A Lactobacillus mutant capable of accumulating long-chain polyphosphates that enhance intestinal barrier function

Inorganic polyphosphate (polyP) was previously identified as a probiotic-derived substance that enhances intestinal barrier function. PolyP-accumulating bacteria are expected to have beneficial effects on the human gastrointestinal tract. In this study, we selected Lactobacillus paracasei JCM 1163 as a strain with the potential to accumulate polyP, because among the probiotic bacteria stored in our laboratory, it had the largest amount of polyP. The chain length of polyP accumulated in L. paracasei JCM 1163 was approximately 700 phosphate (Pi) residues. L. paracasei JCM 1163 accumulated polyP when Pi was added to Pi-starved cells. We further improved the ability of L. paracasei JCM 1163 to accumulate polyP by nitrosoguanidine mutagenesis. The mutant accumulated polyP at a level of 1500 nmol/mg protein—approximately 190 times that of the wild-type strain. PolyP extracted from the L. paracasei JCM 1163 significantly suppressed the oxidant-induced intestinal permeability in mouse small intestine. In conclusion, we have succeeded in breeding the polyP-accumulating Lactobacillus mutant that is expected to enhance intestinal barrier function.     

Homophilic Dscam interactions control complex dendrite morphogenesis

Alternative splicing of the Drosophila gene Dscam results in up to 38,016 different receptor isoforms proposed to interact by isoform-specific homophilic binding. We report that Dscam controls cell-intrinsic aspects of dendrite guidance in all four classes of dendrite arborization (da) neurons. Loss of Dscam in single neurons causes a strong increase in self-crossing. Restriction of dendritic fields of neighboring class III neurons appeared intact in mutant neurons, suggesting that dendritic self-avoidance, but not heteroneuronal tiling, may depend on Dscam. Overexpression of the same Dscam isoforms in two da neurons with overlapping dendritic fields forced a spatial segregation of the two fields, supporting the model that dendritic branches of da neurons use isoform-specific homophilic interactions to ensure minimal overlap. Homophilic binding of the highly diverse extracellular domains of Dscam may therefore limit the use of the same "core" repulsion mechanism to cell-intrinsic interactions without interfering with heteroneuronal interactions.     

Arsonic acid-presenting superparamagnetic iron oxide for pH-responsive aggregation under slightly acidic conditions

We report the synthesis and characterization of the arsonic acid-presenting superparamagnetic iron oxide (SPIO). We used arsonoacetic acid as the ligand for SPIOs in aqueous media. The surface modification of the SPIOs was accomplished via the ligand exchange from undecanoic acid to the carboxyl moiety of arsonoacetic acid. Consequently, the well-dispersed arsonic acid-presenting SPIOs in water were obtained. We found that the dispersion state of the arsonic acid-presenting SPIOs can be sharply regulated by pH changes in the biological significant region. The well dispersion state of the arsonic acid-presenting SPIOs can be maintained at the neutral pH region. In contrast, the arsonic acid-presenting SPIOs can sensitively form the aggregation below pH 6.1. Moreover, these dispersion states can be controlled reversibly by the pH alteration in the narrow region.     

Induction of insulin-like growth factor 2 expression in a mesenchymal cell line co-cultured with an ameloblast cell line

Various growth factors have been implicated in the regulation of cell proliferation and differentiation during tooth development. It has been unclear if insulin-like growth factors (IGFs) participate in the epithelium–mesenchyme interactions of tooth development. We previously produced three-dimensional sandwich co-culture systems (SW) containing a collagen membrane that induce the differentiation of epithelial cells. In the present study, we used the SW system to analyze the expression of IGFs and IGFRs. We demonstrate that IGF2 expression in mesenchymal cells was increased by SW. IGF1R transduces a signal; however, IGF2R does not transduce a signal. Recombinant IGF2 induces IGF1R and IGF2R expression in epithelial cells. IGF1R expression is increased by SW; however, IGF2R expression did not increase by SW. Thus, IGF2 signaling works effectively in SW. These results suggest that IGF signaling acts through the collagen membrane on the interaction between the epithelium and mesenchyme. In SW, other cytokines may be suppressed to induce IGF2R induction. Our results suggest that IGF2 may play a role in tooth differentiation.     

Plasma interleukin (IL)-18 (interferon-gamma-inducing factor) and other inflammatory cytokines in patients with gouty arthritis and monosodium urate monohydrate crystal-induced secretion of IL-18

To determine whether levels of interleukin (IL)-18, together with those of IL-1beta, tumor necrosis factor-alpha, IL-6, and IL-8, are elevated in the plasma of patients with gouty arthritis, the plasma concentrations of those cytokines were measured in 31 males with gouty arthritis. Further, CD14+ cells were obtained from human blood and thioglycolate medium-induced peritoneal cells obtained from caspase 1-deficient mice, and then separately cultured in the presence of monosodium urate monohydrate (MSU) crystals. In addition, in an animal in vivo experiment, MSU crystals were injected into subcutaneous air pouches of IL-18-deficient mice. The plasma concentrations of IL-18, IL-6, and IL-8 were elevated in the presence of gouty arthritis in the gout patients. In the in vitro study, the presence of MSU crystals stimulated CD14+ cells (monocytes) to secrete IL-18 and increased the activity of caspase 1 in CD14+ cells, whereas there was no significant effect on IL-18 messenger RNA in CD14+ cells and only a slight induction of IL-18 secretion from thioglycolate medium-induced caspase 1-deficient peritoneal cells. In the in vivo experiment, MSU crystals injected into the air pouch promoted neutrophil accumulation along with an increase in concentrations of keratinocyte-derived chemokine (KC) and macrophage inflammatory protein (MIP)-1alpha in air-pouch fluids in both IL-18-deficient and wild-type mice. However, there was no increase in the concentration of IL-18 in air-pouch fluids in either mouse strain. Our results suggest that plasma IL-18, IL-6, IL-8, and C-reactive protein (CRP) levels reflect local inflammation associated with gouty arthritis, though IL-18 does not play an important role in neutrophil accumulation. Further, they suggest that MSU crystals accelerate the processing of IL-18 from an inactive to active form via the activation of caspase 1.     

Variable Numbers of Tandem Repeats in Plasmodium falciparum Genes

Genome variation studies in Plasmodium falciparum have focused on SNPs and, more recently, large-scale copy number polymorphisms and ectopic rearrangements. Here, we examine another source of variation: variable number tandem repeats (VNTRs). Interspersed low complexity features, including the well-studied P. falciparum microsatellite sequences, are commonly classified as VNTRs; however, this study is focused on longer coding VNTR polymorphisms, a small class of copy number variations. Selection against frameshift mutation is a main constraint on tandem repeats (TRs) in coding regions, while limited propagation of TRs longer than 975 nt total length is a minor restriction in coding regions. Comparative analysis of three P. falciparum genomes reveals that more than 9% of all P. falciparum ORFs harbor VNTRs, much more than has been reported for any other species. Moreover, genotyping of VNTR loci in a drug-selected line, progeny of a genetic cross, and 334 field isolates demonstrates broad variability in these sequences. Functional enrichment analysis of ORFs harboring VNTRs identifies stress and DNA damage responses along with chromatin modification activities, suggesting an influence on genome mutability and functional variation. Analysis of the repeat units and their flanking regions in both P. falciparum and Plasmodium reichenowi sequences implicates a replication slippage mechanism in the generation of TRs from an initially unrepeated sequence. VNTRs can contribute to rapid adaptation by localized sequence duplication. They also can confound SNP-typing microarrays or mapping short-sequence reads and therefore must be accounted for in such analyses.     

A bit-parallel dynamic programming algorithm suitable for DNA sequence alignment

Myers' elegant and powerful bit-parallel dynamic programming algorithm for approximate string matching has a restriction that the query length should be within the word size of the computer, typically 64. We propose a modification of Myers' algorithm, in which the modification has a restriction not on the query length but on the maximum number of mismatches (substitutions, insertions, or deletions), which should be less than half of the word size. The time complexity is O(m log |Σ|), where m is the query length and |Σ| is the size of the alphabet Σ. Thus, it is particularly suited for sequences on a small alphabet such as DNA sequences. In particular, it is useful in quickly extending a large number of seed alignments against a reference genome for high-throughput short-read data produced by next-generation DNA sequencers.