DETERMINATION OF GUSTATORY SUCROSE THRESHOLD IN WATER BY USE OF A LINEAR SUCROSE GRADIENT

A novel experimental method was developed which allows the determination of the threshold concentration of sucrose by use of a linear sucrose gradient in water. With this method a continuous tasting of the test-liquid is possible. A panel of 15 persons experienced in taste-testing was used. Three gradients of different steepness were applied: 0 to 1.5% (w/w) sucrose in 2 min (I), 3 min (II) and 4 min (III). The results of the new method were compared with those of the standard method (DIN). With gradients I and II we found values which were significantly higher than those of the standard method (I: 0.49% (w/w); II: 0.46% (w/w); DIN: 0.31% (w/w)), whereas with gradient III the same threshold value was found as with the DIN-Method (III: 0.32% (w/w)).     

MOLECULAR HETEROCHRONIES AND HETEROTOPIES IN EARLY ECHINOID DEVELOPMENT

By comparing the spatial and temporal distribution of three proteins during early development in seven echinoid species, we demonstrate that both heterochronies and heterotopies in gene-product expression have accompanied the radiation of post-Paleozoic echinoids. All three proteins examined showed significant alterations in time of expression, site of expression, or both. These molecular heterochronies and heterotopies indicate that early development is not necessarily as evolutionarily conservative as morphology of embryos alone would suggest. Evolutionary alterations in early development may be more common than is generally assumed.     

On the ultrastructure of hyalin, a cell adhesion protein of the sea urchin embryo extracellular matrix

Hyalin is a large (ca. 350 x 10(3) kD by gel electrophoresis) molecule that contributes to the hyalin layer surrounding the sea urchin embryo. In previous work a mAb (McA Tg-HYL), specific for hyalin, was found to inhibit cell-hyalin adhesion and block morphogenesis of whole embryos (Adelson, D. L., and T. D. Humphreys. 1988. Development. 104:391-402). In this report, hyalin ultrastructure was examined via rotary shadowing. Hyalin appeared to be a filamentous molecule approximately 75-nm long with a globular "head" about 12 nm in diameter that tended to form aggregates by associating head to head. Hyalin molecules tended to associate with a distinct high molecular weight globular particle ("core"). In fractions containing the core particle often more than one hyalin molecule were seen to be associated with the core. The core particle maintained a tenacious association with hyalin throughout purification procedures. The site(s) of McA Tg-HYL binding to the hyalin molecule were visualized by decorating purified hyalin with the antibody and then rotary shadowing the complex. In these experiments, McA Tg-HYL attached to the hyalin filament near the head region in a pattern suggesting that more than one antibody binding site exists on the hyalin filament. From the ultrastructural data and from the cell adhesion data presented earlier we conclude that hyalin is a filamentous molecule that binds to other hyalin molecules and contains multiple cell binding sites. Attempts were made to demonstrate the existence of lower molecular weight hyalin precursors. Whilst no such precursors could be identified by immunoprecipitation of in vivo labeled embryo lysates, immunoprecipitation of in vitro translation products suggested such precursors (ca 40 x 10(3) kD) might exist.     

Long-term platinum retention after treatment with cisplatin and oxaliplatin

Background  

The aim of this study was to evaluate long-term platinum retention in patients treated with cisplatin and oxaliplatin.     

A novel combinatorial biocatalytic approach for producing antibacterial compounds effective against Mycobacterium tuberculosis (TB)

Two bacterial hosts expressing cloned aromatic oxygenases were used to catalyze the oxidation and polymerization of indole and related substrates, creating mixtures of indigoid compounds comprised of novel dimers and trimers. Crude extracts and purified compounds were tested for their ability to inhibit the growth of Gram-positive organisms, in general, and Mycobacterium tuberculosis (TB), in particular. Of the 74 compounds tested against M. tuberculosis, ~66 % had minimum inhibitory concentrations (MIC) of 5 μg/ml or less. The most effective antibiotic found was designated SAB-P1, a heterodimer of indole and anthranil, which had a MIC of 0.16 μg/ml, and did not inhibit kidney cells (IC⁵⁰) at concentrations of >8 μg/ml. Combinatorial biocatalysis was used to create a series of halogenated derivatives of SAB-P1 with a wider therapeutic window. None of the derivatives had MIC values that were superior to SAB-P1, but some had a wider therapeutic window because of decreased kidney cell toxicity. Generally, the indigoid dimers that were effective against TB appeared to be specific for TB. Some of the trimers generated, however, had a broader spectrum of activity inhibiting not only TB (MIC?=?1.1 μg/ml) but also the growth of Mycobacterium smegmatis MC2 155, Bacillus cereus, Enterococcus faecalis, Staphylococcus epidermidis, Bacillus subtilis 168, and Clostridium acetobutylicum. The structure of two of the novel dimers (SAB-C4 and SAB-P1) and a trimer (SAB-R1) were solved using X-ray crystallography.     

Delimitation of the maritime boundary in the gulf of Maine area

The compulsory dispute settlement regime included in the 1982 Law of the Sea Convention is recognized as one of the most comprehensive in a modern international convention. Yet, in the recent application of this regime, the question has arisen as to whether the procedural prerequisites associated with the LOS Convention's compulsory dispute settlement mechanism are so arduous as to avoid binding and compulsory jurisdiction in most instances. This article addresses that question by examining, in particular, the reasoning of the Southern Bluefin Tuna arbitration tribunal, which found Article 281 of Section 1 of the LOS Convention to bar jurisdiction to the compulsory dispute settlement mechanism prescribed by the Convention, and offers suggestions as to how states might distinguish or overcome the barriers imposed by the Southern Bluefin Tuna tribunal in future cases.     

LvNotch signaling mediates secondary mesenchyme specification in the sea urchin embryo

Cell-cell interactions are thought to regulate the differential specification of secondary mesenchyme cells (SMCs) and endoderm in the sea urchin embryo. The molecular bases of these interactions, however, are unknown. We have previously shown that the sea urchin homologue of the LIN-12/Notch receptor, LvNotch, displays dynamic patterns of expression within both the presumptive SMCs and endoderm during the blastula stage, the time at which these two cell types are thought to be differentially specified (Sherwood, D. R. and McClay, D. R. (1997) Development 124, 3363-3374). The LIN-12/Notch signaling pathway has been shown to mediate the segregation of numerous cell types in both invertebrate and vertebrate embryos. To directly examine whether LvNotch signaling has a role in the differential specification of SMCs and endoderm, we have overexpressed activated and dominant negative forms of LvNotch during early sea urchin development. We show that activation of LvNotch signaling increases SMC specification, while loss or reduction of LvNotch signaling eliminates or significantly decreases SMC specification. Furthermore, results from a mosaic analysis of LvNotch function as well as endogenous LvNotch expression strongly suggest that LvNotch signaling acts autonomously within the presumptive SMCs to mediate SMC specification. Finally, we demonstrate that the expansion of SMCs seen with activation of LvNotch signaling comes at the expense of presumptive endoderm cells, while loss of SMC specification results in the endoderm expanding into territory where SMCs usually arise. Taken together, these results offer compelling evidence that LvNotch signaling directly specifies the SMC fate, and that this signaling is critical for the differential specification of SMCs and endoderm in the sea urchin embryo.     

High-throughput single-nucleotide polymorphism genotyping by fluorescent competitive allele-specific polymerase chain reaction (SNiPTag)

Single nucleotide polymorphisms (SNPs) are becoming widely recognized as the new currency for gene mapping as increasing numbers are discovered. Here we outline a method for their rapid analysis based on an allele-specific polymerase chain reaction (PCR) which employs a competitive approach, whereby both allele-specific primers are present in the same reaction and carry different fluorescent labels. This procedure is simple and amenable to high-throughput genotyping using conventional automated sequencing equipment, and no post-PCR modifications are required. Verification of the procedure was carried out by comparison of results derived by this method with those from restriction enzyme digestion of the ALDH2 exon 12 functional polymorphism (Glu-487-Lys) in 109 individuals. Additionally, we have examined all combinations of nucleotide substitutions and shown them to be differentiated by this method. As proof of concept, several assays were combined and loaded on a single gel lane/capillary to substantially improve throughput. This was made possible by designing the PCR products to be of different lengths and no interference was observed between products differing in size by only six nucleotides. We outline a number of test assays for well-characterized SNPs in human candidate genes for behavioral disorders.