Interference of hepatitis C virus replication in cell culture by antisense peptide nucleic acids targeting the X-RNA.

The RNA-dependent RNA polymerase (RdRp) of hepatitis C virus (HCV) is the essential catalytic enzyme for viral genome replication. It initiates minus-strand RNA synthesis from a highly conserved 98-nt sequence, called the X-RNA, at the 3'-end of the plus-strand viral genome. In this study, we evaluated the antiviral effects of peptide nucleic acids (PNAs) targeting the X-RNA. Our in vitro RdRp assay results showed that PNAs targeting the three major stem-loop (SL) domains of X-RNA can inhibit RNA synthesis initiation. Delivery of X-RNA-targeted PNAs by fusing the PNAs to cell-penetrating peptides (CPPs) into HCV-replicating cells effectively suppressed HCV replication. Electrophoretic mobility shift assays revealed that the PNA targeting the SL3 region at the 5'-end of X-RNA dissociated the viral RdRp from the X-RNA. Furthermore, delivery of the SL3-targeted PNA into HCV-infected cells resulted in the suppression of HCV RNA replication without activation of interferon ß expression. Collectively, our results indicate that the HCV X-RNA can be effectively targeted by CPP-fused PNAs to block RNA-protein and/or RNA-RNA interactions essential for viral RNA replication and identify X-RNA SL3 as an RdRp binding site crucial for HCV replication. In addition, the ability to inhibit RNA synthesis initiation by targeting HCV X-RNA using antisense PNAs suggests their promising therapeutic potential against HCV infection.

Glucocorticoid receptor represses the Dex-mediated induction of human androgen response element-linked Luc activity.

A human androgen response element (hARE), identified within intron 8 of the human sterol regulatory element-binding protein cleavage-activating protein, interacts with both glucocorticoid receptor (GR) and androgen receptors (AR). The aim of this study was to test the hypothesis that human GR (hGR) might modulate the expression of a hARE-linked reporter gene by dexamethasone (Dex). The hypothesis was tested by: a) co-transfecting HepG2 cells with a hGR and a luciferase (Luc)-reporter gene for performing in vitro investigations and b) by their co-injection into the tail vein of mice for in vivo investigation. In vitro co-transfected cells and the in vivo co-injected mice were then treated with Dex. Our results have led us to concluded that both transfection and injection of the hGR leads to a repression in the Dex-mediated induction of hARE-linked Luc activity both in vitro and in vivo settings. These findings suggest that this assay system allows screening of drug candidates affecting to a signal transduction pathway of the GR and AR and may help in the future discovery and analysis of novel and selection of GR and AR agonists.

Tau overexpression in transgenic mice induces glycogen synthase kinase 3beta and beta-catenin phosphorylation.

The abnormal phosphorylations of tau, GSK3beta, and beta-catenin have been shown to perform a crucial function in the neuropathology of Alzheimer's disease (AD). The primary objective of the current study was to determine the manner in which overexpressed htau23 interacts and regulates the behavior and phosphorylation characteristics of tau, GSK3beta, and beta-catenin. In order to accomplish this, transgenic mice expressing neuron-specific enolase (NSE)-controlled human wild-type tau (NSE/htau23) were created. Transgenic mice evidenced the following: (i) tendency toward memory impairments at later stages, (ii) dramatic overexpression of the tau transgene, coupled with increased tau phosphorylation and paired helical filaments (PHFs), (iii) high levels of GSK3beta phosphorylation with advanced age, resulting in increases in the phosphorylations of tau and beta-catenin, (iv) an inhibitory effect of lithium on the phosphorylations of tau, GSK3beta, and beta-catenin, but not in the non-transgenic littermate group. Therefore, the overexpression of NSE/htau23 in the brains of transgenic mice induces abnormal phosphorylations of tau, GSK3beta, and beta-catenin, which are ultimately linked to neuronal degeneration in cases of AD. These transgenic mice are expected to prove useful for the development of new drugs for the treatment of AD.

Application of inulin-type fructans in animal feed and pet food.

The inulin-type fructans are non-digestible oligosaccharides that are fermented in the gastrointestinal tract of farm animals and pets. This review focuses on the various effects of inulin-type fructans in pigs, poultry, calves and companion animals. Effects of the inulin-type fructans on gut microflora, digestion and availability of nutrients, gut morphology, fermentation characteristics and animal performance are discussed. Inulin-type fructans can support animal performance and health by affecting nutrient digestion, gut microflora and gut morphology, although results vary depending on composition of the basal diet, inclusion level, type of fructan, adaptation period and experimental hygienic conditions.

Effects of feeding antibiotic-free creep feed supplemented with oligofructose, probiotics or synbiotics to suckling piglets increases the preweaning weight gain and composition of intestinal microbiota.

The objective of this study was to determine whether feeding an antibiotic-free creep feed supplemented with either oligofructose, probiotics or synbiotics to suckling piglets influences growth performance, the gut microflora, gut morphology and hematological traits at weaning. Twenty sows with 10 piglets each were randomly assigned to one of four treatments. The treatments consisted of a control (antibiotic-free) diet, 0.2% oligofructose (OF), 0.3% probiotics or 0.5% synbiotics (mixture of 0.2% OF+0.3% probiotics). Piglets were offered the diet ad libitum from 7 d after birth until one day after weaning (21 d of age). At the day after weaning, blood samples were collected from the jugular vein to determine the immune response. Digesta samples of the ileum and colon were collected to determine the microbial composition. Tissue segments from the duodenum and ileum were collected for morphometric measurements of the small intestine. The average daily weight gain was significantly higher for piglets fed the OF or synbiotics diet compared with the pigs fed the control diet. The hematological traits (the concentration of lymphocytes and neutrophils in whole blood) were not affected by the diet. Piglets fed the OF, probiotics or synbiotics diet had a significantly decreased number of total coliform bacteria in the colon. Feeding OF, probiotics or synbiotics significantly increased the population of bifidobacteria in the ileum compared to the control. In the colon, the probiotics and synbiotics diet significantly increased the number of bifidobacteria compared with the control diet. The results of this experiment showed that supplementation of oligofructose or synbiotics to an antibiotic-free creep feed during the preweaning period affected gut microbial population and performance of piglets.

Thoracoscopic plication of diaphragmatic eventration using endostaplers.

Unilateral diaphragmatic eventration and paralysis require plication in cases of progressive dyspnea on exertion and recurrent respiratory infection. The patient, a 40-year-old woman, who had complained of worsening dyspnea on exertion and elevation of the left diaphragm on chest radiographs for 4 years, underwent plication by thoracoscopy with knifeless endostaplers. Improvements in pulmonary functions and dyspnea on exertion have been maintained for 14 months.

Beta-glucuronidase inhibitory activity and hepatoprotective effect of 18 beta-glycyrrhetinic acid from the rhizomes of Glycyrrhiza uralensis.

An inhibitor of beta-glucuronidase from the rhizomes of Glycyrrhiza uralensis was isolated and its hepatoprotective activity on CCI4-induced hepatotoxicity of rats was investigated. From the water-soluble extract of G. uralensis, glycyrrhizin was isolated as a potent inhibitor of beta-glucuronidase. When glycyrrhizin was orally administered, it had a hepatoprotective activity. However, when glycyrrhizin was intraperitoneally administered, it did not have a hepatoprotective activity. 18 beta-Glycyrrhetinic acid, which is a major metabolite of glycyrrhizin by human intestinal bacteria, was also a potent inhibitor of beta-glucuronidase. When 18 beta-glycyrrhetinic acid was intraperitoneally administered, it also had some hepatoprotective activity. These results suggest that glycyrrhizin may be a natural prodrug for the observed hepatoprotective effect in rats and that serum beta-glucuronidase levels have implications for the liver injury, as reductions of its activity by administration of inhibitors such as G. uralensis or its derived products and silymarin correlate with reductions in biochemical indices of liver injury.

Beta-glucuronidase-inhibitory activity and hepatoprotective effect of Ganoderma lucidum.

To prove the relationship between the fluctuation in serum beta-glucuronidase level and hepatotoxicity, an inhibitor of beta-glucuronidase from G. lucidum was isolated and its hepatoprotective activity was investigated. The ether fraction of G. lucidum, which had potent beta-glucuronidase-inhibitory activity, protected against CCl4-induced liver injury. From this ether fraction, ganoderenic acid A, was isolated as the potent inhibitor of beta-glucuronidase. It had a potent hepatoprotective effect against CCl4-induced liver injury. These results suggest that the beta-glucuronidase seems to be closely related to liver injury, which could be prevented by beta-glucuronidase inhibitors.

The role of intestinal bacteria in the transformation of sodium picosulfate.

Sodium picosulfate, a laxative, was biotransformed to 4,4'-dihydroxydiphenyl-(2 pyridyl)-methane by intestinal flora that produced a novel sulfotransferase (not sulfatase). The biotransformation was activated by adding phenolic compounds such as phenol, acetaminophen and flavonoids. The enzyme activity related to this biotransformation was the highest in the contents of the caecum region of the intestine. The enzyme activity was 3.0 mumole/hr/g wet feces in humans and 0.75 in rats (pH 8.0). The optimal pH was 9.0.