Biomedical subjects

Y Fu

Publications and source records attributed to Y Fu.

At least 91 records · Page 5Linked to original sources

PubMed1998

[The microsatellite instability and loss of heterozygosity in chromosome translocation breakpoint in leukemia].

OBJECTIVE: To investigate the microsatellite instability (MSI) and loss of heterozygosity (LOH) near the chromosome translocation breakpoints of various types of leukemia. METHODS: PCR amplification of 11 microsatellite loci closely linked to chromosome translocation breakpoints in 30 leukemia patients. RESULTS: Twenty one in 30 cases of leukemia had MSI in from 1 to 14 microsatellite loci. Eleven in 15 cases of acute leukemia had microsatellite instability (MSI) in more than 1 locus; among them 4 cases had MSI in from 7 to 11 loci. In order of MSI presence rate were D22S315>D9S179>D16S515>D8S559>D12S89. In six cases of chronic myeloid leukemia, 4 cases had MSI in more than 3 loci. Thirteen (2/15) percent of acute leukemia patients had loss of heterozygosity (LOH); the rate was higher than that in other leukemia types. CONCLUSION: The results of MSI and LOH in many cases indicate that there are a lot of allelic alterations in leukemia patients, but those changes have no special relationship with chromosome translocation breakpoints in various leukemia types.

Chromosome Breakage↗

PubMed1998

Cloning and disruption of caPLB1, a phospholipase B gene involved in the pathogenicity of Candida albicans.

The Candida albicans PLB1 gene was cloned using a polymerase chain reaction-based approach relying on degenerate oligonucleotide primers designed according to the amino acid sequences of two peptide fragments obtained from a purified candidal enzyme displaying phospholipase activity (Mirbod, F., Banno, Y., Ghannoum, M. A., Ibrahim, A. S., Nakashima, S., Yasuo, K., Cole, G. T., and Nozawa, Y. (1995) Biochim. Biophys. Acta 1257, 181-188). Sequence analysis of a 6.7-kilobase pair EcoRI-ClaI genomic clone revealed a single open reading frame of 1818 base pairs that predicts for a pre-protein of 605 residues. Comparison of the putative candidal phospholipase with those of other proteins in data base revealed significant homology to known fungal phospholipase Bs from Saccharomyces cerevisiae (45%), Penicillium notatum (42%), Torulaspora delbrueckii (48%), and Schizosaccharomyces pombe (38%). Thus, we have cloned the gene encoding a C. albicans phospholipase B homolog. This gene, designated caPLB1, was mapped to chromosome 6. Disruption experiments revealed that the caplb1 null mutant is viable and displays no obvious phenotype. However, the virulence of strains deleted for caPLB1, as assessed in a murine model for hematogenously disseminated candidiasis, was significantly attenuated compared with the isogenic wild-type parental strain. Although deletion of caPLB1 did not produce any detectable effects on candidal adherence to human endothelial or epithelial cells, the ability of the caplb1 null mutant to penetrate host cells was dramatically reduced. Thus, phospholipase B may well contribute to the pathogenicity of C. albicans by abetting the fungus in damaging and traversing host cell membranes, processes which likely increase the rapidity of disseminated infection.

Amino Acid Sequence↗

PubMed1998

Mechanism of reductive activation of potato tuber ADP-glucose pyrophosphorylase.

The potato tuber (Solanum tuberosum L.) ADP-glucose pyrophosphorylase activity is activated by a incubation with ADP-glucose and dithiothreitol or by ATP, glucose- 1-phosphate, Ca2+, and dithiothreitol. The activation was accompanied by the appearance of new sulfhydryl groups as determined with 5, 5'-dithiobis(2-nitrobenzoic acid). By analyzing the activated and nonactivated enzymes on SDS-polyacrylamide gel electrophoresis under nonreducing conditions, it was found that an intermolecular disulfide bridge between the small subunits of the potato tuber enzyme was reduced during the activation. Further experiments showed that the activation was mediated via a slow reduction and subsequent rapid conformational change induced by ADP-glucose. The activation process could be reversed by oxidation with 5, 5'-dithiobis(2-nitrobenzoic acid). Incubation with ADP-glucose and dithiothreitol could reactivate the oxidized enzyme. Chemical modification experiments with [14C]iodoacetic acid and 4-vinylpyridine determined that the intermolecular disulfide bridge was located between Cys12 of the small subunits of the potato tuber enzyme. Mutation of Cys12 in the small subunit into either Ala or Ser eliminated the requirement of DTT on the activation and prevented the formation of the intermolecular disulfide of the potato tuber enzyme. The mutants had instantaneous activation rates as the wild-type in the reduced state. A two-step activation model is proposed.

Adenosine Triphosphate↗

PubMed1998

Characterization and chromosomal localization of PTP-NP-2, a new isoform of protein tyrosine phosphatase-like receptor, expressed on synaptic boutons.

Recently, there have been several reports describing the cloning and characterization of the novel family of protein tyrosine phosphatase-like receptor molecules (known as IA-2 and PTP-NP/PTP-IAR/IA-2beta/phogrin), which may act as autoantigens in diabetes. Here, we report the molecular characterization and chromosomal localization of a new isoform of this family in brain termed PTP-NP-2 (for PTP-NP tyrosine phosphatase isoform), and its function in rat primary hippocampal neurons. PTP-NP-2 has 48% identity to IA-2. The principal difference between PTP-NP-2 and PTP-NP is a 17-amino-acid insert near the N-terminus of PTP-NP that is absent in PTP-NP-2. Genomic DNA analysis indicates that the 17-amino-acid insert is coded by a separate exon, suggesting that both IA-2beta and PTP-NP-2 are isoforms arising by alternate splicing of the same gene. Reverse transcriptase-PCR revealed that both isoforms are present in human SH-SY5Y neuroblastoma cells. PTP-NP-2 mRNA expression is highly restricted, with a 5.5-kb specific transcript in human fetal and adult brain and 5.5 and 3. 8 kb in human adult pancreas. SH-SY5Y neuroblastoma and U87-MG glioblastoma cells showed specific transcripts of 5.5 and 3.8 kb, respectively, indicating the existence of several isoforms of this molecule in the nervous system. The human gene encoding PTP-NP-2 was assigned to human chromosome 7q22-qter using Southern blot analysis of genomic DNAs from rodent/human somatic hybrid cell lines. Confocal microscopy analyses of rat primary hippocampal neurons revealed that PTP-NP-2 is abundantly expressed on synaptic boutons in primary neurons. Wild-type PTP-NP-2 showed no measurable tyrosine phosphatase activity using an in-vitro pNPP assay. Examination of the PTP-NP-2 catalytic consensus sequence revealed that this sequence differed from the typical tyrosine phosphatase-domain consensus sequence by an alanine to aspartate change (amino acid 930). Mutation of aspartate 930 to alanine produced a catalytically active enzyme, suggesting that native PTP-NP and its isoform PTP-NP-2 are catalytically inactive receptor protein tyrosine phosphatase homologues. Taken together, these results indicate that the tyrosine phosphatase PTP-NP-2 is a new isoform of PTP-NP tyrosine phosphatase, is expressed on synaptic boutons and may participate in the regulation of synaptic bouton endocytosis.

Adult↗

PubMed1998

Delivery of epitopes by the Salmonella type III secretion system for vaccine development.

Avirulent strains of Salmonella typhimurium are being considered as antigen delivery vectors. During its intracellular stage in the host, S. typhimurium resides within a membrane-bound compartment and is not an efficient inducer of class I-restricted immune responses. Viral epitopes were successfully delivered to the host-cell cytosol by using the type III protein secretion system of S. typhimurium. This resulted in class I-restricted immune responses that protected vaccinated animals against lethal infection. This approach may allow the efficient use of S. typhimurium as an antigen delivery system to control infections by pathogens that require this type of immune response for protection.

Animals↗

PubMed1998

Identification of novel susceptibility loci for inflammatory bowel disease on chromosomes 1p, 3q, and 4q: evidence for epistasis between 1p and IBD1.

The idiopathic inflammatory bowel diseases, Crohn's disease (CD) and ulcerative colitis (UC), are chronic, frequently disabling diseases of the intestines. Segregation analyses, twin concordance, and ethnic differences in familial risks have established that CD and UC are complex, non-Mendelian, related genetic disorders. We performed a genome-wide screen using 377 autosomal markers, on 297 CD, UC, or mixed relative pairs from 174 families, 37% Ashkenazim. We observed evidence for linkage at 3q for all families (multipoint logarithm of the odds score (MLod) = 2.29, P = 5.7 x 10(-4)), with greatest significance for non-Ashkenazim Caucasians (MLod = 3.39, P = 3.92 x 10(-5)), and at chromosome 1p (MLod = 2.65, P = 2.4 x 10(-4)) for all families. In a limited subset of mixed families (containing one member with CD and another with UC), evidence for linkage was observed on chromosome 4q (MLod = 2.76, P = 1.9 x 10(-4)), especially among Ashkenazim. There was confirmatory evidence for a CD locus, overlapping IBD1, in the pericentromeric region of chromosome 16 (MLod = 1.69, P = 2.6 x 10(-3)), particularly among Ashkenazim (MLod = 1.51, P = 7.8 x 10(-3)); however, positive MLod scores were observed over a very broad region of chromosome 16. Furthermore, evidence for epistasis between IBD1 and chromosome 1p was observed. Thirteen additional loci demonstrated nominal (MLod > 1.0, P < 0.016) evidence for linkage. This screen provides strong evidence that there are several major susceptibility loci contributing to the genetic risk for CD and UC.

Chromosome Mapping↗

PubMed1998

Antigen processing and autoimmunity. Evaluation of mRNA abundance and function of HLA-linked genes.

Quantitative defects in the density of conformationally correct human lymphocyte antigen (HLA) class I complexes on the surface of lymphocytes are apparent in patients with diverse HLA-linked autoimmune diseases, including Type I diabetes and Sjögren's syndrome. First, HLA class I expression was investigated in individuals with two rare and genetically divergent polyglandular autoimmune diseases. Polyglandular failure patients whose disease showed HLA linkage, but not those whose disease was not HLA linked, exhibited decreased HLA class I expression on the surface of their lymphocytes as well as a reduced abundance of transcripts of the HLA-linked genes Tap1 and Tap2, both of which encode proteins that contribute to HLA class I processing. Second, lymphocytes from patients with insulin-dependent diabetes mellitus (IDDM), Sjögren's syndrome, Graves' disease, and Hashimoto's disease showed varying degrees of decreased abundance of mRNAs that encode Tap1, Tap2, Lmp2, or Lmp7 (the latter two proteins also contribute to HLA class I processing). Third, in twins discordant for IDDM, reduced transcript abundance was preferential to diabetic subjects. Fourth, functional assays of isolated diabetic proteasomes, the peptide cutting complex containing LMP2 and LMP7 proteins, revealed altered peptidase activity. These data suggest that defective transcription of HLA class I-processing genes could contribute to the quantitative defect in cell-surface expression in autoimmune lymphocytes of HLA-controlled disease.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1998

Desensitization and resensitization of norepinephrine release in the rat hippocampus with repeated nicotine administration.

Desensitization of norepinephrine release was investigated with repeated intravenous (i.v.) infusions of nicotine and in vivo microdialysis of the hippocampus. At 100 min intervals, rats received three infusions of one of the following doses of nicotine: 0.045, 0.09 or 0.135 mg/kg. Doses of 0.09 mg/kg or higher increased norepinephrine release (F= 2.41, P < 0.05). However, the norepinephrine response to the second or third infusion was significantly reduced, compared to the first. The extent of desensitization and rate of resensitization was investigated further by administering consecutive infusions of nicotine (0.135 mg/kg) 40, 60, 100 or 200 min apart. Less norepinephrine was released after a second nicotine infusion given 40 to 100 min later, but this was not reduced further by a third infusion. Norepinephrine release was unchanged with a 200 min inter-infusion interval. Therefore, in the hippocampus, maximal desensitization of nicotine-stimulated norepinephrine release occurs as early as 40 min and persists for at least 100 min; thereafter, resensitization becomes the dominant process.

Animals↗

PubMed1998

Rabies virus entry into endosomes in IMR-32 human neuroblastoma cells.

Early events in rabies virus entry into cultured IMR-32 human neuroblastoma cells were investigated. After adsorption of rabies virus to the cell surface in the cold and warming to 37 degrees C in the presence of tracers for early endosomes, rabies virus and tracers were localized by immunofluorescence microscopy. After 5 min, rabies virus colocalized with Lucifer Yellow, Texas Red-dextran, rhodamine-wheat germ agglutinin, and transferrin receptor in puncta in the cell body, neurites, and nerve terminals. Rabies virus did not colocalize with lysosomal glycoprotein. An acidotropic probe revealed that some of the virus-containing puncta were acidified. Rabies virus also colocalized with synapsin I, a synaptic vesicle marker, in swellings along processes, indicating some virus enters nerve terminals. Electron microscopy revealed the presence of rabies virus within irregular membrane compartments located near the cell surface in the cell body and neurites. The membrane of the virus particle was often continuous with that of the vacuole. It is concluded that rabies virus enters IMR-32 neuroblastoma cells by adsorptive endocytosis and that, shortly after entry, rabies virus is located within and fuses with acidic endosomes.

Adsorption↗

PubMed1998

American families with Crohn's disease have strong evidence for linkage to chromosome 16 but not chromosome 12.

BACKGROUND & AIMS: Two European genome-wide screens for inflammatory bowel disease have identified two significant regions of linkage on chromosomes 16 (IBD1) and 12 (IBD2) and two regions with suggestive levels of significance (chromosomes 3p and 7q). The aim of this study was to determine if there was evidence for linkage to these regions in non-Jewish and Ashkenazi Jewish families multiplex for Crohn's disease from the United States. METHODS: One hundred forty-eight affected relative pairs, 34% Ashkenazim, were genotyped with 10-14 highly polymorphic markers overlying each candidate region. Nonparametric multipoint and two-point linkage analyses were performed. RESULTS: Significant evidence for replication of linkage was found only for the chromosome 16 locus, IBD1, maximal at D16S769 (nonparametric linkage score [NPL], 2.49; P = 0.007). Analysis by ethnicity showed stronger evidence for Ashkenazim (D16S769; NPL = 2. 52; P = 0.007) than for non-Jewish white populations (D16S401; NPL = 1.40; P = 0.082). There was no significant evidence for replication on chromosome 12 (IBD2). Minimal evidence for extension of linkage evidence was observed for the chromosomes 3p and 7q regions. CONCLUSIONS: American families, particularly Ashkenazim, have significant evidence for the Crohn's disease susceptibility locus, IBD1, on chromosome 16, but not for IBD2 on chromosome 12.

Chromosome Mapping↗

PubMed1998

The effect of high altitude and caloric restriction on reactive carbonyl derivatives and activity of glutamine synthetase in rat brain.

Both exposure to high altitude (HA) and caloric restriction (CR) may influence free radical generation. The aim of the present study was to evaluate the effects of 4 wk chronic exposure to 4,000m of HA and CR (60% of CR of control (C) rats) on protein oxidation in brain. Eighteen rats with similar body mass were assigned to CR, HA and C rats. Reactive carbonyl derivatives (RCD), marker of protein oxidation, were measured by Western blot. In addition, the activity and protein content of glutamine synthetase (GS) were determined. The body mass of C rats was significantly higher (P< 0.001) than that of HA and CR groups. The quantified signal intensity of RCD was significantly stronger in C rats than in HA and CR rats. The activity of GS was significantly increased in CR rats, while the protein content of GS was decreased in HA rats compared to C group. The data suggest that both HA and CR decreases the accumulation of RCD in the brain, however the mechanism of the decrease seems to be different during HA and CR.

Altitude↗

PubMed1998

Response of the hypothalamo-pituitary-adrenal axis to nicotine.

Nicotine has been shown to be a potent stimulus for the secretion of the stress-responsive hormones, adrenocorticotropin (ACTH) and prolactin. This paper reviews the findings by our laboratory and others that demonstrate the polysynaptic pathways involved in the neuroendocrine responses to systemic nicotine. It will focus primarily on the hypothalamo-pituitary-adrenal (HPA) axis and the effect of nicotine on ACTH secretion, with supplementary information on prolactin secretion, where relevant. Data are presented demonstrating that nicotine acts via a central mechanism to stimulate indirectly the release of ACTH from the anterior pituitary corticotropes. Nicotine does not appear to act directly at the hypothalamic paraventricular nucleus (PVN), the site of the corticotropin-releasing hormone (CRH) neurons crucial to the regulation of ACTH. However, brainstem catecholaminergic regions projecting to the PVN showed a regionally selective and dose-dependent sensitivity to nicotine, particularly the noradrenergic/adrenergic nucleus tractus solitarius (NTS). A reduction in the modulatory effect of these catecholamines (by neurotoxic lesion, synthetic enzyme inhibitors or adrenergic receptor antagonists) resulted in an inhibition of nicotine-stimulated ACTH secretion. In addition, blockade of nicotinic cholinergic receptors (NAchRs) in the brainstem by the antagonist, mecamylamine, resulted in a dose-dependent reduction in norepinephrine (NE) release from terminals in the PVN, and a concomitant reduction in plasma ACTH. The differential sensitivity of these receptors to the nicotinic agonists, cytisine and nicotine, reflects the heterogeneity of the NAchR subtypes involved. The desensitization characteristics of the neuroendocrine responses to both acute and chronic nicotine exposure are indicative of an alteration in these NAchRs.

Animals↗

PubMed1998

Left ventricular geometric patterns and QT dispersion in borderline and mild hypertension: their evolution and regression.

To investigate whether QT dispersion increases in borderline and mild hypertension during a longitudinal observation of > 3 years and whether it is improved with medications, left ventricular geometric patterns and QT dispersion were studied with special regard to their longitudinal changes in 85 male borderline and mild hypertensive subjects with left ventricular mass index < 125 g/m2. These subjects were followed for > 3 years without medication. Thirty-two patients with a left ventricular mass index > 125 g/m2 at the end of follow-up period were further observed using antihypertensive drugs for an additional 3 years. Echocardiograms and electrocardiograms were obtained at the beginning and end of the follow-up period. At the end of the follow-up period, subjects were classified into four groups based on ventricular geometric patterns determined by left ventricular mass index and relative wall thickness in diastole. The QT dispersion was greater in patients with concentric hypertrophy (56+/-18 msec) than in patients with normal geometry (41+/-17 msec) (P < .05) and increased significantly in the former group during the follow-up period. After medication, the left ventricular mass index regressed and the QT dispersion decreased (from 55+/-21 to 50+/-26 msec, P < .01) in these patients. Thus, these findings suggest that changes in the QT dispersion reflect both concentric evolution and regression of left ventricular hypertrophy.

Adult↗

PubMed1998

No association between alpha-adducin 460 polymorphism and essential hypertension in a Japanese population.

Many unknown genetic factors are involved in the pathogenesis of hypertension. Recently, the reverse genetic approach revealed that some genetic variants, such as angiotensinogen, lipoprotein lipase, and alpha-adducin gene polymorphisms, increase the risk for hypertension. Both in rat and human, the genetic predisposition to hypertension was confirmed only for angiotensinogen and alpha-adducin genes. Adducin is a membrane cytoskeletal protein, which is thought to regulate sodium transport. Abnormalities of membrane sodium transport in the kidney play an important role in hypertension. A recent report by Cusi et al showed that the Trp allele of alpha-adducin polymorphism (Gly 460 Trp) is associated with an increased risk of hypertension in whites, which led us to carry out a case-control study to examine whether the same association is observed in the Japanese population. We recruited 170 hypertensive and 194 normotensive Japanese subjects and compared the genotype distribution of alpha-adducin 460 polymorphism between cases and controls and between whites and Japanese. Trp allele frequency of controls in the Japanese subjects was twice as high as in the whites. However, no association was observed between alpha-adducin polymorphism and hypertension. Furthermore, alpha-adducin 460 polymorphism was not associated with any clinical characteristics. Accordingly, we concluded that alpha-adducin 460 polymorphism is not a major genetic risk for hypertension in Japanese people.

Alleles↗

PubMed1998

Methylenetetrahydrofolate reductase gene polymorphism: relation to blood pressure and cerebrovascular disease.

Hyperhomocysteinemia is reported to be associated with an increase in the incidence of ischemic heart disease and cerebrovascular disease. Genetic aberrations in methylenetetrahydrofolate reductase (MTHFR) may account for reduced enzyme activity and elevated plasma homocysteine level. A recent report revealed that a common mutation (677C to T; Ala to Val) in the MTHFR gene is associated with decreased specific MTHFR activity and with increased risk for coronary artery disease in the homozygous state (Val/Val). In the present study, we investigated whether the MTHFR gene is a genetic risk factor for cerebrovascular disease (CVD). To undertake a case-control study, we selected the patients with cerebral infarction (n = 48) or cerebral hemorrhage (n = 35) and examined the association between MTHFR gene polymorphism and CVD. The genotype distribution of the MTHFR gene was not significantly different between cases and controls. Because the possibility of matching the morbidity of the effects of hypertension, the lack of association could not be excluded in the first study; however, we also examined whether the MTHFR mutation was associated with any clinical risk factor for CVD or with hypertension. It turned out that the subjects with the Val allele of the MTHFR gene had significantly lower blood pressure than the subjects with other genotypes in the general population (P = .02), and that the frequency of the Val/Val genotype in hypertensive subjects (n = 173) was significantly lower than in control subjects (n = 184) (P = .03). From these results, we conclude that the Val/Val homozygous state of the MTHFR gene increased the risk of thrombosis, but reduced the blood pressure, which resulted in the lack of increased risk for CVD.

Cerebrovascular Disorders↗

PubMed1998

Cytokine regulation of secondary lymphoid organ development.

Lymphotoxin and tumor necrosis factor provide essential signals for the formation of secondary lymphoid tissue structures. Lymphotoxin in its membrane form (LT alpha 1 beta 2 heterotrimer) is required for the development of lymph nodes and Peyer's patches and supports the development of normal spleen structure. In the spleen, lymphotoxin acts during embryonic development to support the formation of distinct B and T cell zones. Lymphotoxin also acts in a tonic fashion-supporting the formation and maintenance of the follicular dendritic cell network and of primary B cell follicle structure. The cells that deliver the tonic lymphotoxin signal supporting follicular dendritic cell structure are B cells; thus, B cells participate fundamentally in the development of the lymphoid tissue structure in which they subsequently mature.

Animals↗

PubMed1998

Viral sequences enable efficient and tissue-specific expression of transgenes in Xenopus.

Expression of transgenes within a single generation by direct DNA injection into vertebrate embryos has been plagued by inefficient and nonuniform gene expression. We report a novel strategy for efficient and stable expression of transgenes driven by both ubiquitous and tissue-specific promoters by direct DNA injection into developing Xenopus laevis embryos. This strategy involves flanking expression cassettes of interest with inverted terminal repeat sequences (ITRs) from adeno-associated virus. Our results suggest that the ITR strategy may be generally applicable to other systems, such as zebra fish and embryonic stem cells, and may enable tissue-specific expression of transgenes in problematic contexts.

Actins↗

PubMed1998

The Salmonella typhimurium tyrosine phosphatase SptP is translocated into host cells and disrupts the actin cytoskeleton.

The Salmonella typhimurium protein tyrosine phosphatase SptP is a target of the centisome 63 type III protein secrtion system. This system is essential for the interaction of these bacteria with host cells. We have shown here by a combination of biochemical and microscopy techniques that S. typhimurium directs the translocation of SptP into cultured epithelial cells. Translocation requires the function of the secreted proteins, SipB, SipC and SipD, as strains carrying mutations in any of the genes encoding these proteins fail to translocate SptP. Microinjection of purified GST-SptP into cultured cells results in the disruption of the actin cytoskeleton and the disappearance of stress fibres. These changes are reversible, as microinjected cells regain the normal appearance of their actin cytoskeleton upon prolonged incubation. Microinjection of the catalytically active GST-SptP(C481S) protein results in changes similar to those induced by the wild-type toxin. Furthermore, microinjection of a fusion protein between GST and the first 285 amino acids of SptP also leads to identical disruption of the host cell actin cytoskeleton, indicating that the amino-terminal half of SptP is sufficient to mediate this effect. However, microinjection of a fusion protein between GST and the last 259 amino acids of SptP also disrupted the normal appearance of the cytoskeleton. These results support the hypothesis that SptP is an effector protein arranged in modular domains that may co-operate with each other to exert relate functions.

Actins↗