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Biomedical subjects

K Awai

Publications and source records attributed to K Awai.

At least 19 recordsLinked to original sources
PubMed2006

Lipid trafficking between the endoplasmic reticulum and the chloroplast.

The photosynthetic (thylakoid) membrane of plants is one of the most extensive biological cell membrane systems found in Nature. It harbours the photosynthetic apparatus, which is essential to life on Earth as carbon dioxide is fixed and atmospheric oxygen released by photosynthesis. Lipid biosynthetic enzymes of different subcellular compartments participate in the biogenesis of the thylakoid membrane system. This process requires the extensive exchange of lipid precursors between the chloroplast and the ER (endoplasmic reticulum). The underlying lipid trafficking phenomena are not yet understood at the mechanistic level, but genetic mutants of the model plant Arabidopsis thaliana with disruptions in lipid trafficking between the ER and the chloroplast have recently become available. Their study has led to the identification of components of the lipid transfer machinery at the inner chloroplast envelope.

Arabidopsis↗
PubMed2003

[Development of computerized method for automated classification of the body parts in digital radiographs].

PURPOSE: In picture archiving and communication systems (PACS), the information on the body parts included in radiographs is often not or incorrectly recorded in an image header. In order to apply the computer-aided diagnosis (CAD) system in the PACS environment, the body parts in radiographs need to be recognized correctly by computer. The purpose of this study is to develop a computerized method for correctly classifying the body parts in digital radiographs based on a template matching technique. METHODS/MATERIALS: The image database used in this study was 1032 digital radiographs (14 x 17 inches) obtained with a computed radiography, and included 505 chest of postetroanterior view, 39 chest of lateral view, 241 abdomen, 108 pelvis, 10 upper limbs, 125 lower limbs, and 4 thoracic spine. In this method, test images were classified into four body parts, i.e., (1) chest, (2) abdomen, (3) pelvis, and (4) upper/lower limbs and thoracic spine. This computerized method was tested with 852 images, since 180 images were employed for creation of 98 templates, which represented the average radiographs for various body parts. Our approach was to examine the similarity of a given test image with templates by use of the cross-correlation values as the similarity measures. The body part of the test image was identified as the body part in the template yielding the maximum correlation value. Our method consisted of the following five steps. First, test images were classified into one of three groups; i.e. 1) chest and abdomen, 2) pelvis, and 3) upper/lower limbs and thoracic spine by using the templates obtained from images with the average size and position. Second, the remaining uncertain images were classified by using additional templates in various directions. Third, the chest and abdomen group was separated into two subgroups; i.e.chest and abdomen. Fourth, in order to classify some uncertain images, templates were shifted horizontally and vertically. Fifth, outer pixels of templates were eliminated to avoid the misclassification due to x-ray collimation. RESULTS: Our preliminary results indicated that the body parts for 850 cases (99.8%) were correctly classified with our method. CONCLUSIONS: This method would be useful for automated identification of the body parts in radiographs when various CAD systems would be implemented in the PACS environment.

Diagnosis, Computer-Assisted↗
PubMed2001

Two types of MGDG synthase genes, found widely in both 16:3 and 18:3 plants, differentially mediate galactolipid syntheses in photosynthetic and nonphotosynthetic tissues in Arabidopsis thaliana.

In Arabidopsis, monogalactosyldiacylglycerol (MGDG) is synthesized by a multigenic family of MGDG synthases consisting of two types of enzymes differing in their N-terminal portion: type A (atMGD1) and type B (atMGD2 and atMGD3). The present paper compares type B isoforms with the enzymes of type A that are known to sit in the inner membrane of plastid envelope. The occurrence of types A and B in 16:3 and 18:3 plants shows that both types are not specialized isoforms for the prokaryotic and eukaryotic glycerolipid biosynthetic pathways. Type A atMGD1 gene is abundantly expressed in green tissues and along plant development and encodes the most active enzyme. Its mature polypeptide is immunodetected in the envelope of chloroplasts from Arabidopsis leaves after cleavage of its transit peptide. atMGD1 is therefore likely devoted to the massive production of MGDG required to expand the inner envelope membrane and build up the thylakoids network. Transient expression of green fluorescent protein fusions in Arabidopsis leaves and in vitro import experiments show that type B precursors are targeted to plastids, owing to a different mechanism. Noncanonical addressing peptides, whose processing could not be assessed, are involved in the targeting of type B precursors, possibly to the outer envelope membrane where they might contribute to membrane expansion. Expression of type B enzymes was higher in nongreen tissues, i.e., in inflorescence (atMGD2) and roots (atMGD3), where they conceivably influence the eukaryotic structure prominence in MGDG. In addition, their expression of type B enzymes is enhanced under phosphate deprivation.

Amino Acid Sequence↗
PubMed2001

Monitoring of methyl jasmonate-responsive genes in Arabidopsis by cDNA macroarray: self-activation of jasmonic acid biosynthesis and crosstalk with other phytohormone signaling pathways.

Jasmonates mediate various physiological events in plant cells such as defense responses, flowering, and senescence through intracellular and intercellular signaling pathways, and the expression of a large number of genes appears to be regulated by jasmonates. In order to obtain information on the regulatory network of jasmonate-responsive genes (JRGs) in Arabidopsis thaliana (Arabidopsis), we screened 2880 cDNA clones for jasmonate responsiveness by a cDNA macroarray procedure. Since many of the JRGs reported so far have been identified in leaf tissues, the cDNA clones used were chosen from a non-redundant EST library that was prepared from above-ground organs. Hybridization to the filters was achieved using alpha-33P-labeled single-strand DNAs synthesized from mRNAs obtained from methyl jasmonate (MeJA)-treated and untreated Arabidopsis seedlings. Data analysis identified 41 JRGs whose mRNA levels were changed by more than three fold in response to MeJA. This was confirmed by Northern blot analysis by using eight representatives. Among the 41 JRGs identified, 5 genes were JA biosynthesis genes and 3 genes were involved in other signaling pathways (ethylene, auxin, and salicylic acid). These results suggest the existence of a positive feedback regulatory system for JA biosynthesis and the possibility of crosstalk between JA signaling and other signaling pathways.

Acetates↗
PubMed2001

Accumulation of plant galactolipid affects cell morphology of Escherichia coli.

Monogalactosyldiacylglycerol (MGDG) is a major constituent of thylakoid membrane in chloroplasts. Therefore, it is considered to have an important role in the maintenance of the complicated structure of the thylakoid membrane. We have succeeded in cloning the enzyme for MGDG synthesis and overexpressed it in Escherichia coli. In this study we analyzed the morphology of the E. coli harboring the gene. The fatty acid composition of its membrane lipids did not differ between the wild type and transformant, except for the appearance of MGDG. However, transformant cells appeared to be elongated. DAPI staining revealed the entire intracellular region of filamentous cells to be stained; therefore, the elongation of the cells is probably due to a defect in cell division. Atomic force microscopy revealed that the transformant had a smooth but scratched surface. It was concluded that the excessive accumulation of a non-bilayer lipid, MGDG, interfered with the translocation of proteins across the plasma membrane, including those for cell division.

Escherichia coli↗
PubMed2001

Selenium-based digital radiography versus high-resolution storage phosphor radiography in the detection of solitary pulmonary nodules without calcification: receiver operating characteristic curve analysis.

OBJECTIVE: The purpose of this study was to compare selenium-based digital radiography with high-resolution storage phosphor radiography for the detection of solitary pulmonary nodules without calcification. MATERIALS AND METHODS: One hundred twenty-four patients underwent selenium-based digital radiography, high-resolution storage phosphor radiography, and chest CT for evaluation of pulmonary nodules. Thirty-one patients with pulmonary nodules smaller than 3 cm in diameter and 40 patients with normal lungs were selected for receiver operating characteristic curve analysis. Five board-certified radiologists who were unaware of the CT results independently reviewed each of the hard copies of selenium-based digital radiography and storage phosphor radiography, identified pulmonary nodules, and graded their confidence for the presence of each nodule. For each radiologist, we calculated the areas under the receiver operating characteristic curve (AUC) for selenium-based digital radiography and storage phosphor radiography. RESULTS: The average performance of selenium-based digital radiography (AUC = 0.72) was higher than that of high-resolution storage phosphor radiography (AUC = 0.64), which is statistically significant (p <0.05). CONCLUSION: Our results indicate that selenium-based digital radiography is superior to high-resolution storage phosphor radiography for detecting solitary pulmonary nodules without calcification.

Adult↗
PubMed2000

Identification and light-induced expression of a novel gene of NADPH-protochlorophyllide oxidoreductase isoform in Arabidopsis thaliana.

In Arabidopsis thaliana, we identified a novel gene of a NADPH-protochlorophyllide oxidoreductase (POR) isoform, which catalyzes the light-dependent protochlorophyllide a reduction in the chlorophyll (Chl) biosynthetic pathway. The deduced amino acid sequence of the novel POR isoform (PORC) showed significant identities ( approximately 75%) with the previously isolated two POR isoforms of A. thaliana. Contrasting with these POR isoforms, the PORC transcript increased in etiolated seedlings by illumination, and was dominantly expressed in immature and mature tissues. The present results demonstrated that Chl biosynthesis and chloroplast biogenesis in A. thaliana are controlled by three POR isoforms, which are differentially controlled by light and development.

Amino Acid Sequence↗
PubMed2000

The multigenic family of monogalactosyl diacylglycerol synthases.

Because the synthesis of monogalactosyldiacylglycerol (MGDG) is unique to plants, identified as an important marker of the plastid envelope, involved in a key step of plastid biogenesis and is the most abundant lipid on earth, MGDG synthase activity was extensively analysed at the biochemical and physiological levels. In the present paper, we present our current knowledge on the MGDG synthase's function, structure and topology in envelope membranes, and discuss possible roles in plant cell glycerolipid metabolism. The recent discovery of a multigenic family of MGDG synthases raised the possibility that multiple isoenzymes might carry out MGDG synthesis in various tissues and developmental stages.

Amino Acid Sequence↗
PubMed2000

Transcriptional regulation by light and phytohormones of the MGD gene in cucumber.

Monogalactosyldiacylglycerol (MGDG) synthase catalyses formation of MGDG, a major structural lipid of chloroplasts. We have already cloned a cDNA for the synthase from a cucumber cDNA library and shown that expression of this gene is regulated by light and a phytohormone, cytokinin. In the present study, we report the molecular basis for transcriptional regulation by light and cytokinin in detail. First, in terms of the enzyme activity, gradual increases in activity mediated by light and cytokinin treatments were observed. At the same time, however, the changes in the mRNA level showed different profiles, with a transient peak during the early stages of light and cytokinin treatment. The interval between the peak level of mRNA and enzyme activation implies the existence of a post-transcriptional regulatory system. In addition, a genomic clone of MGDG synthase isolated from a cucumber genome library was used for a motif search in databases, and this revealed that putative cis-acting elements for light and phytohormones exist in the 5'-upstream region of the MGD cucumber gene. Detailed analysis of this region for light- and cytokinin-responsive activity was performed using a -90 truncated 35 S minimal promoter/luciferase (LUC) reporter gene. It resulted in high levels of LUC expression in etiolated cucumber cotyledons in response to illumination and cytokinin treatment. These results indicate that this 5'-upstream region is involved in light- and cytokinin-enhanced MGD gene expression, and that light- and cytokinin-responsive enhancements of the MGDG synthase activity are regulated, at least in part, at the level of transcription.

Adenine↗
PubMed2000

Genome-wide expression-monitoring of jasmonate-responsive genes of Arabidopsis using cDNA arrays.

Jasmonates are generally considered to mediate signalling, such as defence responses, flowering and senescence. However, factors involved in the jasmonate signal-transduction pathway remain unclear. To clarify the functions and signalling mechanisms of jasmonates on a genome-wide level, we adopted a cDNA macroarray technique. We prepared nylon filters of a cDNA macroarray on which 2880 independent expressed sequence tag clones of Arabidopsis were blotted, and hybridized (33)P-labelled single-strand DNAs synthesized from mRNAs of methyl jasmonate (MeJA)-treated and untreated Arabidopsis plants to the nylon filters. By analysing the data from the cDNA macroarray, we identified many function-known and unknown genes as MeJA-responsive genes, and confirmed that the profiles of the expression showed good agreement with Northern-blot analysis. These results demonstrate the efficiency of the cDNA macroarray for systematically analysing jasmonate-responsive genes on a genome-wide scale.

Acetates↗
PubMed1999

Biochemical and topological properties of type A MGDG synthase, a spinach chloroplast envelope enzyme catalyzing the synthesis of both prokaryotic and eukaryotic MGDG.

MGDG synthase, the enzyme that catalyzes the synthesis of the major chloroplast membrane lipid monogalactosyldiacylglycerol (MGDG), is encoded by a multigenic family. We have analyzed the biochemical properties, subcellular localization and membrane topology of a spinach chloroplast MGDG synthase, a representative member of the type A family from Spinacia oleracea (soMGD A), using a recombinant protein that was functionally overexpressed in Escherichia coli and specific polyclonal antibodies. We demonstrated that soMGD A could catalyze the synthesis of both 'prokaryotic' and 'eukaryotic' MGDG molecular species in vitro, with a selectivity for diacylglycerol similar to that of purified chloroplast envelope MGDG synthase activity. Furthermore, soMGD A was shown to be sensitive to chemical reagents (dithiothreitol, N-ethylmaleimide and o-phenanthroline) known to affect MGDG synthesis by the partially purified enzyme, as well as in isolated chloroplast envelope membranes. In spinach chloroplasts, soMGD A was localized by Western blot analysis in the inner envelope membrane. Topological studies demonstrated that soMGD A is a monotopic enzyme, embedded within one leaflet of the inner envelope membrane from spinach chloroplasts, a structure which may involve amphipathic alpha helices. We further demonstrated that in vitro, soMGD A precursor is imported and processed to its correct mature form in intact chloroplasts. These results show that soMGD A corresponds to a mature polypeptide of approximately 45 kDa. In addition, inactivation kinetics after gamma-ray irradiation strongly suggest that both native chloroplast envelope MGDG synthase and recombinant soMGD A have a functional molecular mass of 95-100 kDa, indicating that they are probably active as homodimers made of two 45-kDa subunits. This study suggests that, in spite of the growing evidence that MGDG synthesis is catalyzed by a multigenic family of enzymes, in spinach leaves both prokaryotic and eukaryotic MGDG syntheses could be attributable to a unique dimeric enzyme, provided that diacylglycerol is transported from the outer membrane to the inner membrane of the chloroplast envelope.

Amino Acid Sequence↗
PubMed1999

[Utility of thin-section/high-pitch helical CT for the assessment of small lung nodules].

To determine the utility of thin-section/high-pitch CT for the assessment of small lung nodules, a lung phantom was scanned on 1 mm width collimation at variable pitches of 1.0, 1.5, 2.0, 2.5 and 3.0. Images obtained at pitch 3.0 were degraded visually. Secondly a bead phantom was scanned on 1 mm width collimation at pitches of 1.0, 1.5, 2.0 and 3.0. Focus size was determined with a variable setting of small and large. We obtained maximum and full width at half maximum (FWHM) scans and performed a clinical study of small lung nodules on 1 mm width at pitch 2.0 on small focus. Thirdly, we estimated the image quality of 1 mm/2 mm helical scans in comparison with 1 mm/1 mm helical scans. Comparison was made using axial, MPR and 3D-CT (MIP) images. The axial and MIP images were almost equal qualitatively, whereas MPR images were degraded, especially in the near region of the heart and great vessels, due to motion artifacts. We emphasize the utility of 1 mm/2 mm helical CT in scanning a much larger volume with the same breath holding.

Adult↗
PubMed1997

Development and characterization of cationic liposomes conjugated with HVJ (Sendai virus): reciprocal effect of cationic lipid for in vitro and in vivo gene transfer.

Today, nonviral gene transfer vectors attract more attention as a therapeutic strategy for human diseases, because viral vectors such as adenoviral and herpes viral vectors have been proven to have problems, especially in immunogenicity and cytotoxicity. However, the main limitation of nonviral vectors has been low efficiency of gene expression. To overcome this defect, we have developed a new class of transfection vehicles, HVJ-cationic liposomes. The use of the cationic lipid DC-cholesterol facilitates efficient entrapment of negatively charged macromolecules (plasmid DNA, oligodeoxynucleotides, and proteins) and efficient interaction with negatively charged plasma membranes of cultured cells in vitro. Moreover, the fusogenic envelope proteins of hemagglutinating virus of Japan (HVJ) enhance delivery of the enclosed materials into cells. Using firefly luciferase as a marker, we optimized the liposome formula. As a result, we have succeeded in obtaining 100-800 times higher gene expression in vitro than with the conventional HVJ-anionic liposomes. However, in vivo gene transfer experiments have revealed that the use of cationic lipid instead of anionic lipid reduced the transgene expression dramatically in organs such as muscle and liver. We further discovered that the use of anionic liposomes with a viral-mimic king lipid composition increased transfection efficiency by several times in vivo. We conclude that the alternative usage of transfer vectors, for example, HVJ-anionic liposomes for in vivo delivery to extended areas of organs and HVJ-cationic liposomes for in vitro delivery (and possibly for in vivo delivery to a restricted area of organs), is of significance.

Animals↗
PubMed1997

Protection against verocytotoxin in mice induced by liposome-coupled verocytotoxin.

Purified verocytotoxins (VTs), VT1 and VT2, were coupled to liposomes via glutaraldehyde. During the coupling procedure, both VT1 and VT2 were detoxified. Intraperitoneal injection in BALB/c mice with either VT1-liposome or VT2-liposome induced a substantial amount of anti-VT1 or anti-VT2 IgG antibody production, respectively. Mice immunized with VT2-liposome were protected against intravenous challenge with a lethal dose of VT2 and the degree of protection correlated well with the amount of IgG induced against VT2. Although VT1-liposome failed to induce protection against VT1, the decrease of the body weight observed after the toxin challenge correlated inversely with the amount of anti-VT1 IgG induced, suggesting that VT1 neutralizing antibody was present in VT1-liposome-immune mice. In addition, VT-liposome conjugate induced no detectable anti-VT IgE antibody production. These results demonstrate the potential ability of VT-liposome conjugates for the production of VT vaccine which induces protection against VTs.

Animals↗
PubMed1995

[Experimental study on early detection of alloxan-induced pulmonary injury by magnetic resonance imaging].

We studied the early detection of alloxan-induced pulmonary injury by magnetic resonance imaging in vivo. Permeability edema was induced in ten rats by intravenous injection of alloxan at 100 mg/kg. T1-and T2-weighted images were acquired in five rats every 30 min for 120 min after alloxan injection. Five rats served as controls. The rats were sacrificed immediately after imaging and examined microscopically. CT images were also acquired in five rats every 30 min for 120 min after alloxan injection. Five rats served as controls. The rats were sacrificed immediately after imaging, and the wet-to-dry ratio of the lung was measured. In T1-weighted images, relative signal intensity from the lung with permeability edema rose from 30 min to 120 min, and was greater than that from normal lung every time. In T2-weighted images, there was no statistically significant difference in relative signal intensity of the lung between permeability edema and the control during 120 min. In CT images, there was also no statistically significant difference in lung density between permeability edema and the control during 120 min. There was no statistically significant difference in the wet-to-dry lung ratio between edematous lung and normal lung. In histological study, mild congestion and interstitial edema were observed in edematous lung. These results suggest the potential capability of MR imaging in detecting the early phase of permeability pulmonary edema.

Alloxan↗
PubMed1995

[Distal pulmonary vessels demonstration by magnetic resonance angiography and its usefulness in the diagnosis of peripheral nodules].

To evaluate the usefulness of MRA in the diagnosis of peripheral pulmonary nodules, 18 lung cancers and 12 old inflammatory nodules were studied employing multiplanar fast radiofrequency spoiled gradient echo sequence (SPGR) with phased array coils. For each case, 12 images in coronal and sagittal sections were obtained after administration of Gd-DTPA, and original images, maximum intensity projection (MIP) images and multiplanar reconstruction (MPR) images were used for the interpretation. In ten cases, MRA images were compared with resected specimens and we found that the minimum diameter of pulmonary vessels which could be demonstrated by our method was about 1.5mm. As the results of the interpretation of MRA images in 30 cases, venous involvement, which is one of the classical important findings indicative of malignancy, was clearly demonstrated in 15 of 18 (83%) lung cancers, whereas such involvement was detected in only 3 of 12 (25%) inflammatory nodules. Pulmonary MRA, especially MIP and MPR images could make it easy to identify involved vessels, and proved to be useful in differentiating pulmonary nodules.

Adenocarcinoma↗