Background and Aims. Embryonic stem (ES) cells have a pluripotent ability to differentiate into a variety of cell lineages in vitro.We have recently found the emergence of cell clusters that show the cellular uptake of indocyanine green (ICG) in the culture of differentiated ES cells. ICG is clinically used as a test substance to evaluate liver function because it is eliminated exclusively by hepatocytes. The aim of the present study was to investigate the hepatic characteristics of ICG-stained cells.Methods. Embryoid bodies (EBs), formed by a 5-day hanging drop culture of ES cells, were allowed to outgrow in the placed culture. Gene expression of hepatocyte markers was analyzed by reverse transcriptase-polymerase chain reaction, and albumin production was examined immunohistochemically. Morphology and cellular components were investigated by electron microscopy. ICG-stained cells were further transplanted into the portal vein of mice.

Fish protein has been shown to decrease serum cholesterol content by inhibiting absorption of cholesterol and bile acid in laboratory animals, though the mechanism underlying this effect is not yet fully understood. The purpose of this study was to elucidate the mechanism underlying the inhibition of cholesterol and bile acid absorption following fish protein intake. Male Wistar rats were divided into 2 dietary groups of 7 rats each, 1 group receiving a diet consisting of 20% casein and the other receiving a diet consisting of 10% casein and 10% fish protein. Both experimental diets also contained 0.5% cholesterol and 0.1% sodium cholate. After the rats had been on their respective diets for 4 wk, their serum and liver cholesterol contents and fecal cholesterol, bile acid, and nitrogen excretion contents were measured. Fish protein consumption decreased serum and liver cholesterol content and increased fecal cholesterol and bile acid excretion and simultaneously increased fecal nitrogen excretion. In addition, fish protein hydrolyzate prepared by in vitro digestion had lower micellar solubility of cholesterol and higher binding capacity for bile acids compared with casein hydrolyzate. These results suggest that the hypocholesterolemic effect of fish protein is mediated by increased fecal cholesterol and bile acid excretion, which is due to the digestion products of fish protein having reduced micellar solubility of cholesterol and increased bile acid binding capacity.

We have attempted to generate embryonic stem (ES) cell-derived hepatocytes expressing liver-specific functional properties by use of ES cell technology. It was found that ES cells are allowed to differentiate into hepatocytes possessing high metabolic activities when hepatocyte nuclear factor (HNF)-3beta-transfected ES cells are cultured in alpha-MEM medium supplemented with 10% fetal bovine serum (FBS) and fibroblast growth factor (FGF)-2 in the three-dimensional cell culture system at 5% CO2. The differentiated cells induced albumin, triacylglycerol, urea, and glycogen synthesis as well as further expression of metabolic proteins and serum factors as markers of hepatocytic differentiation for at least 4 months. The cells differentiated from HNF-3beta-transfected ES cells also had hepatocyte-like ultrastructural characteristics, including several endoplasmic reticula, mitochondrion, and glycogen. Our findings indicate that generation of hepatocytes maintaining high metabolic functions developed from mouse ES cells will facilitate the study of the basic mechanism for hepatogenesis and will certainly provide new opportunities for tissue transplantation.

Background and Aims. The purpose of the present study was to examine the efficacy of transplantation of mouse embryonic-stem-(ES)-cell-derived tyrosine hydroxylase-positive (TH

Fish consumption is well known to provide health benefits in both experimental animals and human subjects. Numerous studies have demonstrated the beneficial effects of various protein hydrolysates on lipid metabolism. In this context, this study examined the effect of fish protein hydrolysates (FPH) on cholesterol metabolism compared with the effect of casein. FPHs were prepared from Alaska pollock meat using papain as a protease. Male Wistar rats were divided into the following four dietary groups of seven rats each: either casein (20%) or FPH (10%) + casein (10%), with or without 0.5% cholesterol and 0.1% sodium cholate. Serum and liver lipid levels, fecal cholesterol and bile acid excretions, and the hepatic expression of genes encoding proteins involved in cholesterol homeostasis were examined. In rats fed the FPH diets compared with casein diets with or without cholesterol and sodium cholate, the indexes of cholesterol metabolism-namely, serum cholesterol, triglyceride, and low-density lipoprotein-cholesterol levels-were significantly lower, whereas fecal cholesterol and bile acid excretions were higher. Rats fed the FPH diets compared with casein with cholesterol exhibited a lower liver cholesterol level via an increased liver cholesterol 7α-hydroxylase (CYP7A1) expression level. This study demonstrates that the intake of FPH has hypocholesterolemic effects through the enhancement of fecal cholesterol and bile acid excretions and CYP7A1 expression levels. Therefore, fish peptides prepared by papain digestion might provide health benefits by decreasing the cholesterol content in the blood, which would contribute to the prevention of circulatory system diseases such as arteriosclerosis.

BackgroundDuring the 2013 outbreak, 4638 infection cases and 32 deaths have been recorded in the southern part of Laos. In recent years, the chikungunya virus (CHIKV) emerged in the part of the country bordering Cambodia. Dengue virus (DENV) and CHIKV are transmitted by common mosquito vectors. Both diseases have similar clinical presentations; therefore, CHIKV infections might go undiagnosed in DENV-endemic areas. Thus, rapid detection and accurate diagnosis are crucial for differentiating between the two viruses (DENV and CHIKV). In this study, we demonstrated that CHIKV and two serotypes of DENV are circulating in Laos. In addition, we encountered patients that had been concurrently infected with multiple DENV serotypes or DENV and CHIKV.MethodsPlasma samples were collected from 40 patients with suspected DENV infections during an outbreak between July and August 2013. The reverse transcription polymerase chain reaction was performed to detect the four DENV serotypes and CHIKV using specific primers. Specifically, the complete envelope gene sequences of the viruses were sequenced and subjected to phylogenetic analysis.ResultsForty acute-phase plasma samples from patients with suspected dengue infections were tested for the presence of DENV viral RNA using molecular methods. Among the 40 samples, 14 samples were positive for DENV, 2 samples were positive for both viruses (DENV-2 and DENV-3), whereas DENV-1 and DENV-4 were not detected during the study period. We also encountered 10 samples that were positive for CHIKV. Of the 10 CHIKV-positive samples, 3 samples were co-infected by DENV-2, and 2 samples were co-infected by DENV-3. Phylogenetic analysis revealed that the 2013 dengue outbreak in Laos involved DENV-2 genotype Asian I and DENV-3 genotype II. Moreover, the Laotian CHIKV strains grouped together with those isolated during outbreaks on the Indian Ocean Islands within the East Central South African genotype.ConclusionsThese findings revealed that two serotypes (DENV-2 and DENV-3) and CHIKV were detected. Furthermore, infection of multiple DENV serotypes and CHIKV was also observed in the 2013 dengue outbreak. This is the first documented evidence of co-infection with CHIKV and one of two DENV serotypes.

Protamine has been widely used as a pharmaceutical product and natural food preservative. However, few studies have been conducted to assess the beneficial function of dietary protamine. This study examined the effects of dietary salmon protamine on serum and liver lipid levels and the expression levels of genes encoding proteins involved in lipid homeostasis in the liver of rats. Groups of male Wistar rats were fed AIN93G diet containing 2% or 5% protamine. After 4 weeks of feeding these diets, markedly decreased serum and liver cholesterol (CHOL) and triacylglycerol levels were noted. Increased activity of liver carnitine palmitoyltransferase-2 and acyl-CoA oxidase, which are key enzymes of fatty acid β-oxidation in the mitochondria and peroxisomes, was found in rats fed on protamine. Furthermore, rats fed protamine showed enhanced fecal excretion of CHOL and bile acid and increased liver mRNA expression levels of ATP-binding cassette (ABC) G5 and ABCG8, which form heterodimers and play a major role in the secretion of CHOL into bile. The decrease in triacylglycerol levels in protamine-fed rats was due to the enhancement of liver β-oxidation. Furthermore, rats fed protamine exhibited decreased CHOL levels through the suppression of CHOL and bile acid absorption and the enhancement of CHOL secretion into bile. These results suggest that dietary protamine has beneficial effects that may aid in the prevention of lifestyle-related diseases such as hyperlipidemia and atherosclerosis.

This study examined the effects of fish protein in combination with fish oil on rat lipid metabolism. Male Wistar rats were divided into four groups and fed an AIN93G-based diet with casein (20%) + soybean oil (7%), casein (10%) + fish protein (10%) + soybean oil (7%), casein (20%) + soybean oil (5%) + fish oil (2%), and casein (10%) + fish protein (10%) + soybean oil (5%) + fish oil (2%) for 4 weeks. The dietary combination of fish protein and fish oil decreased the contents of serum triacylglycerol, serum cholesterol, liver triacylglycerol and liver cholesterol in addition to altering liver lipid fatty acid composition. These effects are partly due to the increase in fecal cholesterol, bile acid excretion, and increased enzyme activities of fatty acid β-oxidation in the liver. These data suggest that combined intake of fish protein and fish oil lead to both hypocholesterolemic and hypotriglyceridemic in serum and the liver, while sole intake of fish protein or fish oil decrease only cholesterol and triglyceride levels, respectively. These results suggest that combined intake of fish protein and fish oil may play beneficial roles in the prevention of lifestyle-related diseases as compared with sole fish protein intake.