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Biomedical subjects

Y Hotta

Publications and source records attributed to Y Hotta.

At least 91 records · Page 5Linked to original sources
PubMed1996

Protective effects of dimethyl amiloride against postischemic myocardial dysfunction in rabbit hearts: phosphorus 31-nuclear magnetic resonance measurements of intracellular pH and cellular energy.

The effects of 5-(N,N-dimethyl)amiloride, a potent and specific Na(+)-H+ exchange inhibitor, were investigated in isolated perfused rabbit hearts subjected to ischemia and reperfusion. Phosphorus 31-nuclear magnetic resonance spectroscopy was used to monitor intracellular pH, creatine phosphate, beta-adenosine triphosphate, and inorganic phosphate. After cardioplegic arrest with St. Thomas' Hospital solution, normothermic (37 degrees C) global ischemia was induced for 45 minutes, and the hearts were reperfused for 50 minutes. Dimethyl amiloride at 10 mumol/L, which has minimal inotropic and chronotropic effects on the nonischemic heart, was added to the cardioplegic solution. Treatment with dimethyl amiloride reduced the elevation of left ventricular end-diastolic pressure during and after the ischemia and improved the postischemic recovery of developed pressure from 76% +/- 3.2% at 30 minutes of reperfusion in control hearts (n = 6) up to 99% +/- 1.9% in hearts treated with dimethyl amiloride (n = 8). Dimethyl amiloride did not affect the decline in intracellular pH during ischemia for up to 30 minutes but enhanced the intracellular acidosis thereafter. The intracellular pH at the end of ischemia was 6.21 +/- 0.05 in control hearts compared with 5.24 +/- 0.17 in hearts treated with dimethyl amiloride (p < 0.05). During reperfusion, intracellular pH of hearts treated with dimethyl amiloride was less than control for 5 minutes, but subsequent recovery of intracellular pH was similar to control. Treatment with dimethyl amiloride did not affect creatine phosphate breakdown, inorganic phosphate accumulation, and beta-adenosine triphosphate depletion during 45 minutes of ischemia. The creatine phosphate resynthesis and inorganic phosphate reduction during reperfusion were also unaffected. These findings suggest that Na(+)-H+ exchange plays an important role not only during reperfusion but also during ischemia for the development of postischemic cardiac dysfunction most likely by inducing primary Na+ and secondary Ca2+ overload. Specific Na(+)-H+ exchange inhibitors like dimethyl amiloride would have a potential therapeutic profile in cardiac surgery, especially if added before ischemia.

Acidosis↗
PubMed1996

A single Drosophila melanogaster myosin light chain kinase gene produces multiple isoforms whose activities are differently regulated.

BACKGROUND: Myosin light chain kinase (MLCK) specifically phosphorylates the myosin regulatory light chain in a calcium/calmodulin (Ca2+/CaM)-dependent manner in animal cells. The roles of MLCK are not fully understood, particularly in nonmuscle cells. Therefore, we cloned and characterized a Drosophila MLCK gene as the first step in a genetic analysis of this process. RESULTS: Four transcripts are produced from this gene. These transcripts encode at least three isoforms (isoform-I, -II and -III), which share a kinase domain, a fibronectin type III motif and an immunoglobulin C2 motif. However, regulatory regions differ between isoform-I/II and -III due to the alternative splicing of the exon encoding a CaM-binding domain. As a result, isoform-I and -II are Ca2+/CaM-dependent forms, whereas isoform-III is a Ca2+/CaM-independent form. Northern blotting and in situ hybridization showed that the expressions of these isoforms are distinctly regulated in stage- and tissue-dependent manners. Isoform-I seems to be expressed ubiquitously, while isoform-III is expressed predominantly in muscle tissues. In contrast to these isoforms, isoform-II is specific to late pupa and adult. CONCLUSION: In Drosophila, a single MLCK gene produces multiple isoforms whose regulatory regions and expression patterns are different. These differences suggest various cellular functions of MLCK in Drosophila.

Amino Acid Sequence↗
PubMed1996

Analysis of phosducin as a candidate gene for retinopathies.

Phosducin, a retina-expressed gene mapped to chromosome 1q25-32.1, was analyzed as a candidate gene for retinopathies. The phosducin gene was cloned and characterized, and PCR primers were designed. Eighty-three patients with various retinopathies and 45 control subjects (24 American, 21 Japanese) were analyzed for mutations in the phosducin gene by PCR, denaturing gradient gel electrophoresis (DGGE), and sequencing. A heterozygous sequence variant changing a glycine to arginine at codon 178 was found in one Usher syndrome type II (USH2) patient, while the other USH2 patients did not show any coding sequence variant. A heterozygous sequence variant changing an asparagine to lysine at codon 174 was found in a patient with a severe retinal degeneration in the category of diseases known as acute zonal occult outer retinopathy (AZOOR). Three non-coding sequence variants were found. Two of these were always present together and found in 20.8% of American and 2.4% of Japanese control subjects, reflecting a difference in population pools. In conclusion, the phosducin gene did not show mutations consistent with it being the causative gene for USH2, but its possible pathogenicity in AZOOR or other retinopathies remains an open question which may be answered by further analysis.

Base Sequence↗
PubMed1996

Cloning and high expression of rabbit FKBP25 in cornea.

Rabbit 25 kDa FK-506/rapamycin binding protein (FKBP25) cDNA was isolated during a screening of rabbit corneal endothelial cDNA library. The cDNA clone was a 955 bp fragment with a 670 bp open reading frame and a 285 bp 3' non-coding region. The cDNA and deduced amino acid sequences were 91.7 and 96.4% identical, respectively, to previously reported human FKBP25B sequences. The lysine-rich region KK(X)7KK(X)26KKKK, which plays an important role in nuclear translocation in rabbit FKBP25, was recognized. Reverse transcription-polymerase chain reaction (RT-PCR) amplification of rabbit FKBP25 and FKBP12 in selected rabbit organs, including total cornea, showed higher expressions in cornea, retina, cerebrum and cerebellum than other compared tissues. RT-PCR amplification in rabbit cultured corneal cells showed expressions in all cultured corneal epithelial, stromal, endothelial, and especially in SV40-adenovirus vector-immortalized epithelial cells. The higher expressions of FKBP25 and FKBP12 suggest significant roles for FKBPs in those tissues.

Amino Acid Sequence↗
PubMed1996

Recovery of alveolar bone by the guided bone regeneration technique.

Bone defects associated with the removal of endosteal implants often create challenging problems for clinicians. The guided bone regeneration (GBR) technique has been recently used in the treatment of such defects, and promising results have been obtained. This report describes the use of Gore-Tex tissue augmentation material (GTAM) membranes after the removal of Bioceram screw implants in two patients who wore the implants for 10 years and seven years, respectively. After removal, the residual bone defects were treated by the GBR technique. Bone regeneration was confirmed in the defects around the newly placed implants, and decreased bone mass was avoided in both patients. In case 1, a free gingival graft was used to increase the area of attached gingiva around the implants, since the status of peri-implant soft tissue is also crucial to the outcome of therapy. This technique was very effective in maintaining an adequate width of attached gingiva.

Adult↗
PubMed1995

glial cells missing: a binary switch between neuronal and glial determination in Drosophila.

In the Drosophila CNS, both neurons and glial are derived from neuroblasts. We have identified a gene, glial cells missing (gcm), that encodes a novel nuclear protein expressed transiently in early glial cells. Its mutation causes presumptive glial cells to differentiate into neurons, whereas its ectopic expression forces virtually all CNS cells to become glial cells. Thus, gcm functions as a binary switch that turns on glial fate while inhibiting default neuronal fate of the neuroblasts and their progeny. Similar results are also obtained in the PNS. Analyses of the mutant revealed that "pioneer neurons" can find correct pathways without glial cells and that neurons and glia have a common molecular basis for individual identity.

Amino Acid Sequence↗
PubMed1995

Expression profiles of a human gene identified as a structural homologue of meiosis-specific recA-like genes.

Using the cDNA clone from mouse testis which carries the conserved sequences among meiosis-specific recA-like genes, a highly homologous cDNA clone was isolated from a cDNA library of human testis. The clone had the coding capacity of a protein consisting of 340 amino acid residues, which coincides with the average size of putative eukaryotic recA-like proteins. When expression of the corresponding gene, named HsLIM15, in various tissues was examined by reverse transcription-PCR, products of two different sizes were detected in testis: While the longer was predominantly seen in the testis, the shorter was commonly to all the tissues including the testis. Analysis of the sequences indicated that the longer product corresponded to the above cDNA clone, and the shorter one was its deletion product missing an internal 165 bp portion. The result strongly suggests that the mRNA species coding for the putative meiosis-specific RecA-like protein in human is predominantly expressed in testis possibly as an alternative splicing product of a ubiquitously expressed gene.

Adenosine Triphosphatases↗
PubMed1995

Different effects of isoproterenol and dihydroouabain on cardiac Ca2+ transients.

Cytosolic fura-2 Ca2+ transient signals (TCa) and the left ventricular pressure or contraction of myocardium under the positive inotropic effects of the beta-adrenoceptor agonist, isoproterenol, and the cardiac glycoside, dihydroouabain, were measured simultaneously and the results were compared. TCa was observed preceding the onset of force development and showed a steeper rise and slower decay than did the contraction curve of papillary muscle. Isoproterenol increased the steepness and the amplitude of TCa, reflecting the speed and peak force of contraction, and clearly biphasic TCa were observed with biphasic contractions developed at low frequency. Ryanodine reduced not only the early component of the contraction but also TCa, without affecting the diastolic Ca2+ level. These effects of isoproterenol were attributed to the enhanced uptake of Ca2+ by the sarcoplasmic reticulum. In contrast, dihydroouabain elevated the Ca2+ level at diastole without any change in the amplitude of TCa, suggesting that dihydroouabain inhibits the membrane Na pump thereby increasing the intracellular Ca2+ via Na(+)-Ca2+ exchange. Furthermore, a comparison of the time course of the isometric twitch curve with that of TCa in rested state contraction indicated that there are distinct differences between the mechanisms of the positive inotropic effects of isoproterenol and of dihydroouabain.

Adrenergic beta-Agonists↗
PubMed1995

Characterization of a mouse recA-like gene specifically expressed in testis.

Using the sequence information on a class of recA-like genes, which have been isolated from meiosis-specific cDNA libraries of Lilium and budding yeast and identified in various plant cells, a new gene was screened from a mouse testis cDNA library. The putative product of the gene, termed MmLim15, was 37.8 kDa consisting of 340 amino acid residues. The predicted amino acid sequence showed 72-78% similarity to those of Lim15 from Lilium and other meiosis-specific RecA-like proteins. Similarity was also found to another class of recA-like proteins, Rad51 and its homologues, which have been detected in both budding and fission yeasts and various animal cells, but to a lesser extent and the similarity patterns were somewhat different from those for Lim15 type proteins, revealing that the two classes of recA-like genes are phylogenetically separate. Reverse transcription-PCR using poly(A)+ RNAs from various tissues as templates indicated that transcription of MmLim15 occurred specifically in the testis, suggesting that the MmLim15 product participates in meiotic recombination.

Adenosine Triphosphatases↗
PubMed1995

Localization of RecA-like recombination proteins on chromosomes of the lily at various meiotic stages.

The Rad51 and Lim15 proteins of lily, which are homologs of the bacterial RecA protein, were found on chromosomes in various stages of meiotic prophase 1. The presence of both Rad51 and Lim15 proteins as discrete foci on leptotene and zygotene chromosomes and their colocalization suggest that meiotic recombination begins at the leptotene stage with the cooperation of these proteins and continues in zygotene. Localization of the foci on or adjacent to the chromosomes suggests that these proteins bind to the chromatin loops that extend from the axial cores. The proteins in these foci may participate in the searching and pairing of homologous DNA sequences, as the RecA protein does. The different pattern of localization of the Rad51 protein between the leptotene and pachytene stages and the absence of the Lim15 protein in the pachytene stage suggest that the Rad51 protein plays different roles in these stages.

Amino Acid Sequence↗
PubMed1995

Fasciclin III as a synaptic target recognition molecule in Drosophila.

Fasciclin III, a cell adhesion molecule of the immunoglobulin superfamily, is expressed by motor neuron RP3 and its synaptic targets (muscle cells 6 and 7) during embryonic neuromuscular development of Drosophila. We report here that RP3 often incorrectly innervates neighbouring non-target muscle cells when these cells misexpress fasciclin III, but still innervates normal targets in the fasciclin III null mutant. Fasciclin III manipulations do not influence target selections by other motor neurons, including fasciclin III-expressing RP1. We propose that fasciclin III acts as a synaptic target recognition molecule for motor neuron RP3, and also that its absence can be compensated for by other molecule(s).

Animals↗
PubMed1995

Molecular heterogeneity among primary motoneurons and within myotomes revealed by the differential mRNA expression of novel islet-1 homologs in embryonic zebrafish.

Zebrafish embryos have three or four identifiable primary motoneurons per hemisegment. We previously reported that, while several ventral cells initially express the zebrafish Islet-1 (Isl-1) gene, a member of the LIM/homeobox gene family, the expression of this gene becomes restricted to a single or a pair of cells slightly anterior to each segment border by 16 hr after fertilization. Double staining by in situ hybridization and immunohistochemistry strongly suggested that these cells were mainly rostral primary motoneurons. Here, we have isolated two novel zebrafish cDNA clones for more Isl-1 family genes, termed zfIsl-2 and zfIsl-3. zfIsl-2 mRNA starts to be expressed in the ventral midsegmental cells per hemisegment around 15 hr. Double labeling experiments have shown that these midsegmental cells are the caudal primary motoneuron (CaP) and its variant equivalence pair. Our results revealed the heterogeneity in the expressed genes among primary motoneurons before the fates of the primary motoneurons are irreversibly determined, and further suggest the involvement of the Isl-1 and zfIsl-2 genes in the determination of cellular identities by primary motoneurons in embryonic zebrafish. zfIsl-3 mRNA is not expressed in motoneurons but is expressed at 17 hr, mainly in the ventral myotomes. This suggests that zfIsl-3 may be involved in the regional specification of the myotome and also in target recognition by CaP. zfIsl-2 is also expressed throughout the developing eye and tectal region of the midbrain, the target for the retinal axons. In the ventral spinal cord of the spadetail mutant embryo, which has defects in the somites, the cells expressing zfIsl-2 mRNA significantly decreased in number in contrast to the increase in cells expressing Isl-1 mRNA, suggesting the influence of the somites on the expression of both genes.

Amino Acid Sequence↗
PubMed1995

Missense mutation of rhodopsin gene codon 15 found in Japanese autosomal dominant retinitis pigmentosa.

Heterozygous missense mutation in codon 15 of the rhodopsin gene was detected in a patient with autosomal dominant retinitis pigmentosa (ADRP), where a transition of adenine to guanine at the second nucleotide in codon 15 (AAT-->AGT), corresponding to a substitution of serine residue for asparagine residue (Asn-15-Ser) was detected. None of the remaining unrelated 42 ADRP, 24 autosomal recessive RP (ARRP) and 34 normal individuals had this alteration. Her funduscopic findings were sectorial in type similar to that of the patients with the same mutation found in an Australian pedigree (Sullivan et al., 1993). This study shows phenotypic similarities in patients with the same mutation of a different ancestry.

Amino Acid Sequence↗
PubMed1995

Meiosis specific transcription and functional proteins.

We have discussed and/or demonstrated the following: 1. Many enzymes and structural proteins have been identified as meiosis-specific proteins. These can be classified according to their metabolic behavior. 2. We obtained and analyzed 18 cDNA clones from lily meiocytes. One of them, LIM15, was similar to known genes like RecA, RAD57, and DMC1/ISC2, and might function in pairing and recombination. 3. Transcription of these genes is regulated by their regulator region(s). When such a regulator, mei2 promoter sequence isolated from S. pombe, was ligated with the proper vector and transfected, it functioned specifically in meiotic cells but not in the somatic cells tested. 4. Presence of a new lamin, lamin B3 was identified in mammalian spermatocytes and the transfection of lamin B3 gene (inserted into vector) into somatic cells alters the nuclear shape, possibly expressing a characteristic shape of meiotic nuclei. Lamin B3 was synthesized after meiosis-specific processing of lamin B2 mRNA. Other protein specific to meiotic nuclear-skeleton (MNS1) were found and characterized. All these events were studied basically focussing on homologous pairing and recombination which take place in meiosis I. We recognize the necessity of further studies on these and other events like the structure and segregation of chromosomes and the suppression of somatic gene expression during meiosis.

Amino Acid Sequence↗
PubMed1995

Structural analysis of a recA-like gene in the genome of Arabidopsis thaliana.

A recA-like gene was identified in the genome of Arabidopsis thaliana by means of PCR using primers designed on the basis of previously reported amino acid sequences of eukaryotic RecA-like proteins. The structure of the gene, termed ArLIM15, was investigated by comparing the primary structure of the genomic DNA with that of the corresponding cDNA. The open reading frame, which was split into 15 exons, was established to have the capacity for encoding a 37.3-kDa polypeptide. The amino acid sequence of the putative product of ArLIM15 showed a high degree of similarity to that of LIM15 in the monocotyledonous plant Lilium, including a 93% identity, and to those of other recA-like genes in yeasts and vertebrates with identities of 69-71%. Phylogenetic analysis indicated ArLIM15 to be much closer to meiosis-specific LIM15 and DMC1 in Saccharomyces cerevisiae than to RAD51 in S. cerevisiae and its homologues on an evolutionary scale.

Amino Acid Sequence↗