Wnt and BMP signaling govern lineage segregation of melanocytes in the avian embryo

Recent studies show that specification of some neural crest lineages occurs prior to or at the time of migration from the neural tube. We investigated what signaling events establish the melanocyte lineage, which has been shown to migrate from the trunk neural tube after the neuronal and glial lineages. Using in situ hybridization, we find that, although Wnts are expressed in the dorsal neural tube throughout the time when neural crest cells are migrating, the Wnt inhibitor cfrzb-1 is expressed in the neuronal and glial precursors and not in melanoblasts. This expression pattern suggests that Wnt signaling may be involved in specifying the melanocyte lineage. We further report that Wnt-3a-conditioned medium dramatically increases the number of pigment cells in quail neural crest cultures while decreasing the number of neurons and glial cells, without affecting proliferation. Conversely, BMP-4 is expressed in the dorsal neural tube throughout the time when neural crest cells are migrating, but is decreased coincident with the timing of melanoblast migration. This expression pattern suggests that BMP signaling may be involved in neural and glial cell differentiation or repression of melanogenesis. Purified BMP-4 reduces the number of pigment cells in culture while increasing the number of neurons and glial cells, also without affecting proliferation. Our data suggest that Wnt signaling specifies melanocytes at the expense of the neuronal and glial lineages, and further, that Wnt and BMP signaling have antagonistic functions in the specification of the trunk neural crest.     

Molecular aspects of self-incompatibility in Brassica species.

Many flowering plants possess self-incompatibility (SI) systems to prevent inbreeding. SI in Brassica species is controlled by a single S locus with multiple alleles. In recent years, much progress has been made in determining the male and female S determinant in Brassica species. In the female, a gain-of-function experiment clearly demonstrated that SRK was the sole S determinant, and that SLG enhanced the SI recognition process. By contrast, the male S determinant (termed SP11/SCR) was identified in the course of genome analysis of S locus to be a small cysteine-rich protein, which was classified as a pollen coat protein. This SP11/SCR may function as a ligand for the S domain of SRK in the SI recognition reaction of Brassica species.     

Alteration of the Systemic and Microcirculation by a Single Oral Dose of Flavan-3-Ols

Several systematic reviews have reported that flow mediated dilatation (FMD) was significantly increased in subjects after ingestion of chocolate that contains flavan-3-ols; however, the mechanisms responsible for this effect are not clear. In this study, we evaluated the effects of a single oral dose of flavan-3-ols on the systemic circulation and microcirculation in the cremaster muscle using intravital video microscopy in vivo. The cremaster muscle in rats was spread over a plastic chamber and a gastric tube was placed into the stomach. Blood flow in the cremasteric artery was determined using a laser Doppler flowmeter, while blood pressure and heart rate were measured by the tail-cuff method. Red blood cell velocity in arterioles and blood flow in the artery were significantly increased 5 min after the administration of 10 mg/kg flavan-3-ols compared with distilled water treatment. The number of capillaries recruited in the cremaster muscle was also significantly increased 15 min after treatment. Microscopic observation confirmed that increased shear stress on endothelial cells was maintained during the measurement period. The mean arterial blood pressure and heart rate were also significantly elevated soon after administration and returned to baseline before the end of the observation period. Plasma nitrate and nitrite levels, and NO phosphorylation of aortic tissue were significantly increased at 60 min after administration of flavan-3-ols. According to these results, a single oral dose of flavan-3-ols elevates blood pressure and flow transiently, and these effects induce NO production through increased shear stress on endothelial cells.     

Kawasaki Disease-Specific Molecules in the Sera Are Linked to Microbe-Associated Molecular Patterns in the Biofilms

Background

Kawasaki disease (KD) is a systemic vasculitis of unknown etiology. The innate immune system is involved in its pathophysiology at the acute phase. We have recently established a novel murine model of KD coronary arteritis by oral administration of a synthetic microbe-associated molecular pattern (MAMP). On the hypothesis that specific MAMPs exist in KD sera, we have searched them to identify KD-specific molecules and to assess the pathogenesis.

Methods

We performed liquid chromatography-mass spectrometry (LC-MS) analysis of fractionated serum samples from 117 patients with KD and 106 controls. Microbiological and LC-MS evaluation of biofilm samples were also performed.

Results

KD samples elicited proinflammatory cytokine responses from human coronary artery endothelial cells (HCAECs). By LC-MS analysis of KD serum samples collected at 3 different periods, we detected a variety of KD-specific molecules in the lipophilic fractions that showed distinct m/z and MS/MS fragmentation patterns in each cluster. Serum KD-specific molecules showed m/z and MS/MS fragmentation patterns almost identical to those of MAMPs obtained from the biofilms formed in vitro (common MAMPs from Bacillus cereus, Yersinia pseudotuberculosis and Staphylococcus aureus) at the 1^st study period, and from the biofilms formed in vivo (common MAMPs from Bacillus cereus, Bacillus subtilis/Bacillus cereus/Yersinia pseudotuberculosis and Staphylococcus aureus) at the 2^nd and 3^rd periods. The biofilm extracts from Bacillus cereus, Bacillus subtilis, Yersinia pseudotuberculosis and Staphylococcus aureus also induced proinflammatory cytokines by HCAECs. By the experiments with IgG affinity chromatography, some of these serum KD-specific molecules bound to IgG.

Conclusions

We herein conclude that serum KD-specific molecules were mostly derived from biofilms and possessed molecular structures common to MAMPs from Bacillus cereus, Bacillus subtilis, Yersinia pseudotuberculosis and Staphylococcus aureus. Discovery of these KD-specific molecules might offer novel insight into the diagnosis and management of KD as well as its pathogenesis.     

Transfer of cholesterol and diacylglycerol from lipophorin to Bombyx mori ovarioles in vitro: role of the lipid transfer particle

The objective of this study was to characterize the transfer of diacylglycerol (DAG) and cholesterol from larval Bombyx mori lipophorin to ovarioles. Transfer studies were carried out by incubating pupal ovarioles (5-day) with [(3)H]-cholesterol and [(3)H]-DAG-labeled lipophorin under different conditions. Transfer of both cholesterol and DAG exhibited hyperbolic dependency on lipophorin concentration with apparent Km values of 0.83 +/- 0.17 mg/ml and 0.74 +/- 0.16 mg/ml, respectively. Pretreatment of ovarioles with anti-lipid transfer particle (LTP) IgG significantly inhibited transfer of labeled DAG to ovarioles (75%) and not cholesterol. Injection of B. mori pupae (day 4) with anti-LTP IgG significantly affected the weight (65%), number of eggs (49%), amount of lipid (74%), and protein (65%) of the adult ovaries. Matured eggs had a very faint yellow color and deformed shape compared to controls. The inhibitory effect demonstrates the active role LTP plays in growth of ovaries, development, and oogenesis. The effect on vitellogenin shortage on egg development and maturation was determined by implanting ovaries in male recipients that lack vitellogenin. An 80% decline in egg production was observed. However, the mature eggs were normal in shape, color, and lipid content. Thus, restricting lipid or protein delivery to developing ovaries would dramatically affect choriogenesis.     

Glycosidase inhibition by cyclic sulfonium compounds

Inhibitory activities of various cyclic sulfonium compounds including salacinol against several glycosidases were studied and some compounds showed significant inhibition. The sulfonium ion structure was found to be essential for the inhibitory activity. Specific inhibition of salacinol toward rice alpha-glucosidase was ascribed to the tether arm.     

Mutational analysis of conserved aspartic acid residues in the <Emphasis Type="Italic">Methanothermobacter thermautotrophicus</Emphasis> MCM helicase

Minichromosome maintenance (MCM) helicases are thought to function as the replicative helicases in archaea and eukarya, unwinding the duplex DNA in the front of the replication fork. The archaeal MCM helicase can be divided into three parts, the N-terminal, catalytic, and C-terminal regions. The N-terminal part of the protein is divided into three domains, A, B, and C, and was shown to be involved in protein multimerization and binding to single- and double-stranded DNA. Two Asp residues found in domain C are conserved among MCM proteins from different archaea. These residues are located in a loop at the interface with domain A. Mutations of these residues in the Methanothermobacter thermautotrophicus MCM protein, Asp202 and Asp203, to Asn result in a significant reduction in the ability of the enzyme to bind DNA and in lower thermal stability. However, the mutant proteins retained helicase and ATPase activities. Further investigation of the DNA binding revealed that the presence of ATP rescues the DNA binding deficiencies by these mutant proteins. Possible roles of these conserved residues in MCM function are discussed.     

Prevention of allergic asthma by vaccination with transgenic rice seed expressing mite allergen: induction of allergen-specific oral tolerance without bystander suppression

This study tested the feasibility of oral immunotherapy for bronchial asthma using a newly developed subunit vaccine in which a fragment (p45-145) of mite allergen (Der p 1) containing immunodominant human and mouse T cell epitopes was encapsulated in endoplasmic reticulum-derived protein bodies of transgenic (Tg) rice seed. Allergen-specific serum immunoglobulin responses, T cell proliferation, Th1/Th2 cytokine production, airway inflammatory cell infiltration, bronchial hyper-responsiveness (BHR) and lung histology were investigated in allergen-immunized and -challenged mice. Prophylactic oral vaccination with the Tg rice seeds clearly reduced the serum levels of allergen-specific IgE and IgG. Allergen-induced CD4(+) T cell proliferation and production of Th2 cytokines in vitro, infiltration of eosinophils, neutrophils and mononuclear cells into the airways and BHR were also inhibited by oral vaccination. The effects of the vaccine were antigen-specific immune response because the levels of specific IgE and IgG in mice immunized with Der f 2 or ovalbumin were not significantly suppressed by oral vaccination with the Der p 1 expressing Tg rice. Thus, the vaccine does not induce nonspecific bystander suppression, which has been a problem with many oral tolerance regimens. These results suggest that our novel vaccine strategy is a promising approach for allergen-specific oral immunotherapy against allergic diseases including bronchial asthma.