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N Kondo

Publications and source records attributed to N Kondo.

At least 19 recordsLinked to original sources
PubMed2001

Various cells retrovirally transduced with N-acetylgalactosoamine-6-sulfate sulfatase correct Morquio skin fibroblasts in vitro.

Gene therapy may provide a long-term approach to the treatment of mucopolysaccharidoses. As a first step toward the development of an effective gene therapy for mucopolysaccharidosis type IVA (Morquio syndrome), a recombinant retroviral vector, LGSN, derived from the LXSN vector, containing a full-length human wildtype N-acetylgalactosamine-6-sulfate sulfatase (GALNS) cDNA, was produced. Severe Morquio and normal donor fibroblasts were transduced by LGSN. GALNS activity in both Morquio and normal transduced cells was several fold higher than normal values. To measure the variability of GALNS expression among different transduced cells, we transduced normal and Morquio lymphoblastoid B cells and PBLs, human keratinocytes, murine myoblasts C2C12, and rabbit synoviocytes HIG-82 with LGSN. In all cases, an increase of GALNS activity after transduction was measured. In Morquio cells co-cultivated with enzyme-deficient transduced cells, we demonstrated enzyme uptake and persistence of GALNS activity above normal levels for up to 6 days. The uptake was mannose-6-phosphate dependent. Furthermore, we achieved clear evidence that LGSN transduction of Morquio fibroblasts led to correction of the metabolic defect. These results provide the first evidence that GALNS may be delivered either locally or systematically by various cells in an ex vivo gene therapy of MPS IVA.

Animals↗
PubMed2001

Temperature-sensitive phenotype of Chinese hamster ovary cells defective in PEX5 gene.

SK32 mutant cells, which were isolated as peroxisome-deficient Chinese hamster ovary (CHO) cells by an advantage of a visible peroxisome form of green fluorescent protein (GFP), were found to suffer from a functional loss of PEX5 gene encoding for PTS1R. The sequence analysis of cDNA indicated that PEX5 gene encoded for the two isoforms composed of 603 amino acids (PTS1RS) and 640 amino acids (PTS1RL). The mutation changed glycine to arginine at amino acid position 343 of PTS1RL (corresponding to the position 306 of PTS1RS) in SK32 cells. The mutant cells exhibited a temperature-sensitive (TS) phenotype on the peroxisomal localizations of the recombinant GFP and urate oxidase appending a genuine peroxisome targeting signal 1 (PTS1), a tripeptide of Ser-Lys-Leu (SKL) at the C-terminus, but did not on that of catalase harboring a divergent PTS1, Lys-Ala-Asn-Leu (KANL) sequence. 3-ketoacyl-CoA thiolase (hereafter referred to as thiolase), which harbors an extension sequence (PTS2) at the N-terminus, never appeared to be affected on the peroxisomal localization in the mutant cells. When thiolase was examined on the molecular size in the mutant cells, the enzyme existed as the larger precursor form in the peroxisomes at 37 degrees C and a considerable part (almost half) was converted to the mature size at 30 degrees C. These results indicate that the amino acid substitution, Gly306Arg in PTS1RS and/or Gly343Arg in PTSRL, gives rise to TS phenotype on the peroxisomal translocation of PTS1 proteins and the maturation of PTS2 protein.

Amino Acid Sequence↗
PubMed2001

HNF-1 regulates the liver-specific transcription of the chipmunk HP-20 gene.

The chipmunk hibernation-specific protein HP-20 is a component of the 140 kDa complex that drastically decreases in the blood during hibernation, and its gene is expressed specifically in the liver. To reveal molecular mechanisms underlying the liver-specific transcription of the HP-20 gene, we isolated chipmunk HP-20 genomic clones. The HP-20 gene spans approximately 6 kb, and consists of three exons. The transcription start site, as determined by 5' RACE-PCR analysis, was found to be 160 bp upstream of the translation initiation codon. Transient transfection studies in HepG2 cells revealed that the 57 bp 5' flanking sequence was sufficient for the liver-specific promoter activity. A database search revealed that this region contains a potential binding site for hepatocyte nuclear factor-1 (HNF-1). In a gel retardation assay, in vitro-synthesized HNF-1 bound to the 5' flanking sequence from -52 to -26. A similar shifted band was also observed with HepG2 nuclear extracts, and this complex was super-shifted by an anti-(HNF-1) Ig. When transfected into COS-7 cells, HNF-1 transactivated transcription from the HP-20 gene promoter, and this activity was abolished by a mutation of the HNF-1 binding site, indicating that HNF-1 plays an important role in HP-20 gene expression.

5' Flanking Region↗
PubMed2001

The N-terminal internal region of BLM is required for the formation of dots/rod-like structures which are associated with SUMO-1.

Bloom Syndrome (BS) is a human autosomal genetic disorder characterized by a predisposition to a variety of malignant tumors. The gene responsible for BS encodes a protein (BLM) consisting of 1417 amino acids with a nuclear localization signal in the C-terminal region, which is a member of the RecQ helicase family. We previously showed, using a yeast two-hybrid system, that BLM interacted with Ubc9, which is the conjugating enzyme of SUMO-1 (small ubiquitin-related modifier-1). In the present study, we exogenously expressed a green fluorescent protein-tagged Bloom syndrome protein, GFP-BLM, in human 293EBNA cells and found that it formed dots/rod-like structures associated with SUMO-1 in the nucleus. Deletion experiments indicated that the region from amino acids 238 to 586 of BLM is required for the formation of dots/rod-like structures associated with SUMO-1, and the DNA helicase domain, but not the helicase activity itself, slightly affected the formation and/or stability of these structures. Expression of a GFP-BLM which contained the 238-586 region, but lacked the C-terminal nuclear localization signal, resulted in localization to the cytoplasm without the formation of dots/rod-like structures and association with SUMO-1, indicating that these events occur only in the nucleus.

Adenosine Triphosphatases↗
PubMed2001

Phenotype-genotype relationships in peroxisome biogenesis disorders of PEX1-defective complementation group 1 are defined by Pex1p-Pex6p interaction.

The peroxisome biogenesis disorders (PBDs), including Zellweger syndrome (ZS), neonatal adrenoleucodystrophy (NALD) and infantile Refsum disease (IRD), are fatal autosomal recessive diseases caused by impaired peroxisome biogenesis, of which 12 genotypes have been reported. ZS patients manifest the severest clinical and biochemical abnormalities, whereas those with NALD and IRD show less severity and the mildest features respectively. We have reported previously that temperature-sensitive peroxisome assembly is responsible for the mildness of the clinical features of IRD. PEX1 is the causative gene for PBDs of complementation group E (CG-E, CG1 in the U.S.A. and Europe), the PBDs of highest incidence, encoding the peroxin Pex1p of the AAA ATPase family. It has been also reported that Pex1p and Pex6p interact with each other. In the present study we investigated phenotype-genotype relationships of CG1 PBDs. Pex1p from IRD such as Pex1p with the most frequently identified mutation at G843D was largely degraded in vivo at 37 degrees C, whereas a normal level of Pex1p was detectable at the permissive temperature. In contrast, PEX1 proteins derived from ZS patients, including proteins with a mutation at L664P or the deletion of residues 634-690, were stably present at both temperatures. Pex1p-G843D interacted with Pex6p at approx. 50% of the level of normal Pex1p, whereas Pex1p from ZS patients mostly showing non-temperature-sensitive peroxisome biogenesis hardly bound to Pex6p. Taking these results together, we consider it most likely that the stability of Pex1p reflects temperature-sensitive peroxisome assembly in IRD fibroblasts. Failure in Pex1p-Pex6p interaction gives rise to more severe abnormalities, such as those manifested by patients with ZS.

ATPases Associated with Diverse Cellular Activitie↗
PubMed2001

Evidence for metaboreceptor stimulation of sweating in normothermic and heat-stressed humans.

1. Isometric handgrip (IHG) exercise increases sweat rate and arterial blood pressure, and both remain elevated during post-exercise ischaemia. The purpose of this study was to identify whether the elevation in arterial blood pressure during post-exercise ischaemia contributes to the increase in sweating. 2. In normothermia and during whole-body heating, 2 min IHG exercise at 40% maximal voluntary contraction, followed by 2 min post-exercise ischaemia, was performed with and without bolus intravenous administration of sodium nitroprusside during the ischaemic period. Sodium nitroprusside was administered to reduce blood pressure during post-exercise ischaemia to pre-exercise levels. Sweat rate was monitored over two microdialysis membranes placed in the dermal space of forearm skin. One membrane was perfused with the acetylcholinesterase inhibitor neostigmine, while the other was perfused with the vehicle. 3. In normothermia, IHG exercise increased sweat rate at the neostigmine-treated site but not at the control site. Sweat rate remained elevated during post-exercise ischaemia even after mean arterial blood pressure returned to the pre-IHG exercise baseline. Subsequent removal of the ischaemia stimulus returned sweat rate to pre-IHG exercise levels. Sweat rate during post-exercise ischaemia without sodium nitroprusside administration followed a similar pattern. 4. During whole-body heating, IHG exercise increased sweat rate at both neostigmine-treated and untreated sites. Similarly, regardless of whether mean arterial blood pressure remained elevated or was reduced during post-exercise ischaemia, sweat rate remained elevated during the ischaemic period. 5. These results suggest that sweating in non-glabrous skin during post-IHG exercise ischaemia is activated by metaboreflex stimulation and not via baroreceptor loading.

Adult↗
PubMed2001

Urinary organic acids in peroxisomal disorders: a simple screening method.

Using GC-MS, we studied urinary organic acids in 20 Japanese patients with peroxisomal disorders, including Zellweger syndrome (ZS), neonatal adrenoleukodystrophy, and single deficiency of peroxisomal beta-oxidation enzymes. Non-ketotic dicarboxylic aciduria with elevated sebacate/adipate molar ratio was observed in 19 of the 20 patients. Elevation of 2-hydroxysebacate and epoxydicarboxylic acids were seen in 13 and 18, respectively. Tyrosyluria was remarkable in all patients. In two ZS patients, we tracked the time course from birth to infancy, and all the above stated findings were detected, except for one sample. Urinary organic acid analysis is indeed useful for screening subjects with peroxisomal disorders.

Acids↗
PubMed2001

The clinical phenotype and outcome of mitochondrial acetoacetyl-CoA thiolase deficiency (beta-ketothiolase or T2 deficiency) in 26 enzymatically proved and mutation-defined patients.

Mitochondrial acetoacetyl-CoA thiolase (T2 enzyme) deficiency (MIM 203750) is an autosomal recessive disorder of isoleucine and ketone-body metabolism. We determined the molecular basis of T2 enzyme deficiency in 26 patients at the levels of skin fibroblast enzyme activity, protein integrity, and DNA nucleotide sequence. Thirty different disease-associated alleles were identified. From these data we predicted that T2 in 6 of the 26 patients would have a mild effect on the enzyme protein and 20 would have a severe effect from their mutant genotypes. The corresponding clinical data were collected (by interviews and questionnaires) for the patients in the two groups. We found that genotype does not predict clinical severity and mutant sibs can have different clinical phenotypes; there were no consistent differences in clinical severity between patients with null-conferring or residual-conferring genotypes for T2 activity; only the absence of or a low urinary excretion of tiglyglycine during ketoacidosis correlated with a mild genotype. In general, T2 deficiency has a favorable outcome and 23 of 26 patients developed normally; one died during the first ketoacidotic episode and two have developmental delay. The median age at onset for the first ketoacidotic episode is 15 months (range 3 days to 48 months). The frequency of attacks falls with age, the last in our series occurring at 10 years of age; 11 patients had only one episode and 3 patients had none. We conclude that clinical consequences of T2 deficiency are avoidable with early diagnosis, appropriate management of ketoacidosis, and modest protein restriction.

Acetyl-CoA C-Acyltransferase↗
PubMed2001

A novel single-base substitution (380C>T) that activates a 5-base downstream cryptic splice-acceptor site within exon 5 in almost all transcripts in the human mitochondrial acetoacetyl-CoA thiolase gene.

Most mutation-related aberrant splicing occurs in the conserved splice-acceptor and -donor sites and some exonic mutations also affect splicing. We identified and characterized a point mutation (380C>T) in a Spanish patient (GK25) with mitochondrial acetoacetyl-CoA thiolase (T2) deficiency. GK25 is a homozygote of 380C>T, which activates a cryptic splice-acceptor site 5 bases downstream from 380C>T within exon 5, causing aberrant splicing in 94% of transcripts. The aberrant splicing results in a 17-amino acids deletion, including the active-site 126Cys. The 380C>T mutation also results in A127V mutation in 6% of transcripts. Transient expression analysis showed that the A127V mutation did not retain T2 activity, indicating that 380C>T was a null mutation. Although this cryptic splice site has a higher Shapiro and Senapathy's score (86) in even a normal sequence than the authentic splice-acceptor site of intron 4 (78), it is not used in normal controls. While the 380C>T mutation increases the score slightly (90), the cryptic splice site is used in almost all transcripts in GK25 fibroblasts. This is an example in which a point mutation activates a cryptic splice-acceptor site motif that is used preferentially over the upstream authentic splice site.

Acetyl-CoA C-Acyltransferase↗
PubMed2001

The cell surface-expressed HSC70-like molecule preferentially reacts with the rat T-cell receptor Vdelta6 family.

We previously showed that the cell surface-expressed Mr 70,000 heat shock cognate (hsc70, a constitutively expressed member of the hsp70 family) protein-like molecule (#067 molecule) interacts with rat CD3+, CD4-, CD8-, T-cell receptor (TCR)alphabeta-, natural killer recetor-P1- T cells. This 70hsc-like molecule was also suggested to present cellular peptide antigens to these T cells. In the present study, we identified the genetic structure of the TCR by establishing T-cell hybridomas between these T cells and mouse BW5147 cells. Our data indicated that these T cells preferentially used TCRs with the Vdelta6 family. Analysis of the nucleotide sequence of the CDR3 junctional portion showed that there are substantial diversities, with insertion of seven to nine amino acid residues. These data provide indirect evidences for our hypothesis that an hsc70-like molecule could be presented together with cellular peptide antigens to particular T cells with TCR gammadelta chains. Since the expression of this hsc70-like #067 antigen on the cell surface is usually induced along with cell transformation by activated oncogenes, T cells with the TCR Vdelta6 family are likely to contribute to host resistance to tumor cells.

Amino Acid Sequence↗
PubMed2001

The peroxin Pex6p gene is impaired in peroxisomal biogenesis disorders of complementation group 6.

Human genetic peroxisomal biogenesis disorders (PBDs), such as Zellweger syndrome, comprise 13 different complementation groups (CGs). Eleven peroxin genes, termed PEXs, responsible for PBDs have been identified, whereas pathogenic genes for PBDs of 2CGs, CG-A (the same CG as CG8 in the United States and Europe) and CG6, remained unidentified. We herein provide several lines of novel evidence indicating that PEX6, the pathogenic gene for CG4, is impaired in PBD of CG6. Expression of PEX6 restored peroxisome assembly in fibroblasts from a CG6 PBD patient. This patient was a compound heterozygote for PEX6 gene alleles. Accordingly, by merging CG6 with CG4, human PBDs are now classified into 12CGs.

ATPases Associated with Diverse Cellular Activitie↗
PubMed2001

Acute idiopathic blind spot enlargement syndrome: prolonged retinal dysfunction revealed by multifocal electroretinogram technique.

PURPOSE: To report the multifocal electroretinographic findings of a patient with acute idiopathic blind spot enlargement syndrome recorded at 2 weeks and 4 months after the onset. METHODS: A 35-year-old patient underwent static visual field (Humphrey, 30-2) and multifocal electroretinogram testing at 2 weeks and 4 months after the onset of acute idiopathic blind spot enlargement. The stimulus array for the multifocal electroretinogram consisted of 61 hexagons, and the total recording time was about 4 minutes. RESULTS: At 2 weeks, the patient had a large, well-demarcated scotoma centered on the blind spot, and its margin extended to within 5 degrees of fixation. The multifocal electroretinograms were depressed in the areas corresponding to the scotoma. At 4 months, her visual symptoms disappeared and static visual fields showed normal sensitivities at almost all locations. However, the multifocal electroretinograms still revealed reduced focal responses in a broad area around the blind spot. CONCLUSION: These results suggest that multifocal electroretinograms may be used to detect subclinical retinal dysfunction in patients with acute idiopathic blind spot enlargement.

Acute Disease↗
PubMed2001

Excision of an invasive thymoma: a cure for ulcerative colitis?

Thymoma is often associated with other diseases, including autoimmune diseases and endocrine disorders. Ulcerative colitis is a nonspecific inflammatory disease and is thought to be an immune-mediated disorder; however, coincidence of thymoma and ulcerative colitis is rare. We treated a 51-year-old woman with invasive thymoma and ulcerative colitis. After excision of the invasive thymoma with partial resections of pericardium, left lung, brachiocephalic vein, and left phrenic nerve, the ulcerative colitis was cured.

Colitis, Ulcerative↗
PubMed2001

Multifocal pupillary light response fields in normal subjects and patients with visual field defects.

The optimal conditions for recording focal pupillary light responses with a multifocal stimulation technique were determined, and the technique was applied to normal subjects and patients with visual field defects. Thirty-seven hexagonal stimuli were presented on a TV monitor with a visual field of 40 degrees diameter under a constant background illumination. Using a slow (4.7 Hz) m-sequence, reliable focal responses were obtained in both normal subjects and patients. The pupillary field and visual field were well correlated in patients with retinal diseases, but the correlation was not strong in patients with optic-nerve diseases. Pupillary light responses were reduced in the blind hemifield in patients with post-geniculate lesions. These results indicate that the multifocal stimulation technique can be used clinically to obtain a pupillary field for objective visual field testing.

Adult↗
PubMed2001

The clinical course of childhood and adolescent adrenoleukodystrophy before and after Lorenzo's oil.

The clinical course of Japanese patients with childhood and adolescent adrenoleukodystrophy (ALD) who were treated with Lorenzo's oil after the progression of early symptoms was investigated by Kaplan-Meier analyses. Many of the early symptoms, including visual, personality, hearing, mental and speech disturbances, had already appeared when the oil therapy was started. These early symptoms appeared significantly faster than bed-ridden state or dysphagia (P<0.01). Gait disturbance appeared faster than bed-ridden state and dysphagia (P<0.05), and tended to appear later than the early symptoms. These results would reflect the early natural course of childhood and adolescent ALD before treatment, and provide a basis for the evaluation of therapeutic trials for ALD.

Adolescent↗
PubMed2001

Epileptic seizures and structural abnormalities in a patient with holoprosencephaly.

In a patient with holoprosencephaly, partial seizures had various initial ictal symptoms, and ictal EEGs showed epileptogenic foci in the right and left brain. Partial seizures did not culminate in secondary generalized tonic-clonic convulsions. Characteristic malformed structures contribute to the absence of secondary generalization and the presence of Jacksonian-type propagation.

Brain↗