Calculation of line integrals in boundary integral methods

We propose quadrature rules for the approximation of line integrals possessing logarithmic singularities and show their convergence. In some instances a superconvergence rate is demonstrated.

Wolbachia infections are distributed throughout insect somatic and germ line tissues

Wolbachia are intracellular microorganisms that form maternally-inherited infections within numerous arthropod species. These bacteria have drawn much attention, due in part to the reproductive alterations that they induce in their hosts including cytoplasmic incompatibility (CI), feminization and parthenogenesis. Although Wolbachia's presence within insect reproductive tissues has been well described, relatively few studies have examined the extent to which Wolbachia infects other tissues. We have examined Wolbachia tissue tropism in a number of representative insect hosts by western blot, dot blot hybridization and diagnostic PCR. Results from these studies indicate that Wolbachia are much more widely distributed in host tissues than previously appreciated. Furthermore, the distribution of Wolbachia in somatic tissues varied between different Wolbachia/host associations. Some associations showed Wolbachia disseminated throughout most tissues while others appeared to be much more restricted, being predominantly limited to the reproductive tissues. We discuss the relevance of these infection patterns to the evolution of Wolbachia/host symbioses and to potential applied uses of Wolbachia.

In vitro cultivation of Wolbachia pipientis in an Aedes albopictus cell line

A continuous cell line, Aa23, was established from eggs of a strain of the Asian tiger mosquito, Aedes albopictus, naturally infected with the intracellular symbiont Wolbachia pipientis. The resulting cell line was shown to be persistently infected with the bacterial endosymbiont. Treatment with antibiotics cured the cells of the infection. In the course of establishing this cell line it was noticed that RFLPs in the PCR products of two Wolbachia genes from the parental mosquitoes were fixed in the infected cell line. This indicates that the mosquito host was naturally superinfected with different Wolbachia strains, whereas the infected cell line derived from these mosquitoes only contained one of the original Wolbachia strains. The development of anin vitroculture system for this fastidious microorganism should facilitate molecular analysis of the reproduction distorting phenotypes it induces in natural arthropod hosts.

The metal directed assembly of a trinuclear macrocyclic copper(II) complex

A trinuclear macrocyclic complex is reported from the metal directed condensation between melamine, formaldehyde and the Cu-II complex of a linear tetraamine.

A model for assembly and activation of the GM-CSF, IL-3 and IL-5 receptors: Insights from activated mutants of the common beta subunit

Granulocyte-macrophage colony stimulating factor (GM-CSF), Interleukin-3 (IL-3) and Interleukin-5 (IL-5) have overlapping, pleiotropic effects on hematopoietic cells, including neutrophils, eosinophils, monocytes and early progenitor cells. The high-affinity receptors for human GM-CSF, IL-3, and IL-5 share a common beta-subunit (h beta(c)), which is essential for signalling and plays a major role in recruiting intracellular signalling molecules. While activation of the cytoplasmic tyrosine kinase JAK2 appears to be the initiating event for signalling, the immediate events that trigger this are still unclear. We have isolated a number of activated mutants of h beta(c), which can be grouped into classes defined by their state of receptor phosphorylation, their requirement for alpha subunit as a cofactor, and their activities in primary cells and cell lines. We discuss these findings with regard to the stoichiometry, activation, and signalling of the normal GM-CSF/IL-3/IL-5 receptor complexes. Specifically, this work has implications for the role of the ligand-specific alpha-subunits in initiating the signalling through the beta-subunit, the role of beta subunit dimerization as a receptor trigger, and the function of receptor tyrosine phosphorylation in generating growth and survival signals. Based on the properties of the activated mutants and the recent structures of erythropoietin receptor (Epo-R) complexes, we propose a model in which (1) activation of h beta(c) can occur via alternative states that differ with respect to stoichiometry and subunit assembly, but which all mediate proliferative responses, and (2) each of the different classes of activated mutants mimics one of these alternative states. (C) 2000 International Society for Experimental Hematology. Published by Elsevier Science Inc.

The line structure of balanced ternary designs with block size three, index three and rho(2) = 1, 2

In this paper necessary and sufficient conditions for a vector to be the fine structure of a balanced ternary design with block size 3, index 3 and rho(2) = 1 and 2 are determined with one unresolved case.

Removal of a cysteine ligand from rubredoxin: assembly of Fe2S2 and Fe(S-Cys)(3)(OH) centres

The electron transfer protein rubredoxin from Clostridium pasteurianum contains an Fe(S-Cys)(4) active site. Mutant proteins C9G, C9A, C42G and C42A, in which cysteine ligands are replaced by non-ligating Gly or Ala residues, have been expressed in Escherichia coli. The C42A protein expresses with a (Fe2S2)-S-III cluster in place. In contrast, the other proteins are isolated in colourless forms, although a (Fe2S2)-S-III cluster may be assembled in the C42G protein via incubation with Fe-III and sulfide. The four mutant proteins were isolated as stable mononuclear Hg-II forms which were converted to unstable mononuclear Fe-III preparations that contain both holo and apo protein. The Fe-III systems were characterized by metal analysis and mass spectrometry and by electronic, electron paramagnetic resonance, X-ray absorption and resonance Raman spectroscopies. The dominant Fe-III form in the C9A preparation is a Fe(S-Cys)(3)(OH) centre, similar to that observed previously in the C6S mutant protein. Related centres are present in the proteins NifU and IscU responsible for assembly and repair of iron-sulfur clusters in both prokaryotic and eukaryotic cells. In addition to Fe(S-Cys)(3)(OH) centres, the C9G, C42G and C42A preparations contain a second four-coordinate Fe-III form in which a ligand appears to be supplied by the protein chain. Electronic supplementary material to this paper can be obtained by using the Springer Link server located at http://dx.doi.org/10.1007/s00775-0020355-1.

Identification of cellular changes associated with increased production of human growth hormone in a recombinant Chinese hamster ovary cell line

A proteomics approach was used to identify the proteins potentially implicated in the cellular response concomitant with elevated production levels of human growth hormone in a recombinant Chinese hamster ovary (CHO) cell line following exposure to 0.5 mM butyrate and 80 muM zinc sulphate in the production media. This involved incorporation of two-dimensional (2-D) gel electrophoresis and protein identification by a combination of N-terminal sequencing, matrix-assisted laser desorption/ionisation-time of flight mass spectrometry, amino acid analysis and cross species database matching. From these identifications a CHO 2-D reference,map and annotated database have been established. Metabolic labelling and subsequent autoradiography showed the induction of a number of cellular proteins in response to the media additives butyrate and zinc sulphate. These were identified as GRP75, enolase and thioredoxin. The chaperone proteins GRP78, HSP90, GRP94 and HSP70 were not up-regulated under these conditions.

The land assembly problem revisited

As in the standard land assembly problem, a developer wants to buy two adjacent blocks of land belonging to two different owners. The value of the two blocks of land to the developer is greater than the sum of the individual values of the blocks for each owner. Unlike the land assembly literature, however, our focus is on the incentive that each lot owner has to delay the start of negotiations, rather than on the public goods nature of the problem. An incentive for delay exists, for example, when owners perceive that being last to sell will allow them to capture a larger share of the joint surplus from the development. We show that competition at point of sale can cause equilibrium delay, and that cooperation at point of sale will eliminate delay. This suggests that strategic delay is another source for the inefficient allocation of land, in addition to the public-good type externality pointed out by Grossman and Hart [Bell Journal of Economics 11 (1980) 42] and O'Flaherty [Regional Science and Urban Economics 24 (1994) 287]. (C) 2004 Elsevier B.V. All rights reserved.

Process development for a recombinant Chinese Hamster Ovary (CHO) cell line utilizing a metal induced and amplified metallothionein expression system

The suspension Chinese Hamster Ovary cell line, 13-10-302, utilizing the metallothionein (MT) expression system producing recombinant human growth hormone (hGH) was studied in a serum-free and cadmium-free medium at different fermentation scales and modes of operation. Initial experiments were carried out to optimize the concentration of metal addition to induce the MT promoter. Subsequently, the cultivation of the 13-10-302 cell line was scaled up from spinner flasks into bioreactors, and the cultivation duration was extended with fed-batch and perfusion strategies utilizing 180 muM zinc to induce the promoter controlling expression of recombinant hGH. It was shown that a fed-batch process could increase the maximum cell numbers twofold, from 3.3 to 6.3 x 10(6) cell/mL, over those obtained in normal batch fermentations, and this coupled with extended fermentation times resulted in a fourfold increase in final hGH titer, from 135 +/- 15 to 670 +/- 70 mg/L at a specific productivity q(hGH) value of 12 pg cell(-1)d(-1). The addition of sodium butyrate increased the specific productivity of hGH in cells to a value of approximately 48 pg cell(-1)d(-1), resulting in a final hGH titer of over a gram per liter during fed-batch runs. A BioSep acoustic cell recycler was used to retain the cells in the bioreactor during perfusion operation. It was necessary to maintain the specific feeding rates (SFR) above a value of 0.2 vvd/(10(6) cell/mL) to maintain the viability and productivity of the 13-10-302 cells; under these conditions the viable cell number increased to over 107 cell/mL and resulted in a volumetric productivity of over 120 mg(hGH) L(-1)d(-1). Process development described in this work demonstrates cultivation at various scales and sustained high levels of productivity under cadmium free condition in a CHO cell line utilizing an inducible metallothionein expression system. (C) 2004 Wiley Periodicals, Inc.

The alpha-galactosyl derivatives of ganglioside GD(1b) are essential for the organization of lipid rafts in RBL-2H3 mast cells

Gangliosides are complex glycosphingolipids that are important in many biological processes. The present study investigated the role of gangliosides in the organization of lipid rafts in RBL-2H3 mast cells and in the modulation of mast cell degranulation via Fc epsilon RI. The role of gangliosides was examined using two ganglioside deficient cell lines (B6A4A2III-E5 and B6A4C1III-D1) as well as the parent cell line (RBL-2H3). All three cell lines examined express Fc epsilon RI, Lyn, Syk and LAT. However, only in RBL-2H3 cells were Fc epsilon RI, LAT and alpha-galactosyl derivatives of ganglioside GD(1b) mobilized to lipid raft domains following Fc epsilon RI stimulation. The inhibition of glycosphingolipid synthesis in RBL-2H3 cells also resulted in a decrease in the release of beta-hexosaminidase activity after Fc epsilon RI activation. The two mutant cell lines have a reduced release of beta-hexosaminidase activity after Fc epsilon RI stimulation, but not after exposure to calcium ionophore. These results indicate that the alpha-galactosyl derivatives of ganglioside GD(1b) are important in the initial events of Fc epsilon RI signaling upstream of Ca(2+) influx. Since the initial signaling events occur in lipid rafts and in the mutant cell lines the rafts are disorganized, these results also suggest that these gangliosides contribute to the correct assembly of lipid rafts and are essential for mast cell activation via Fc epsilon RI. (c) 2008 Published by Elsevier Inc.