01.06 A Large Animal Model of Liver Injury Induced by Retrorsine and Partial Hepatectomy

K. Inomata1, H. Yagi1, K. Tajima1, H. Shimoda1, T. Hibi1, Y. Abe1, M. Kitago1, M. Shinoda1, O. Itano1, Y. Kitagawa1 1Keio Universiry,Surgery,Shinanomachi, Shinjukuku, TOKYO, Japan

Introduction: Recently, regenerative medicine approaches have been made great advantage, therefore evaluation of its feasibility in animal model is absolutely necessary for clinical application. However, pre-clinical model for the study of liver regeneration is yet to be established. Regeneration process of the liver requires continuous regenerative signals induced by hepatic injury, on the other hand, transplanted cells require functional advantage compared to the remnant cells to repopulate efficiently in recipient’s body. One of the sufficient models for these demands is clearly retrorsine with partial heaptectomy (PH) model. However, no study has been reported using large animal including porcine. Here we demonstrate the first established porcine model with retrorsine and PH.

Methods: Göttingen miniature pigs were treated with 2 intra-peritoneal administrations of retrorsine in different doses; 30mg, 50mg and 100mg/kg respectively, or saline as a control, 2 weeks apart. Four weeks after the second administration of retrorsine, animals underwent PH in different volumes; 50%, 60% and 85%. The animals were sacrificed on 10, 14 and 28 days after the PH and the resected liver volume and histological alterations were evaluated. Blood samples were obtained every 7 days until and every 3 days after the PH to analyze hepatobilially enzymes and synthesized proteins. The quantity of serum and liver tissue retrorsine concentration was determined by LC-MS/MS. Hematoxylin and eosin staining and immunohistochemical staining for Ki-67 and EpCAM were performed on the liver tissue.

Results: The Animals injected 100 mg/kg retrorsine or performed 85% PH were resulted in dead with severe liver injury. Blood test demonstrate that distinct liver disorder was sustained after PH. Liver regeneration was inhibited in animals as 20% volume reduction at day 10 of 60% PH animals with pre-surgical administration of 50mg/kg retrorsine. The histological study demonstrated the explosive puff and cytoplasmic vacuoles were remained in retrorsine treated animals at day 10. The immunohistochemical staining showed the expression of Ki-67 in hepatocytes and EpCAM in biliary epithelium were suppressed in retrorsine treated animals. LC-MS/MS determined the quantity of accumulated retrorsine in the liver tissue while retrorsine was not detected in the blood.

Conclusion: We could successfully demonstrate the first large animal model of retrorsine and 60% PH characterized by sustained liver injury with suppression of hepatic regeneration. We believe that it is a promising model for improving preclinical study of liver regenerative medicine.