THE PREPARATION OF FRUCTOSE-MODIFIED CHITOSAN MICROCARRIERS AND CULTURE OF PRIMARY RAT HEPATOCYTES

The fructose modified chitosan microcarries (CMs) were prepared by the reaction of glutaraldehyde with fructose-modified chitosan. Various factors that influence the preparation were studied and the reaction conditions were optimized. Morphology of rat hepatocytes cultured on CMs was observed using phase contrast microscope and scanning electron microscope, and the metabolic activities were measured. Rat hepatocytes cultured on CMs retained the spherical shape as they have in vivo and had high metabolic activities. Fructose can enhance the metabolic activity of hepatocytes and the modified CMs are promising scaffold for hepatocytes attachment.

Restoration of cell polarity and bile excretion function of hepatocytes in sandwich-culture

Objective： To investigate the nature of the restoration of cell polarity and bile excretion function in Sandwich-cultured hepatocytes. Methods ： Freshly isolated hepatocytes from male Sprague-Dawley rats were cultured in a double layer collagen gel Sandwich configuration. Morphological changes were observed under a inverted microscope. The domain specific membrane associated protein DPP IV was tested by immunofluorescence, and the bile excretion function was determined by using fluorescein diacetate. Hepatocytes cultured on a single layer of collagen gel were taken as control. Results.. Adult rat hepatocytes cultured in a double layer collagen gel sandwich configuration regained its morphological and functional polarity and maintained polygonal morphology for at least 4 weeks. Immunofluorescence studies using antibodies against DPP IV showed polarity restoration as early as 48 h. After cultured in the double layer collagen gel Sandwich configuration for 96 h the hepatocytes began to excrete bile; while hepatocytes cultured on a single layer collagen gel had no bile excretion. Conclusion.. Hepatocytes cultured in a double layer collagen gel Sandwich configuration are able to regain their morphological and functional polarity given certain conditions. Hepaotcyte culture is a useful tool for the study of polarity restoration.

LIPOPOLYSACCHARIDE－INDUCED CYTOTOXICITY AGAINST CULTURED MOUSE HEPATOCYTES AND THE ROLE OF NONPARENCHYMAL LIVER CELLS

Lipopolysaccharide (LPS) was found to induce significant hepatocytotoxicity against cultured mouse hepatocytes. Degeneration and necrosis of cultured hepatocytes and decrease of hepatocyte viability were prominent. The aspartate transferase level and 3H-TdR release in the medium were significantly increased after treatment, and the degree of these changes paralleled with LPS concentration. Various other parameters showed no significant difference between the hepatocytes cultured alone and those cocultured with nonparenchymal liver cells. However, if the nonparenchymal liver cells were isolated from mice which had been pretreated with D-galactosamine (GalN) not only was the hepatocyotoxicity induced by LPS enhanced, but the cells also showed certain cytotoxicity against cultured hepatocytes even without LPS, These results suggest that nonparenchymal liver cells might promote LPL-induced hepatocyte injury.

APPLICATION OF CHITOSAN-BASED BIOMATERIALS IN BIOARTIFICIAL LIVE

Bioart,ficial liver support system （BALS） has the potential to provide temporary support for patients with fulminant hepatic failure and consist of viable hepatocytes and scaffolding materials for hepatocytes attachment. Various scaffolding materials are used in BALS, including chitosan, which is easily obtained by deacetylation of chitin and widely applied in biomedical applications. In this paper, we introduce and discuses chitosan-based biomaterials for BALS application.

Fucoidan protects hepatocytes from apoptosis and inhibits invasion of hepatocellular carcinoma by up-regulating p42/44 MAPK-dependent NDRG-1/CAP43

Fucoidan is a traditional Chinese medicine suggested to possess anti-tumor effects.In this study the anti-metastatic effects of fucoidan were investigated in vitro in human hepatocellular carcinoma(HCC) cells(Huh-7 and SNU-761) under normoxic and hypoxic conditions and in vivo using a distant liver metastasis model involving injection of MH134 cells into spleen via the portal vein.Its ability to protect hepatocytes against bile acid(BA)-induced apoptosis was investigated in primary hepatocytes.Fucoidan was found to suppress the invasion of HCC cells through up-regulation of p42/44 MAPKdependent NDRG-1/CAP43 and partly,under normoxic conditions,through up-regulation of p42/44MAPK-dependent VMP-1 expression.It also significantly decreased liver metastasis in vivo.As regards its hepatoprotective effect,fucoidan decreased BA-induced hepatocyte apoptosis as shown by the attenuation of caspase-8,and-7 cleavages and suppression of the mobilization of caspase-8 and Fas associated death domain(FADD) into the death-inducing signaling complex.In summary,fucoidandisplays inhibitory effects on proliferation of HCC cells and protective effects on hepatocytes.The results suggest fucoidan is a potent suppressor of tumor invasion with hepatoprotective effects.

Isolation and cultivation of porcine hepatocytes for extracorporeal artificial liver support system

Objective To develop procedures for the successful harvesting of large quantities of viable and functional pig liver cells from abattoir organs.Methods The procedure included partial liver lobe retrograde perfusion and mechanical/enzymatic digestion of the liver tissue, followed by separation of the hepatocytes, based on size and density, from contaminating cell types.Results Digestion of the partial liver lobe resulted in an average yield of 1.39×109 cells (9.9×106 cells/g liver) with an average viability of 92.5%. The yield and viability of cells were improved by dispase/collagenase resultant digestion. The emergence of blebby cells was blocked by supplying oxygen to the cell isolation buffers. Isolated hepatocytes seeded onto polystyrene surfaces remained viable and functional at a level comparable to that of rat hepatocytes, although their function decreased over time.Conclusions Adult pig hepatocytes can be harvested with high yields and retain viability and differentiated function using this method. Abattoir pig livers can be an excellent source of hepatocytes for use as the biological component of artificial liver assist devices.