Biomedical subjects

Hisatake Kondo

Publications and source records attributed to Hisatake Kondo.

52 records · Page 3Linked to original sources

PubMed2003

Specific localization of epidermal-type fatty acid binding protein in dendritic cells of splenic white pulp.

Dendritic cells in the splenic white pulp of mice were intensely immunoreactive for epidermal-type fatty acid binding protein (E-FABP). This specific immunostaining revealed a clear difference in morphology between the dendritic cells in the periarterial lymphoid sheath (PALS) and follicular dendritic cells in the follicles in terms of cell sizes and process branching. No immunoreactivity was detected in dendritic cells in the marginal zones and the red pulp, although endothelial cells of almost all capillaries in the red pulp were immunoreactive for E-FABP. After peritoneal injection of lipopolysaccharide, the immunoreactive cells in PALS progressively enlarged and became rounded in shape with a peak in size at 24 h postinjection and they eventually resumed the dendritic form at 48 h postinjection. Within each of the enlarged immunoreactive cell perikarya were included small immunonegative apoptotic cells, presumptive lymphocytes. Taken together, E-FABP is useful as a marker for dendritic cells in the splenic white pulp, and may be involved through combination with fatty acids in antigen presentation and retention as well as in cytokine production.

Animals↗

PubMed2003

Coexpression of junctophilin type 3 and type 4 in brain.

Recent studies indicated that junctophilins (JPs) contribute to the formation of junctional membrane structures in excitable cells by interacting with the plasma membrane and spanning the endoplasmic/sarcoplasmic reticulum (ER/SR) membrane. In the brain, functional crosstalk between cell-surface and intracellular channels is proposed in the "subsurface cistern" as the junctional membrane complex observed in neurons. So far, three JPs have been identified as tissue-specific subtypes derived from different genes; JP-1 is specifically expressed in skeletal muscle, JP-2 is detected throughout muscle cell types, and JP-3 is predominantly expressed in the brain. In this paper, we report a novel JP subtype, JP-4, encoded in the human (chromosome 14q11.1) and mouse (chromosome 14C1-2) genomes. Cloning the cDNA showed that JP-4 shares characteristic structural features with other JP subtypes, and Northern and Western blot analyses demonstrated its brain-specific expression. In situ hybridization analysis revealed that both JP-3 and JP-4 mRNAs are expressed in discrete neuronal sites, and their overall regional distribution patterns were similar in the brain. Furthermore, both the JP mRNAs and subsurface cistern showed somatodendritic localization in hippocampal pyramidal neurons. The results obtained suggest the collaborative contribution of JP-3 and JP-4 to the subsurface cistern formation in neurons.

Animals↗

PubMed2003

Structural and functional characterization of mouse glutamate decarboxylase 67 gene promoter.

Neuronal expression of the mouse glutamate decarboxylase 67 (mGAD67) gene occurs exclusively in neurons that synthesize and release GABA (GABAergic neurons). This gene is also expressed in pancreatic islet cells and testicular spermatocytes. In order to elucidate the molecular mechanisms underlying the regulation of mGAD67 gene expression, we isolated and characterized the 5'-flanking region of this gene. Sequence analysis of a 10.2-kb DNA fragment of this gene containing a promoter region (8.4 kb) and noncoding exons 0A and 0B revealed the presence of numerous potential neuron-specific cis-regulatory elements. Functional analysis of the 5'-flanking region of exons 0A and 0B by transient transfection into cultured cells revealed that the region -98 to -52 close to exon 0A is important for the transcriptional activity of both exons 0A and 0B. In addition, we used transgenic mice to examine the expression pattern conferred by the 10.2 kb DNA fragment of the mGAD67 gene fused to the bacterial lacZ reporter gene. Transgene expression was observed in neurons of particular brain regions containing abundant GABAergic neurons such as the basal ganglia, in pancreatic islet cells and in testicular spermatocytes and spermatogonia. These results suggest that the 10.2 kb DNA fragment of the mGAD67 gene contains regulatory elements essential for its targeted expression in GABAergic neurons, islet cells and spermatocytes.

5' Flanking Region↗

PubMed2003

Localization of brain-type fatty acid-binding protein in Kupffer cells of mice and its transient decrease in response to lipopolysaccharide.

Brain-type fatty acid-binding protein (B-FABP) was localized in Kupffer cells of liver of postnatal day 10 (P10) and older mice in immunolight and electron microscopy as well as by in situ hybridization histochemistry. The immunoreaction products were localized in the cytoplasmic matrix but not within the nucleus. After peritoneal injection of lipopolysaccharide (LPS), the immunoreaction for B-FABP decreased markedly in Kupffer cells at 1 h postinjection and thereafter gradually recovered to the preinjection level by 24 h postinjection, although no decrease in the mRNA expression was detected in Northern blotting throughout the course after the injection. The specific localization of B-FABP, but not the other FABPs, in Kupffer cells, and its rapid decrease after LPS injection suggest the intimate involvement of B-FABP in Kupffer cells in the inflammatory reaction, probably through mediation of n-3 polyunsaturated fatty acids, which are strong binders of B-FABP.

Animals↗

PubMed2003

Cytoplasmic matrix in embedment-free electron microscopy: non-molecular biological histology.

Embedment-free electron microscopy using polyethylene glycol as a transient embedment has revealed that slender strands, originally termed microtrabeculae and microtrabecular lattices, interconnect every organelle and conventional cytoskeletons as well as plasma membranes, resulting in the formation of 3-D meshworks in all portions of the cytoplasmic matrix of every cell. The microtrabeculae correspond well to the wispy components in the cytoplasmic matrix of conventionally epoxy-sectioned cell specimens that have been looked at but often neglected because of their poorly defined images due to the presence of embedding media having a substantial electron-scattering property. Because of the occurrence of similar meshworks in specimens that are supposed to be unstructured, such as the intramitochondrial matrix and blood plasma, together with the failure to detect any predictable changes of the microtrabecular lattices by experimental manipulation of cellular environments, it is inaccurate to conclude that all microtrabecular lattice represent structures equivalent to those in a living state of cells simply because of their clear appearance. Instead, three possible interpretations are newly proposed for the biological significance of the microtrabecular lattices. The first is that the appearance of lattices represents the presence of proteins, and that their approximate concentrations are speculated based on the compactness of the lattice. The second is that when an intracellular microdomain composed of more compact lattices is contiguous with another domain composed of looser lattices in a given cell, the former might represent the gelated state and the latter the solated state. Possible examples for these two interpretations are also proposed, possibly leading us to further elaborate the significance of microtrabecular lattices.

Animals↗

PubMed2003

Nuclear localization of diacylglycerol kinase zeta in neurons.

Diacylglycerol kinase (DGK) is involved in intracellular signal transduction as a regulator of levels of diacylglycerol which leads to protein kinase C activation. Previous studies have revealed that DGK consists of a family of isozymes in mammalian species and that most if not all of them show abundant expression in the central nervous system, suggesting the importance of this enzyme in neuronal function. Among the isozymes, DGK zeta (previously also known as DGK-IV for the rat clone) has unique structural features, such as four ankyrin-like repeats and a nuclear localization signal (NLS), and shows intense mRNA expression in neurons of the olfactory bulb, hippocampus and cerebral and cerebellar cortices (Goto, K. & Kondo, H. (1996), Proc. Natl Acad. Sci. USA, 93, 11196-11201). However, previous studies have given conflicting results about whether or not DGK zeta localizes to the nucleus in these cells. In this study, we have used immunohistochemistry with specific antibodies in brain tissues and cDNA transfection into primary cultured neurons to address this question. We have shown that, while DGK zeta is primarily a nuclear protein in neurons, it can also be cytoplasmic in some conditions, and the subcellular location depends not only on the cell type but also on the developmental state or growth conditions of the cell. In addition, we have used deletion mutants to show that nuclear transport of DGK zeta depends on a cooperative interaction between the NLS and the C-terminal region including ankyrin repeats in a manner which suggests that the NLS is a cryptic site whose exposure is regulated by the C-terminal region. Together, these results support the hypothesis that the localization of DGK zeta may be regulated by differential expression of these various proteins which interact with its C-terminal region.

Animals↗

PubMed2003

Cloning and characterization of a novel variant (mM-rdgBbeta1) of mouse M-rdgBs, mammalian homologs of Drosophila retinal degeneration B gene proteins, and its mRNA localization in mouse brain in comparison with other M-rdgBs.

We report the cloning, characterization and localization in the brain of a novel isoform termed mM-rdgBbeta1 (mouse type of mammalian retinal degeneration Bbeta1 protein) in comparison with the localization of three known mammalian homologs (M-rdgBbeta, M-rdgB1, M-rdgB2). mM-rdgBbeta1 cDNA contains a sequence of 119 bp as a form of insertion in the open reading frame of the known mM-rdgBbeta, and encodes a protein of 269 amino acids with a calculated molecular mass of 31.7 kDa, different from the molecular mass of 38.3 kDa of mM-rdgBbeta. It also contains a phosphatidylinositol transfer protein (PITP)-like domain similar to the known three homologs, as well as D-rdgB. The recombinant mM-rdgBbeta1 protein shows the specific binding activity to phosphatidylinositol but not to other phospholipids. This novel molecule is localized not only in the cytoplasm but also in the nucleus, different from the cytoplasmic localization of mM-rdgBbeta. In in situ hybridization analysis, the gene expression for mM-rdgBbeta1 in the brain, though weak, is rather confined to the embryonic stage, different from wider expression of mM-rdgBbeta in the gray matters of pre- and post-natal brains. Taken together, mM-rdgBbeta1 is suggested to play a role in the phosphoinositide-mediated signaling in the neural development.

Amino Acid Sequence↗

PubMed2003

Cloning, characterization and expression of two alternatively splicing isoforms of Ca2+/calmodulin-dependent protein kinase I gamma in the rat brain.

Ca2+/calmodulin-dependent protein kinase I (CaMKI), originally identified as a protein kinase phosphorylating synapsin I, has been shown to constitute a family of closely related isoforms (alpha, beta and gamma). Here, we have isolated and determined the complete primary structures of two alternatively splicing isoforms of CaMKI termed CaMKI gamma 1 and -gamma 2. CaMKI gamma 1 and -gamma 2 contain an identical N-terminal catalytic domain with different C-terminal regions due to the deletion of the 425-bp nucleotide sequence of CaMKI gamma 1 in CaMKI gamma 2. In vitro kinase assay has demonstrated the marked enhancement of the Ca2+/CaM-dependent activity of CaMKI gamma 1 by the preincubation with Ca2+/calmodulin-dependent protein kinase kinase (CaMKK), but no significant activation of CaMKI gamma 2. Northern blot analysis has demonstrated the predominant expression of CaMKI gamma in the brain. RT-PCR analysis has revealed similar expression patterns between CaMKI gamma 1 and CaMKI gamma 2 in various brain regions. In situ hybridization analysis has demonstrated that CaMKI gamma mRNA is expressed in a distinct pattern from other isoforms of CaMKI with predominant expression in some restricted brain regions such as the olfactory bulb, hippocampal pyramidal cell layer of CA3, central amygdaloid nuclei, ventromedial hypothalamic nucleus and pineal gland. In the primary hippocampal neurons and NG108-15 cells, transfected CaMKI gamma 1 and -gamma 2 are localized primarily in the cytoplasm and neurites but not in the nucleus. These findings suggest that both isoforms of CaMKI gamma may be involved in Ca2+ signal transduction in the cytoplasmic compartment of certain neuronal population.

Alternative Splicing↗

PubMed2002

Expression of V-1, a novel catecholamine biosynthesis regulatory protein, is enhanced by hypertension in atrial myocytes of Dahl salt-sensitive rats.

V-1 positively controls catecholamine synthetic gene transcription to promote catecholamine production in PC12D cells. In this study, immunohistochemical analysis revealed that in Wistar rats, V-1 immunoreactivity was localized not only in sympathetic axons but also in the cytoplasm of cardiomyocytes, and that the immunoreactivity in atrial myocytes was more intense than that in ventricular myocytes. Western blot analysis also showed that V-1 expression level in the atrium was higher than that in the ventricle of Wistar rat hearts. When Dahl salt-sensitive (DS) rats were fed an 8% NaCl diet after the age of 6 weeks, blood pressure was raised 230mm Hg at 18 weeks. V-1 expression was shown to be increased in the atrial myocytes of these DS rats, but not in the sympathetic axons, when assayed by immunohistochemistry. These results suggest that in normotensive rats, V-1 is preferentially expressed in the cytoplasm of cardiomyocytes in the atrium rather than in the ventricle. It is also suggested that V-1 expression is increased by hypertension in DS rat atrium.

Animals↗

PubMed2002

Are there gap junctions between chief (glomus, type I) cells in the carotid body chemoreceptor? A review.

Since the dye- and electronic couplings between the carotid body chief cells have been demonstrated, the detection and localization of the gap junctions in the carotid body is crucial to understanding the functional mechanism of chemoreception. However, conventional electron microscopy has been unsuccessful in unquestionably detecting ultrastructural features equivalent to the gap junctions, such as close (2 nm in width) membrane appositions in ultrathin sections and aggregations of intramembranous particles in freeze-fracture replicas of the carotid body. We previously reported using a modified electron microscopic study by chemically fixed and subsequent rapid freezing and freeze-substitution method a number of close membrane appositions comparable to the gap junctions. However, we later found that the freeze-substitution also induces numerous close apposition of the membrane in sites where the gap junctions are not known to occur, indicating that the modified electron microscopy by freeze-substitution is not always confirmative in the detection of the gap junction. With regard to the molecular evidence for the gap junction in the carotid body, there have so far been few data on the immunohistochemical demonstration on connexin 32 and 43 in cultured chief cells, but not in the in situ cells.

Animals↗

PubMed2002

Transient up-regulation of elongation factor-2 kinase (Ca2+/calmodulin-dependent protein kinase III) messenger RNA in developing mouse brain.

Elongation factor-2 kinase (eEF-2K) is a Ca(2+)/calmodulin-dependent protein kinase that is highly specific for eukaryotic elongation factor-2. In the present study, we examined the developmental expression of eEF-2K messenger RNA (mRNA) in the mouse brain. At E13, the expression of eEF-2K mRNA is already evident in both ventricular and mantle zones throughout the neuroaxis with a rostrocaudal increasing gradient. During late embryonic and early postnatal stages, eEF-2K mRNA is transiently up-regulated in the cerebral cortex and hippocampal pyramidal cell layer. After birth, the expression of eEF-2K mRNA gradually decreased throughout the brain. In the mature brain, eEF-2K mRNA is expressed moderately only in the cerebellar and dentate granule cell layers, and weakly in the cerebral cortex, hippocampal pyramidal cell layer and olfactory bulb. These findings suggest that eEF-2K may be involved in early neuronal events such as proliferation, differentiation, and establishment of synaptic connections.

Animals↗

PubMed2002

Localization of mRNAs for subfamily of guanine nucleotide-exchange proteins (GEP) for ARFs (ADP-ribosylation factors) in the brain of developing and mature rats under normal and postaxotomy conditions.

ADP-ribosylation factors (ARFs) play important roles in vesicular trafficking and cytoskeletal regulation and its activation depends on guanine nucleotide-exchange proteins (GEPs). By way of in situ hybridization histochemistry, the localization of mRNAs for subfamily members of low-molecular-weight ARF-GEPs in the rat brain was studied at embryonic and postnatal stages. In the embryonic brain, the gene expression for msec7-1 was distinct in the ventricular zone while that for msec7-1, -3 and EFA6 in the mantle zone. In early postnatal brain, the expression for msec7-1, -2, -3 and EFA6 was seen widely in various loci of the gray matter with different intensity, and the expression of msec7-1 and -2 mRNAs was evident in the cerebellar external granule cell layer. In the adult brain, the gene expression for the four ARF-GEPs decreased more or less in most gray matter and the distinct expression was maintained mainly in the hippocampal and dentate neuronal layers and cerebellar cortex. The expression of EFA6 mRNA was also evident in the molecular layer of the hippocampus and dentate gyrus. No obvious gene expression for cytohesin-4 and ARF-GEP100 was detected in the brain at any stages of development. The present findings suggest that ARF-GEPs are differentially involved in some processes essential to neuronal differentiation and maturation in association with ARFs.

ADP-Ribosylation Factors↗

PubMed2002

Localization of mRNAs for phosphatidylinositol phosphate kinases in the mouse brain during development.

The gene expression for seven phosphatidylinositol phosphate kinases (PIPKs)-types Ialpha, Ibeta, Igamma, types IIalpha, IIbeta, IIgamma, and type III-was examined using in situ hybridization histochemistry, in the mouse brain during normal development. In the embryonic mouse brain, positive expression signals were detected only for the genes encoding PIPK Igamma and PIPK IIbeta in both the cerebral ventricular and mantle zones, with weaker signals in the former zone. On the other hand, the genes encoding all PIPKs were essentially detected in the external granule cell layer which represents the germinal zone for the neuronal granule cells. In the postnatal brain, among the seven PIPKs, the expression for genes encoding PIPK Igamma and IIbeta is evident in most gray matter, while the expression for the other five types was weak in the cortical gray matter and negligible in most non-cortical gray matter such as the diencephalon and brain stem nuclei. While the expression for most PIPKs in the mature hippocampus was distinct, the expression in the CA3 and the dentate gyrus was less definite for the genes encoding PIPK Ialpha and IIgamma, respectively. The distinct expression for the gene encoding PIPK IIalpha was detected in the postnatal white matter such as the cerebellar medulla, the corpus callosum, the hippocampal fimbriae, and the internal capsule.

Animals↗

PubMed2002

Altered water barrier function in epidermal-type fatty acid binding protein-deficient mice.

We have generated mutant mice for epidermal-type fatty acid binding protein by the gene targeting technique and examined the phenotype in detail. Despite a lack in the expression of epidermal-type fatty acid binding protein mRNA and its protein in the skin and other tissues of the mutant mice, the animals appeared normal in gross and histologic examination. Northern blot analysis of other fatty acid binding proteins revealed a distinct elevated gene expression of heart-type fatty acid binding protein in the skin of the homozygous mice. In analyses of the skin, no differences were observed in contents of major fatty acids, electron microscopic appearance as well as inflammatory responses in ear skin between the mutant and wild-type mice. Basal transepidermal water loss of homozygous mice was lower than that of the wild mice. When acetone was applied to the skin for disruption of the water permeability barrier, recovery in transepidermal water loss was delayed, although maximum transepidermal water loss upon acetone treatment was similar between homozygous and wild-type mice in terms of size and time course. The molecular mechanism by which epidermal-type fatty acid binding protein contributes to the water barrier function of the skin remains to be elucidated.

Animals↗

PubMed2002

Analysis on the phenotype of E-FABP-gene knockout mice.

The fatty acids are shown to be critical in the maintenance of the water permeability barrier that is ascribed to the lipids in the intracellular milieu of the cornified cell layer in the epidermis. In view of this importance in the skin, we examined the phenotype of epidermal fatty acid binding protein (E-FABP)-deficient mice. In spite of total lack of E-FABP expression in the various tissues of E-FABP deficient mice, these animals appeared normal in gross and histological examination. In Northern blot analysis for other FABPs, the gene expression of heart (H-)-type FABP is specifically elevated in the liver of neonatal heterozygous and homozygous mice, suggesting the functional compensation of H-FABP for E-FABP deficiency during their development. In functional analyses of the skin, the basal transepidermal water loss (TEWL) of the adult homozygous mice showed lower levels compared with the wild-type mice, and the impairment of recovery in TEWL was observed in the homozygous mice when the lipid barrier of the skin was disrupted by acetone. These results demonstrate that E-FABP is responsible for the water permeability barrier of the skin, although the molecular mechanism remains to be further elucidated.

Animals↗

PubMed2001

Localization of epidermal-type fatty acid binding protein in the thymic epithelial cells of mice.

The immunoreactivity for epidermal-type fatty acid binding protein of epidermis type (E-FABP) was selectively localized in the epithelial cells of both cortex and medulla of mouse thymus. The cortical epithelial cytoreticulum was clearly visible with the intense immunoreactivity and the immunoreactive cytoreticulum extended intricately throughout the thymic cortex to enclose thymocytes. In the thymic medulla, the immunoreactivity was variable in intensity among the epithelial cells and there was a tendency that epithelial cells containing more numerous tonofilament bundles were less immunoreactive. Considering the possibility that FABPs function as intracellular carriers for unsaturated long chain fatty acids, the present finding suggests that E-FABP in the thymic epithelial cells, especially the cortical ones because of their extensive location, are intimately involved in the metabolic processes of fatty acids including production of bioactive substances, such as prostaglandin and leukotriene, which are known to exert some regulation of thymic immune responses.

Animals↗