased during liver re-expression of the self-antigen in adulthood. Adenoviral immunisation allows complete elimination of betaGal-expressing hepatocytes in spite of this increased peripheral tolerance. These results highlight the importance of tolerance against self-antigens and validate the AFP/betaGal mice as a good background to test immunotherapy strategies in hepatocarcinogenesis models.
Asunto(s)
Autoantígenos/uso terapéutico , Inmunoterapia/métodos , Hígado/embriología , beta-Galactosidasa/inmunología , beta-Galactosidasa/uso terapéutico , Adenoviridae/genética , Envejecimiento/metabolismo , Animales , Formación de Anticuerpos , Linfocitos T CD8-positivos/inmunología , ADN/uso terapéutico , Feto/metabolismo , Vectores Genéticos , Hepatocitos/metabolismo , Inmunización , Hígado/patología , Ratones , Ratones Transgénicos , Plásmidos/uso terapéutico , Regiones Promotoras Genéticas , alfa-Fetoproteínas/genética , beta-Galactosidasa/metabolismoRESUMEN
BACKGROUND/AIMS: Mature hepatocytes divide to restore liver mass after injury. However, when hepatocyte division is impaired by retrorsine poisoning, regeneration proceeds from another cell type: the small hepatocyte-like progenitor cells (SHPCs). Our aim was to test whether SHPCs could originate from mature hepatocytes. METHODS: Mature hepatocytes were genetically labeled using retroviral vectors harboring the beta-galactosidase gene. After labeling, retrorsine was administered to rats followed by a partial hepatectomy to trigger regeneration. A liver biopsy was performed one month after surgery and rats were sacrificed one month later. RESULTS: We observed the proliferation of small hepatocytes arranged in clusters in liver biopsies. These cells expressed Ki67 antigen and displayed a high mitotic index. At sacrifice, regeneration was completed and clusters had merged. A significant proportion of clusters also expressed beta-galactosidase demonstrating their origin from labeled mature hepatocytes. Finally, the overall proportion of beta-galactosidase positive cells was identical at the time of hepatectomy as well as in liver biopsy and at sacrifice. CONCLUSIONS: The constant proportion of beta-galactosidase positive cells during the regeneration process demonstrates that mature hepatocytes are randomly recruited to proliferate and compensate parenchyma loss in this model. Furthermore, mature hepatocytes are the source of SHPC after retrorsine injury.