Biomedical subjects

Kuniaki Takahashi

Publications and source records attributed to Kuniaki Takahashi.

At least 19 recordsLinked to original sources

PubMed2006

Differential microarray analysis of Drosophila mushroom body transcripts using chemical ablation.

Mushroom bodies (MBs) are the centers for olfactory associative learning and elementary cognitive functions in the Drosophila brain. As a way to systematically elucidate genes preferentially expressed in MBs, we have analyzed genome-wide alterations in transcript profiles associated with MB ablation by hydroxyurea. We selected 100 genes based on microarray data and examined their expression patterns in the brain by in situ hybridization. Seventy genes were found to be expressed in the posterodorsal cortex, which harbors the MB cell bodies. These genes encode proteins of diverse functions, including transcription, signaling, cell adhesion, channels, and transporters. Moreover, we have examined developmental functions of 40 of the microarray-identified genes by transgenic RNA interference; 8 genes were found to cause mild-to-strong MB defects when suppressed with a MB-Gal4 driver. These results provide important information not only on the repertoire of genes that control MB development but also on the repertoire of neural factors that may have important physiological functions in MB plasticity.

Animals↗

PubMed2006

Control of axonal sprouting and dendrite branching by the Nrg-Ank complex at the neuron-glia interface.

Neurons are highly polarized cells with distinct subcellular compartments, including dendritic arbors and an axon. The proper function of the nervous system relies not only on correct targeting of axons, but also on development of neuronal-class-specific geometry of dendritic arbors [1-4]. To study the intercellular control of the shaping of dendritic trees in vivo, we searched for cell-surface proteins expressed by Drosophila dendritic arborization (da) neurons [5-7]. One of them was Neuroglian (Nrg), a member of the Ig superfamily ; Nrg and vertebrate L1-family molecules have been implicated in various aspects of neuronal wiring, such as axon guidance, axonal myelination, and synapse formation [9-12]. A subset of the da neurons in nrg mutant embryos exhibited deformed dendritic arbors and abnormal axonal sprouting. Our functional analysis in a cell-type-selective manner strongly suggested that those da neurons employed Nrg to interact with the peripheral glia for suppressing axonal sprouting and for forming second-order dendritic branches. At least for the former role, Nrg functioned in concert with the intracellular adaptor protein Ankyrin (Ank) [13]. Thus, the neuron-glia interaction that is mediated by Nrg, together with Ank under some situations, contributes to axonal and dendritic morphogenesis.

Animals↗

PubMed2006

Essential role of the apoptotic cell engulfment genes draper and ced-6 in programmed axon pruning during Drosophila metamorphosis.

Axon pruning is a common phenomenon in neural circuit development. Previous studies demonstrate that the engulfing action of glial cells is essential in this process. The underlying molecular mechanisms, however, remain unknown. We show that draper (drpr) and ced-6, which are essential for the clearance of apoptotic cells in C. elegans, function in the glial engulfment of larval axons during Drosophila metamorphosis. The drpr mutation and glia-specific knockdown of drpr and ced-6 by RNA interference suppress glial engulfment, resulting in the inhibition of axon pruning. drpr and ced-6 interact genetically in the glial action. Disruption of the microtubule cytoskeleton in the axons to be pruned occurs via ecdysone signaling, independent of glial engulfment. These findings suggest that glial cells engulf degenerating axons through drpr and ced-6. We propose that apoptotic cells and degenerating axons of living neurons are removed by a similar molecular mechanism.

Animals↗

PubMed2006

Enterolithiasis in a cat.

An 8-year-old female Persian cat was brought in for evaluation of chronic vomiting. The presence of opaque enteric foreign bodies and intestinal obstruction along with azotaemia, hyperphosphataemia, moderate anaemia and peritoneal fluid were revealed following appropriate diagnostic work-up. Exploratory laparotomy confirmed jejunoileal dilation, ileocaecal stenosis, and numerous foreign objects in the jejunoileum. These foreign objects and ileocaecal stenosis were surgically removed, and intestinal resection and anastomosis was performed. The patient recovered favourably. Analysis revealed that the foreign objects were composed of calcium phosphate and calcium carbonate. Intestinal inflammation and stenosis secondary to enterolithiasis may have developed following ingestion of cat litter or a previous unrelated surgical intervention. We were unable to delineate the inciting pathogenesis in this particular case.

Animals↗

PubMed2006

The Toll immune-regulated Drosophila protein Fondue is involved in hemolymph clotting and puparium formation.

Clotting is critical in limiting hemolymph loss and initiating wound healing in insects as in vertebrates. It is also an important immune defense, quickly forming a secondary barrier to infection, immobilizing bacteria and thereby promoting their killing. However, hemolymph clotting is one of the least understood immune responses in insects. Here, we characterize fondue (fon; CG15825), an immune-responsive gene of Drosophila melanogaster that encodes an abundant hemolymph protein containing multiple repeat blocks. After knockdown of fon by RNAi, bead aggregation activity of larval hemolymph is strongly reduced, and wound closure is affected. fon is thus the second Drosophila gene after hemolectin (hml), for which a knockdown causes a clotting phenotype. In contrast to hml-RNAi larvae, clot fibers are still observed in samples from fon-RNAi larvae. However, clot fibers from fon-RNAi larvae are more ductile and longer than in wt hemolymph samples, indicating that Fondue might be involved in cross-linking of fiber proteins. In addition, fon-RNAi larvae exhibit melanotic tumors and constitutive expression of the antifungal peptide gene Drosomycin (Drs), while fon-RNAi pupae display an aberrant pupal phenotype. Altogether, our studies indicate that Fondue is a major hemolymph protein required for efficient clotting in Drosophila.

Animals↗

PubMed2006

Drosophila immunity: a large-scale in vivo RNAi screen identifies five serine proteases required for Toll activation.

Unlike mammalian Toll-like Receptors, the Drosophila Toll receptor does not interact directly with microbial determinants but is rather activated upon binding a cleaved form of the cytokine-like molecule Spatzle (Spz). During the immune response, Spz is thought to be processed by secreted serine proteases (SPs) present in the hemolymph that are activated by the recognition of gram-positive bacteria or fungi . In the present study, we have used an in vivo RNAi strategy to inactivate 75 distinct Drosophila SP genes. We then screened this collection for SPs regulating the activation of the Toll pathway by gram-positive bacteria. Here, we report the identification of five novel SPs that function in an extracellular pathway linking the recognition proteins GNBP1 and PGRP-SA to Spz. Interestingly, four of these genes are also required for Toll activation by fungi, while one is specifically associated with signaling in response to gram-positive bacterial infections. These results demonstrate the existence of a common cascade of SPs upstream of Spz, integrating signals sent by various secreted recognition molecules via more specialized SPs.

Animals↗

PubMed2006

The MAPKKK Mekk1 regulates the expression of Turandot stress genes in response to septic injury in Drosophila.

Septic injury triggers a rapid and widespread response in Drosophila adults that involves the up-regulation of many genes required to combat infection and for wound healing. Genome-wide expression profiling has already demonstrated that this response is controlled by signaling through the Toll, Imd, JAK-STAT and JNK pathways. Using oligonucleotide microarrays, we now demonstrate that the MAPKKK Mekk1 regulates a small subset of genes induced by septic injury including Turandot (Tot) stress genes. Our analysis indicates that Tot genes show a complex regulation pattern including signals from both the JAK-STAT and Imd pathways and Mekk1. Interestingly, Mekk1 flies are resistant to microbial infection but susceptible to paraquat, an inducer of oxidative stress. These results point to a role of Mekk1 in the protection against tissue damage and/or protein degradation and indicate complex interactions between stress and immune pathways in Drosophila.

Animals↗

PubMed2006

A case of protein-losing enteropathy treated with methotrexate in a dog.

A 9-year-old female Pug was presented to us with chronic diarrhea. Hematologic findings indicated severe hypoproteinemia and hypoalbuminemia, and endoscopy revealed severe edema of the duodenal mucosa. Based on these results and on additional histopathological findings, we made a diagnosis of protein-losing enteropathy caused by lymphocytic-plasmacytic enteritis with lymphangiectasia. The dog was initially treated with prednisolone and cyclosporine. This treatment regimen was not effective. However, when methotrexate was substituted for cyclosporine, progress was obtained and the diagnosis was confirmed.

Animals↗

PubMed2006

Extraction of uranium from simulated ore by the supercritical carbon dioxide fluid extraction method with nitric acid-TBP complex.

The supercritical fluid extraction (SFE) method using CO(2) as a medium with an extractant of HNO(3)-tri-n-butyl phosphate (TBP) complex was applied to extract uranium from several uranyl phosphate compounds and simulated uranium ores. An extraction method consisting of a static extraction process and a dynamic one was established, and the effects of the experimental conditions, such as pressure, temperature, and extraction time, on the extraction of uranium were ascertained. It was found that uranium could be efficiently extracted from both the uranyl phosphates and simulated ores by the SFE method using CO(2). It was thus demonstrated that the SFE method using CO(2) is useful as a pretreatment method for the analysis of uranium in ores.

Journal Article↗

PubMed2005

G-protein gamma subunit 1 is required for sugar reception in Drosophila.

Though G-proteins have been implicated in the primary step of taste signal transduction, no direct demonstration has been done in insects. We show here that a G-protein gamma subunit, Ggamma1, is required for the signal transduction of sugar taste reception in Drosophila. The Ggamma1 gene is expressed mainly in one of the gustatory receptor neurons. Behavioral responses of the flies to sucrose were reduced by the targeted suppression of neural functions of Ggamma1-expressing cells using neural modulator genes such as the modified Shaker K+ channel (EKO), the tetanus toxin light chain or the shibire (shi(ts1)) gene. RNA interference targeting to the Ggamma1 gene reduced the amount of Ggamma1 mRNA and suppressed electrophysiological response of the sugar receptor neuron. We also demonstrated that responses to sugars were lowered in Ggamma1 null mutant, Ggamma1(N159). These results are consistent with the hypothesis that Ggamma1 participates in the signal transduction of sugar taste reception.

Animals↗

PubMed2005

The transmembrane protein, Tincar, is involved in the development of the compound eye in Drosophila melanogaster.

We previously cloned and characterized the Drosophila gene, tincar (tinc), which encodes a novel protein with eight putative transmembrane domains. Here, we have studied the expression pattern and functions of tinc during developmental processes. tinc mRNA is expressed in the central and peripheral nervous systems, and midgut during embryogenesis. In the third-instar larval eye disc, tinc mRNA is strongly expressed in all the differentiating ommatidial cells within and in the vicinity of the morphogenetic furrow. Loss-of-function analysis using the RNA-interference method revealed severe defects of eye morphogenesis during the late developmental stages. Our results suggested that tinc may have an indispensable role in the normal differentiation of ommatidial cells.

Animals↗

PubMed2004

An Alternative Approach to Atopic Dermatitis: Part II-Summary of Cases and Discussion.

In the first part of this Review, we presented case-series where Kampo treatment was introduced for those atopic dermatitis (AD) patients who had failed with conventional therapy, in an attempt to prove that there exists a definite subgroup of AD patients for whom Kampo treatment is effective. In this second part, we will first provide the summary of the results for 140 AD patients we treated in 2000. The results suggest that Kampo treatment is effective for more than half of AD patients who fail with conventional therapy. In the Discussion, we will examine the evidential basis for conventional AD therapy and discuss how Kampo treatment should be integrated into the guidelines for AD therapy. We contend that Kampo treatment should be tried before systematic immunosuppressive agents are considered. As each Kampo treatment is highly individualized, it should be regarded more as 'art' than technology, and special care should be taken to assess its efficacy in clinical trial.

Journal Article↗

PubMed2004

The twisted abdomen phenotype of Drosophila POMT1 and POMT2 mutants coincides with their heterophilic protein O-mannosyltransferase activity.

Walker-Warburg syndrome, caused by mutations in protein O-mannosyltransferase-1 (POMT1), is an autosomal recessive disorder characterized by severe brain malformation, muscular dystrophy, and structural eye abnormalities. As humans have a second POMT, POMT2, we cloned each Drosophila ortholog of the human POMT genes and carried out RNA interference (RNAi) knock-down to investigate the function of these proteins in vivo. Drosophila POMT2 (dPOMT2) RNAi mutant flies showed a "twisted abdomen phenotype," in which the abdomen is twisted 30-60 degrees , similar to the dPOMT1 mutant. Moreover, dPOMT2 interacted genetically with dPOMT1, suggesting that the dPOMTs function in collaboration with each other in vivo. We expressed dPOMTs in Sf21 cells and measured POMT activity. dPOMT2 transferred a mannose to the dystroglycan protein only when it was coexpressed with dPOMT1. Likewise, dPOMT1 showed POMT activity only when coexpressed with dPOMT2, and neither dPOMT showed any activity by itself. Each dPOMT RNAi fly totally reduced POMT activity, despite the specific reduction in the level of each dPOMT mRNA. The expression pattern of dPOMT2 mRNA was found to be similar to that of dPOMT1 mRNA using whole mount in situ hybridization. These results demonstrate that the two dPOMTs function as a protein O-mannosyltransferase in association with each other, in vitro and in vivo, to generate and maintain normal muscle development.

Amino Acid Sequence↗

PubMed2004

An Alternative Approach to Atopic Dermatitis: Part I-Case-Series Presentation.

Atopic dermatitis (AD) is a complex disease of obscure pathogenesis. A substantial portion of AD patients treated with conventional therapy become intractable after several cycles of recurrence. Over the last 20 years we have developed an alternative approach to treat many of these patients by diet and Kampo herbal medicine. However, as our approach is highly individualized and the Kampo formulae sometimes complicated, it is not easy to provide evidence to establish usefulness of this approach. In this Review, to demonstrate the effectiveness of the method of individualized Kampo therapy, results are presented for a series of patients who had failed with conventional therapy but were treated afterwards in our institution. Based on these data, we contend that there exist a definite subgroup of AD patients in whom conventional therapy fails, but the 'Diet and Kampo' approach succeeds, to heal. Therefore, this approach should be considered seriously as a second-line treatment for AD patients. In the Discussion, we review the evidential status of the current conventional strategies for AD treatment in general, and then specifically discuss the possibility of integrating Kampo regimens into it, taking our case-series presented here as evidential basis. We emphasize that Kampo therapy for AD is more 'art' than technology, for which expertise is an essential pre-requisite.

Journal Article↗

PubMed2004

In vivo RNA interference analysis reveals an unexpected role for GNBP1 in the defense against Gram-positive bacterial infection in Drosophila adults.

The Drosophila immune system discriminates between different classes of infectious microbes and responds with pathogen-specific defense reactions via the selective activation of the Toll and the immune deficiency (Imd) signaling pathways. The Toll pathway mediates most defenses against Gram-positive bacteria and fungi, whereas the Imd pathway is required to resist Gram-negative bacterial infection. Microbial recognition is achieved through peptidoglycan recognition proteins (PGRPs); Gram-positive bacteria activate the Toll pathway through a circulating PGRP (PGRP-SA), and Gram-negative bacteria activate the Imd pathway via PGRP-LC, a putative transmembrane receptor, and PGRP-LE. Gram-negative binding proteins (GNBPs) were originally identified in Bombyx mori for their capacity to bind various microbial compounds. Three GNBPs and two related proteins are encoded in the Drosophila genome, but their function is not known. Using inducible expression of GNBP1 double-stranded RNA, we now demonstrate that GNBP1 is required for Toll activation in response to Gram-positive bacterial infection; GNBP1 double-stranded RNA expression renders flies susceptible to Gram-positive bacterial infection and reduces the induction of the antifungal peptide encoding gene Drosomycin after infection by Gram-positive bacteria but not after fungal infection. This phenotype induced by GNBP1 inactivation is identical to a loss-of-function mutation in PGRP-SA, and our genetic studies suggest that GNBP1 acts upstream of the Toll ligand Spätzle. Altogether, our results demonstrate that the detection of Gram-positive bacteria in Drosophila requires two putative pattern recognition receptors, PGRP-SA and GNBP1.

Animals↗

PubMed2004

A Drosophila salivary gland mucin is also expressed in immune tissues: evidence for a function in coagulation and the entrapment of bacteria.

Our studies on the developmental regulation of glycosylation in Drosophila melanogaster led us to identify and characterize gp150, an ecdysone-regulated mucin that is found in hemocytes, the gut (peritrophic membrane) and in the salivary glands. We are particularly interested in mucin immune functions and found that gp150 is released from larval hemocytes, becomes part of the clot and participates in the entrapment of bacteria. By RT-PCR and RNAi experiments, we identified gp150 as the previously described I71-7, an ecdysone-induced salivary glue protein. We discuss the evolutionary and biochemical implications of the dual use of salivary proteins for immune functions in insects. Further molecular characterization of such shared proteins may enable a better understanding of the properties of proteins involved in containment and elimination of microbes, as well as hemostasis and wound repair.

Amino Acid Sequence↗

PubMed2003

Synaptic inputs from low threshold afferents of trunk muscles to motoneurons innervating the longissimus lumborum muscle in the spinal cat.

(1) We studied the reflex actions of group I and II afferents to longissimus lumborum (Long) motoneurons in the L1-L5 spinal segments from the epaxial muscle, m. Long, and the hypaxial muscle, m. obliquus externus abdominus (OEA). (2) Postsynaptic potentials (PSPs) recorded from 140 Long motoneurons in 30 spinal cats were analyzed. Under the present experimental conditions, the stimulation of Long and OEA nerves at an intensity below 1.5 times threshold (T) activated only group I muscle afferents, while stimulations at 2-5T activated group II muscle afferents as well. (3) The incidence of PSPs was related to the proximity of the spinal segments of the nerves stimulated to the spinal segment of the motoneurons; the shorter the distance the larger the PSPs and higher incidence of PSPs. The Long motoneurons received group I afferent input mainly from the same and adjacent segments, and received group II afferent inputs from a wider range of segments. (4) A short (i.e., less than 1.0 ms) latency of excitatory PSPs (EPSPs) evoked by ipsilateral group I afferents of Long at the same or adjacent segment indicated a monosynaptic connection. In general, the central latencies became longer as the distance between spinal segments of stimulated nerves and motoneurons increased. Major PSP components were produced by polysynaptic neuronal pathways. The spatial facilitation between PSPs evoked by afferents of different nerves (i.e., ipsilateral Long (iLong) and contralateral Long (cLong) of the same segment; iLongs of different segments; and iLong and iOEA of the same segment) indicated that they shared common interneurons. (5) Although iLong and iOEA muscle afferents produced predominantly EPSPs, and contralateral muscle afferents elicited predominantly IPSPs in Long motoneurons at each spinal segment, the patterns of convergence from Long and OEA muscle afferents of different spinal segments and of different sides differed considerably among motoneurons. (6) These findings demonstrated various input patterns of individual motoneurons within the same motoneuron pool, which might reflect the complexity of neuronal control of the back muscles for various trunk movements, including lateral and dorsal bending, rotating, and fixation of the trunk.

Action Potentials↗

PubMed2003

Role of 5-aminolevulinic acid (ALA) on active oxygen-scavenging system in NaCl-treated spinach (Spinacia oleracea).

ALA is a key precursor in the biosynthesis of porphyrins such as chlorophyll and heme, and was found to induce temporary elevations in the photosynthesis rate, APX, and CAT; furthermore, treatment with ALA at a low concentration might be correlated to the increase of NaCl tolerance of spinach plants. The photosynthetic rate and the levels of active oxygen-scavenging system in the 3rd leaf of spinach (Spinacia oleracea) plants grown by foliar treatment with 0, 0.18, 0.60 and 1.80 mmol/L 5-aminolevulinic acid under 50 and 100 mmol/L NaCl were analyzed. Plants treated with 0.60 and 1.80 mmol/L ALA showed significant increases in the photosynthetic rate at 50 and 100 mmol/L NaCl, while that of 0.18 mmol/L ALA did not show any changes at 50 mmol/L NaCl and a gradual decrease at 100 mmol/L NaCl. In contrast, the rate with 0 mmol/L ALA showed reduction at both concentrations of NaCl. The increase of hydrogen peroxide content by treatment with 0.60 and 1.80 mmol/L ALA were more controlled than that of 0 mmol/L ALA under both NaCl conditions. These ALA-treated spinach leaves also exhibited a lower oxidized/reduced ascorbate acid ratio and a higher reduced/oxidized glutathione ratio than the 0 mmol/L-treated spinach leaves when grown at both NaCl conditions. With regard to the antioxidant enzyme activities in the leaves, ascorbate peroxidase, catalase, and glutathione reductase activities were enhanced remarkably, most notably at day 3, by treatment with 0.60 and 1.80 mmol/L ALA under both NaCl conditions in comparison to that of 0 and 0.18 mmol/L ALA. These data indicate that the protection against oxidative damage by higher levels of antioxidants and enzyme activities, and by a more active ascorbate-glutathione cycle related to the increase of the photosynthesis rate, could be involved in the increased salt tolerance observed in spinach by treatment with 0.60 to 1.80 mmol/L ALA with NaCl.

Aminolevulinic Acid↗