Studies on antiviral glycosides. II. Mode of action for virucidal effects on Sendai virus.

Treatment of Sendai virus with p-(sec-butyl)-phenyl-6-chloro-6-deoxy-beta-D-glucopyranoside, followed by freezing and thawing resulted in a loss of hemolytic and cell fusion activities as well as infectivity without affecting hemagglutinating and neuraminidase activities. The anti-hemolytic activity of this compound was reversed by the addition of phosphatidyl choline to the virus samples. p-Azidophenyl-6-chloro-6-deoxy-beta-D[3H]glucopyranoside was successfully used for photoaffinity labeling of a specific virion site, and we confirmed the affected site of the glucoside to be the lipid components in the viral envelopes.     

Lipid of Streptomyces sioyaensis

Three ninhydrin-positive lipids of Streptomyces sioyaensis were found. These lipids were called substance A, B and C, tentatively. Study on the distribution of these lipids in Actinomycetales has shown that substance A was common in all of the strains tested, and that substance B was found in the limited strains. The substance C was characteristic only in Streptomyces sioyaensis.     

Combined Treatment with Exendin-4 and Metformin Attenuates Prostate Cancer Growth

Introduction

Recently, the pleiotropic benefits of incretin-based therapy have been reported. We have previously reported that Exendin–4, a glucagon-like peptide–1 (GLP–1) receptor agonist, attenuates prostate cancer growth. Metformin is known for its anti-cancer effect. Here, we examined the anti-cancer effect of Exendin–4 and metformin using a prostate cancer model.

Methods

Prostate cancer cells were treated with Exendin–4 and/or metformin. Cell proliferation was quantified by growth curves and 5-bromo–2′-deoxyuridine (BrdU) assay. TUNEL assay and AMP-activated protein kinase (AMPK) phosphorylation were examined in LNCaP cells. For in vivo experiments, LNCaP cells were transplanted subcutaneously into the flank region of athymic mice, which were then treated with Exendin–4 and/or metformin. TUNEL assay and immunohistochemistry were performed on tumors.

Results

Exendin–4 and metformin additively decreased the growth curve, but not the migration, of prostate cancer cells. The BrdU assay revealed that both Exendin–4 and metformin significantly decreased prostate cancer cell proliferation. Furthermore, metformin, but not Exendin–4, activated AMPK and induced apoptosis in LNCaP cells. The anti-proliferative effect of metformin was abolished by inhibition or knock down of AMPK. In vivo, Exendin–4 and metformin significantly decreased tumor size, and further significant tumor size reduction was observed after combined treatment. Immunohistochemistry on tumors revealed that the P504S and Ki67 expression decreased by Exendin–4 and/or metformin, and that metformin increased phospho-AMPK expression and the apoptotic cell number.

Conclusion

These data suggest that Exendin–4 and metformin attenuated prostate cancer growth by inhibiting proliferation, and that metformin inhibited proliferation by inducing apoptosis. Combined treatment with Exendin–4 and metformin attenuated prostate cancer growth more than separate treatments.     

A clique-based method for the edit distance between unordered trees and its application to analysis of glycan structures

Background

Measuring similarities between tree structured data is important for analysis of RNA secondary structures, phylogenetic trees, glycan structures, and vascular trees. The edit distance is one of the most widely used measures for comparison of tree structured data. However, it is known that computation of the edit distance for rooted unordered trees is NP-hard. Furthermore, there is almost no available software tool that can compute the exact edit distance for unordered trees.

Results

In this paper, we present a practical method for computing the edit distance between rooted unordered trees. In this method, the edit distance problem for unordered trees is transformed into the maximum clique problem and then efficient solvers for the maximum clique problem are applied. We applied the proposed method to similar structure search for glycan structures. The result suggests that our proposed method can efficiently compute the edit distance for moderate size unordered trees. It also suggests that the proposed method has the accuracy comparative to those by the edit distance for ordered trees and by an existing method for glycan search.

Conclusions

The proposed method is simple but useful for computation of the edit distance between unordered trees. The object code is available upon request.

     

Heterozygous mutations of FREM1 are associated with an increased risk of isolated metopic craniosynostosis in humans and mice

The premature fusion of the paired frontal bones results in metopic craniosynostosis (MC) and gives rise to the clinical phenotype of trigonocephaly. Deletions of chromosome 9p22.3 are well described as a cause of MC with variably penetrant midface hypoplasia. In order to identify the gene responsible for the trigonocephaly component of the 9p22.3 syndrome, a cohort of 109 patients were assessed by high-resolution arrays and MLPA for copy number variations (CNVs) involving 9p22. Five CNVs involving FREM1, all of which were de novo variants, were identified by array-based analyses. The remaining 104 patients with MC were then subjected to targeted FREM1 gene re-sequencing, which identified 3 further mutant alleles, one of which was de novo. Consistent with a pathogenic role, mouse Frem1 mRNA and protein expression was demonstrated in the metopic suture as well as in the pericranium and dura mater. Micro-computed tomography based analyses of the mouse posterior frontal (PF) suture, the human metopic suture equivalent, revealed advanced fusion in all mice homozygous for either of two different Frem1 mutant alleles, while heterozygotes exhibited variably penetrant PF suture anomalies. Gene dosage-related penetrance of midfacial hypoplasia was also evident in the Frem1 mutants. These data suggest that CNVs and mutations involving FREM1 can be identified in a significant percentage of people with MC with or without midface hypoplasia. Furthermore, we present Frem1 mutant mice as the first bona fide mouse model of human metopic craniosynostosis and a new model for midfacial hypoplasia.     

Thrombin induces MCP-1 expression through Rho-kinase and subsequent p38MAPK/NF-κB signaling pathway activation in vascular endothelial cells

Thrombin has been shown to increase expression of chemokines such as monocyte chemoattractant protein 1 (MCP-1) in endothelial cells, leading to the development of atherosclerosis. However, the precise mechanism of this induction remains unknown. In the present study, we investigated whether the small G protein RhoA, and its effector, Rho-kinase are involved in MCP-1 induction by thrombin in endothelial cells. Y-27632, a specific Rho-kinase inhibitor, potently inhibited MCP-1 induction by thrombin. Y-27632 significantly decreased the chemotactic activity of thrombin-stimulated supernatants of endothelial cells on monocytes. Importantly, fasudil, a specific Rho-kinase inhibitor, attenuated MCP-1 gene expression in the aorta of db/db mice. Y-27632 attenuated thrombin-mediated phosphorylation of p38MAPK and p65, indicating that Rho-kinase mediates thrombin-induced MCP-1 expression through p38MAPK and NF-κB activation. Our findings demonstrate that the Rho/Rho-kinase signaling pathway plays a critical role in thrombin-mediated MCP-1 expression and function, and suggest that Rho/Rho-kinase may be an important target in the development of new therapeutic strategies for atherosclerosis.     

Urine temperature as an index for the core temperature of industrial workers in hot or cold environments

Workers working in hot or cold environments are at risk for heat stroke and hypothermia. In Japan, 1718 people including 47 workers died of heat stroke in 2010 (Ministry of Health Labour and Welfare, Japan 2011). While the American Conference of Governmental Industrial Hygienists (ACGIH) recommendation lists the abnormal core temperature of workers as a criterion for halting work, no method has been established for reliably measuring core temperatures at workplaces. ISO 9886 (Ergonomics-evaluation of thermal strain by physiological measurements. ISO copyright office, Geneva, pp 3–14; 2004) recognizes urine temperature as an index of core temperature only at normal temperature. In this study we ascertained whether or not urine temperature could serve as an index for core temperature at temperatures above and below the ISO range. We measured urine temperature of 31 subjects (29.8 ± 11.9 years) using a thermocouple sensor placed in the toilet bowl at ambient temperature settings of 40, 20, and 5˚C, and compared them with rectal temperature. At all ambient temperature settings, urine temperature correlated closely with rectal temperature exhibiting small mean bias. Urine temperature changed in a synchronized manner with rectal temperature at 40˚C. A Bland and Altman analysis showed that the limits of agreement (mean bias ± 2SD) between rectal and urine temperatures were −0.39 to +0.15˚C at 40˚C (95%CI −0.44 to +0.20˚C) and −0.79 to +0.29˚C at 5˚C (−0.89 to +0.39˚C). Hence, urine temperature as measured by the present method is a practical surrogate index for rectal temperature and represents a highly reliable biological monitoring index for assessing hot and cold stresses of workers at actual workplaces.     

Polydom/SVEP1 is a ligand for integrin α9β1

A variety of proteins, including tenascin-C and osteopontin, have been identified as ligands for integrin α9β1. However, their affinities for integrin α9β1 are apparently much lower than those of other integrins (e.g. α3β1, α5β1, and α8β1) for their specific ligands, leaving the possibility that physiological ligands for integrin α9β1 still remain unidentified. In this study, we found that polydom (also named SVEP1) mediates cell adhesion in an integrin α9β1-dependent manner and binds directly to recombinant integrin α9β1 with an affinity that far exceeds those of the known ligands. Using a series of recombinant polydom proteins with N-terminal deletions, we mapped the integrin-binding site to the 21st complement control protein domain. Alanine-scanning mutagenesis revealed that the EDDMMEVPY sequence (amino acids 2636-2644) in the 21st complement control protein domain was involved in the binding to integrin α9β1 and that Glu(2641) was the critical acidic residue for the integrin binding. The importance of this sequence was further confirmed by integrin binding inhibition assays using synthetic peptides. Immunohistochemical analyses of mouse embryonic tissues showed that polydom colocalized with integrin α9 in the stomach, intestine, and other organs. Furthermore, in situ integrin α9β1 binding assays using frozen mouse tissues showed that polydom accounts for most, but not all, of the integrin α9β1 ligands in tissues. Taken together, the present findings indicate that polydom is a hitherto unknown ligand for integrin α9β1 that functions as a physiological ligand in vivo.