Toshihiro Amanuma

Engineering functional two- and three-dimensional liver systems in vivo using hepatic tissue sheets

Hepatic tissue engineering using primary hepatocytes has been considered a valuable new therapeutic modality for several classes of liver diseases. Recent progress in the development of clinically feasible liver tissue engineering approaches, however, has been hampered mainly by insufficient cell-to-cell contact of the engrafted hepatocytes. We developed a method to engineer a uniformly continuous sheet of hepatic tissue using isolated primary hepatocytes cultured on temperature-responsive surfaces. Sheets of hepatic tissue transplanted into the subcutaneous space resulted in efficient engraftment to the surrounding cells, with the formation of two-dimensional hepatic tissues that stably persisted for longer than 200 d. The engineered hepatic tissues also showed several characteristics of liver-specific functionality. Additionally, when the hepatic tissue sheets were layered in vivo, three-dimensional miniature liver systems having persistent survivability could be also engineered. This technology for liver tissue engineering is simple, minimally invasive and free of potentially immunogenic biodegradable scaffolds.

Effects of 3‐Aminobenzamide on Induction of Multiorgan Carcinogenesis by N‐Nitrosobis(2‐hydroxypropyl)amine in Hamsters

The effects of an inhibitor of poly(ADP‐ribose)polymerase, 3‐aminobenzamide (ABA), on N‐nitrosobis(2‐hydroxypropyl)amine (BHP)‐induced pancreas, liver, gallbladder and lung carcinogenesis in Syrian golden hamsters were investigated. Animals were given either BHP alone, by subcutaneous injection at a dose of 500 mg/kg body weight, or in combination with an intraperitoneal injection of ABA 30 min after the BHP at a dose of 300 or 600 mgAg body weight once a week for 5 weeks, and then killed 35 weeks after the commencement of the experiment. ABA exerted inhibitory effects on pancreas and lung carcinogenesis induced by BHP, with mean numbers of lesions (including hyperplasias and carcinomas) being significantly decreased compared with the BHP‐alone group values, while no significant effect was observed on liver or gallbladder carcinogenesis. These results suggest that the effects of ABA on carcinogenesis depend on the target organ as well as the chemical carcinogen examined.

Effects of 3-aminobenzamide on the post-initiation phase of N-nitrosobis(2-oxopropyl)amine induced pancreatic carcinogenesis in Syrian hamsters

Effects of 3-aminobenzamide (ABA) on pancreatic carcinogenesis after initiation by N-nitrosobis(2-oxopropyl)amine (BOP) were investigated in Syrian hamsters. Animals were given BOP at a dose of 70 mg/kg body weight by subcutaneous injection and following a 2-week recovery period, were administered basal diet or basal diet containing 0.5, 0.75 and 1.5% ABA for 30 weeks. While the incidences of resultant pancreatic lesions, including hyperplasia, atypical hyperplasia and carcinoma, induced by BOP were not significantly influenced by ABA treatment, the mean numbers of those pancreatic lesions were significantly decreased in a dose-dependent way. The results therefore suggested the possible involvement of poly(ADP-ribosyl)ation in the post-initiation phase of pancreatic carcinogenesis in hamsters.

Adult hepatocytes direct liver organogenesis through non-parenchymal cell recruitment in the kidney

BACKGROUND & AIMSnSince the first account of the myth of Prometheus, the amazing regenerative capacity of the liver has fascinated researchers because of its enormous medical potential. Liver regeneration is promoted by multiple types of liver cells, including hepatocytes and liver non-parenchymal cells (NPCs), through complex intercellular signaling. However, the mechanism of liver organogenesis, especially the role of adult hepatocytes at ectopic sites, remains unknown. In this study, we demonstrate that hepatocytes alone spurred liver organogenesis to form an organ-sized complex 3D liver that exhibited native liver architecture and functions in the kidneys of mice.nnnMETHODSnIsolated hepatocytes were transplanted under the kidney capsule of monocrotaline (MCT) and partial hepatectomy (PHx)-treated mice. To determine the origin of NPCs in neo-livers, hepatocytes were transplanted into MCT/PHx-treated green fluorescent protein transgenic mice or wild-type mice transplanted with bone marrow cells isolated from green fluorescent protein-mice.nnnRESULTSnHepatocytes engrafted at the subrenal space of mice underwent continuous growth in response to a chronic hepatic injury in the native liver. More than 1.5u202fyears later, whole organ-sized liver tissues with greater mass than those of the injured native liver had formed. Most remarkably, we revealed that at least three types of NPCs with similar phenotypic features to the liver NPCs were recruited from the host tissues including bone marrow. The neo-livers in the kidney exhibited liver-specific functions and architectures, including sinusoidal vascular systems, zonal heterogeneity, and emergence of bile duct cells. Furthermore, the neo-livers successfully rescued the mice with lethal liver injury.nnnCONCLUSIONnOur data clearly show that adult hepatocytes play a leading role as organizer cells in liver organogenesis at ectopic sites via NPC recruitment.nnnLAY SUMMARYnThe role of adult hepatocytes at ectopic locations has not been clarified. In this study, we demonstrated that engrafted hepatocytes in the kidney proliferated, recruited non-parenchymal cells from host tissues including bone marrow, and finally created an organ-sized, complex liver system that exhibited liver-specific architectures and functions. Our results revealed previously undescribed functions of hepatocytes to direct liver organogenesis through non-parenchymal cell recruitment and organize multiple cell types into a complex 3D liver at ectopic sites. Transcript profiling: Microarray data are deposited in GEO (GEO accession: GSE99141).

Lack of modifying effects of 6-mercaptopurine in a medium term bioassay system for liver carcinogenesis using male F344 rats

Carcinogenic and modification potential of 6-mercaptopurine (6-MP) was studied in a medium-term bioassay system for rat liver carcinogenesis. F344 male rats were initiated with a single dose (200 mg/kg body wt.) of diethylnitrosamine (DEN) i.p. and fed diets containing either 0.005% or 0.02% 6-MP with or without 0.05% phenobarbital (PB) for 6 weeks. Quantitative data revealed that 6-MP did not enhance the appearance of enzyme-altered preneoplastic foci and nodules even when administered at the highest dose (0.02%) despite showing an immunosuppressive effect and slight liver cell damage. Neither of the doses of 6-MP exerted any significant influence on the enhancing effect of PB when administered simultaneously in the medium-term-bioassay.