Expression screening of a yeast artificial chromosome contig refines the location of the mouse H3a minor histocompatibility antigen gene

The H3 complex, on mouse Chromosome 2, is an important model locus for understanding mechanisms underlying non-self Ag recognition during tissue transplantation rejection between MHC-matched mouse strains. H3a is a minor histocompatibility Ag gene, located within H3, that encodes a polymorphic peptide alloantigen recognized by cytolytic T cells. Other genes within the complex include beta2-microglobulin and H3b. A yeast artificial chromosome (YAC) contig is described that spans the interval between D2Mit444 and D2Mit17, a region known to contain H3a. This contig refines the position of many genes and anonymous loci. In addition, 23 new sequence-tagged sites are described that further increase the genetic resolution surrounding H3a. A novel assay was developed to determine the location of H3a within the contig. Representative YACs were modified by retrofitting with a mammalian selectable marker, and then introduced by spheroplast fusion into mouse L cells. YAC-containing L cells were screened for the expression of the YAC-encoded H3a(a) Ag by using them as targets in a cell-mediated lympholysis assay with H3a(a)-specific CTLs. A single YAC carrying H3a was identified. Based on the location of this YAC within the contig, many candidate genes can be eliminated. The data position H3a between Tyro3 and Epb4.2, in close proximity to Capn3. These studies illustrate how genetic and genomic information can be exploited toward identifying genes encoding not only histocompatibility Ags, but also any autoantigen recognized by T cells.     

Rapid multiplex analysis for the factor V Leiden and prothrombin G20210A mutations associated with hereditary thrombophilia

Thromboembolic episodes are common events and affect approximately one in 1,000 persons annually. Pulmonary embolism alone accounts for 50,000 to 100,000 deaths per year in the United States with > 50% of those being elderly persons. Resistance to activated protein C is the most common inherited disorder associated with hereditary thrombophilia. A missense mutation has been identified in the gene coding for coagulation factor V (codon 506) which renders this procoagulant factor resistant to inactivation by activated protein C resulting in an increased risk for venous thrombosis. Recently, a second polymorphism was identified in the prothrombin gene (factor II) which is also associated with increased risk for venous thrombosis. Because of the high prevalence of these two mutations in the general population as well as in specific patient populations, the ability readily to detect these two mutations must be feasible. In this study, we evaluated 303 patients for the prothrombin mutatin (G20210A) which were previously tested for the factor V mutation using established polymerase chain reaction-mediated restriction fragment length polymorphism assays. In these patients, 30 (9.9%) were found to be heterozygous for the factor V Leiden mutation with no homozygous mutants identified. Twenty individuals (6.6%) were heterozygous for the prothrombin G20210A mutation, and we identified two individuals (0.66%) who were homozygous for the 20210A allele. Of the total 303 individuals screened, two were double heterozygotes for both the factor V Leiden and the prothrombin gene mutations. We also describe a multiplex polymerase chain reaction-mediated restriction fragment length polymorphism assay for detecting both mutations in a single-tube double-enzyme digestion reaction making identification of these two mutations easily achievable.     

Synthesis and biological activities of phenyl piperazine-based peptidomimetic growth hormone secretagogues

A new class of potent, orally active phenyl piperazine-based GH secretagogues have been discovered from attempts to mimic the arrangement of the phenyl substituent in the spiroindanyl piperidine and spiroindoline sulfonamide privileged structures of 4 and 1, respectively. The best of these compounds, 18 (EC50 = 2.8 nM) is nearly as potent as MK-0677 for releasing GH from rat pituitary cells.     

Prenatal diagnosis of trisomy 13 on fetal cells obtained from maternal blood after minor enrichment

In a pilot study to establish fetal nucleated red blood cell (NRBC) detection in maternal blood, trisomy 13 was diagnosed by FISH analysis at 11 weeks' gestation. The NRBCs were detected after a single-step ficoll density gradient enrichment. In blood samples taken both before and after CVS, 52 and 80 NRBCs, respectively, were found to be positive for fetal haemoglobin. In 47 per cent of these cells, FISH analysis for X and Y chromosomes confirmed the fetal sex. Moreover, 48 per cent of these NRBCs showed three fluorescent signals for a chromosome 13 probe, which confirmed the diagnosis of trisomy 13, previously detected at CVS karyotyping. This is the first report of non-invasive prenatal diagnosis of trisomy 13, i.e., pre-CVS, in the first trimester. The high number of fetal NRBCs detected indicates a connection with aneuploidy, probably due to early impairment of the feto-maternal barrier.     

Insulin-like growth factor binding protein-1 at mouse neuromuscular synapses

The insulin-like growth factor (IGF) signaling system includes the growth factors and their cell surface receptors, along with circulating IGF binding proteins (IGFBPs) that may alter and modulate the action of these neurotrophic hormones. These IGFBPs, along with IGFs and receptors, have been detected in various tissues including the brain. In this study, using polyclonal antibody to human IGFBP-1 or bovine IGFBP-2, we found that mouse muscle extracts contain similar-sized proteins that cross-react with these antibodies on Western immunoblots. After establishing that these antibodies reacted with the homologous murine IGFBPs, we performed immunocytochemistry to demonstrate the localization of IGFBP-1 at the neuromuscular junction, a model nicotinic, cholinergic synapse, as well as within intramuscular nerves. IGFBP-2, a distinct macromolecule, is present on the surface of muscle fibers and is not present within synapses or nerves.     

Antimetabolite-induced increases in the invasive capacity of murine leukaemia L1210 cells

Pretreatment of murine leukaemia L1210 cells with non-lethal concentrations of various antimetabolites increased the in vitro invasive capacity of these cells into monolayers of rat embryo fibroblasts. The increase in invasive capacity was partly correlated with the induced cell cycle arrest. The concomitant increase in cell surface fucosylation and inhibition of invasion with sulphate indicate a role for glycoproteins in this process. Our results suggest that treatment with antimetabolites may lead to a more aggressive phenotype by altering cell surface properties.     

A hybrid online-product recommendation system: Combining implicit rating-based collaborative filtering and sequential pattern analysis

Many online shopping malls in which explicit rating information is not available still have difficulty in providing recommendation services using collaborative filtering (CF) techniques for their users. Applying temporal purchase patterns derived from sequential pattern analysis (SPA) for recommendation services also often makes users unhappy with the inaccurate and biased results obtained by not considering individual preferences. The objective of this research is twofold. One is to derive implicit ratings so that CF can be applied to online transaction data even when no explicit rating information is available, and the other is to integrate CF and SPA for improving recommendation quality. Based on the results of several experiments that we conducted to compare the performance between ours and others, we contend that implicit rating can successfully replace explicit rating in CF and that the hybrid approach of CF and SPA is better than the individual ones.     

A vertical and floor line-based monocular SLAM system for corridor environments

In this paper, we propose a vertical and floor line-based monocular simultaneous localization and mapping (SLAM) system which utilizes vertical lines, floor lines, and vanishing points as sensory input to perform robust SLAM in corridor environments. By combining three map feature types, our design can help a robot to perform accurate pose estimation, repeatable loop closure, and to construct a more expressive environmental map. As a primitive element of a geometric structure, a line segment has one additional dimension compared to a point feature, thereby allowing the use of line segments to easily represent a geometric structure using a smaller number of features. This system presents map features on a 2D ground space: the vertical line as a projection point, the floor line as the original line, and the vanishing point as a directional vector. Although the vertical line, floor line, and vanishing point use different parameterization and initialization methods, their measurement models are integrated into a unified extended Kalman filter (EKF) framework. Experimental results show that our system can be deployed in a structured indoor environment as a suitable SLAM solution.     

Synthesis of silver/polymer colloidal composites from surface-functional porous polymer microspheres

This study presents a different colloidal silver (Ag)/polymer system where Ag nanoparticles are deposited uniformly onto surface-functional porous poly(ethylene glycol dimethacrylate-co-acrylonitrile) (poly(EGDMA-co-AN)) microspheres. The formation and morphology of the composite microspheres were characterized from electron microscopy and X-ray diffraction analyses. The significance of the present report is that owing to the high affinity between Ag and nitrile group (CN) on the large surface of the microspheres, the Ag nanoparticles having a face-centered cubic phase were incorporated evenly into the deep pores of the microspheres with fine size and size distribution. In the preservation test, the Ag/poly(EGDMA-co-AN) composite microspheres obtained showed an excellent anti-bacterial performance, elucidating a high applicability for a new preservative.