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Biomedical subjects

Isao Saito

Publications and source records attributed to Isao Saito.

At least 55 records · Page 3Linked to original sources
PubMed2005

A cross-sectional study of alcohol drinking and health-related quality of life among male workers in Japan.

BACKGROUND: Although light and moderate alcohol drinkers are likely to have better subjective health, the sub-scales for subjective health have not been well documented. METHODS: We studied 4,521 male workers aged 25 yr and older with no history of cancer or cardiovascular disease, in 12 occupational groups in Japan. Data were from the High-risk and Population Strategy for Occupational Health Promotion Study (HIPOP-OHP). Drinking status was classified according to daily alcohol intake or frequency of drinking. We assessed the health-related quality of life (HRQOL) based on scores for five scales of the SF-36. RESULTS: Decreased odds ratios of sub-optimal HRQOL conditions, defined as less than the median SF-36 scores, for Role-Physical and General Health were found among persons who consumed 1.0 to 22.9 g/d of alcohol. Odds ratios for sub-optimal Vitality conditions were lowered according to increased levels of alcohol intake. Role-Emotional scores were not associated with alcohol drinking. People who drank 5 to 6 d/wk had higher levels of Role-Physical and Vitality, and those who drank 1 to 2 d/wk had better Vitality and Mental Health scores than non-drinkers. When adjusted for age, marital status, working hours, physical activity at work, self-reported job stress, smoking, regular exercise, hypertension, hyperlipidemia, and diabetes, the associations were almost unchanged except for General Health. CONCLUSIONS: Associations of drinking patterns with subjective health varied in five sub-scales of the SF-36. Overall, alcohol drinkers rated their health as good in comparison with non-drinkers.

Adult↗
PubMed2005

Neurotrophin-4/5-depletion induces a delay in maturation of the periodontal Ruffini endings in mice.

Neurotrophin-4/5 (NT-4/5) - a member of the neurotrophic factors - is a ligand for TrkB, which has been reported to be expressed in the mechanoreceptive Ruffini endings of the periodontal ligament. The present study examined developmental changes in the terminal morphology and neural density in homozygous mice with a targeted disruption of the nt-4/5 gene and wild-type mice by immunohistochemistry for protein gene product 9.5 (PGP 9.5), a general neuronal marker, and by quantitative analysis using an image analyzer. Postnatal development of terminal Schwann cells was also investigated by enzymatic histochemistry for non-specific cholinesterase activity (ChE). Furthermore, the immuno-expression of TrkB and low affinity nerve growth factor receptor (p75-NGFR) was surveyed in the periodontal Ruffini endings as well as trigeminal ganglion. At postnatal 1 week, the lingual periodontal ligament of both types of mice contained PGP 9.5-positive nerve fibers showing a tree-like ramification with axonal swellings in their course. In both types of mice at 2 weeks of age, comparatively thick nerve fibers with a smooth outline increased in number, and frequently ramified to form nerve terminals with dendritic profiles. However, no typical Ruffini endings with irregular outlines observed in the adult wild-type mice were found in the periodontal ligament at this stage. At postnatal 3 weeks, typical Ruffini endings with irregular outlines were discernable in the periodontal ligament of the wild-type mice while the dendritic endings showing smooth outlines were restricted to the homozygous mice. After postnatal 8 weeks, both types of mice showed an increase in the number of Ruffini endings, but the morphology differed between the wild-type and NT-4/5 homozygous mice. In the wild-type mice, a major population of the Ruffini endings expanded their axonal branches and developed many microprojections, resulting in a reduction of endings with smooth outlines. In contrast, we failed to find such typical Ruffini endings in the periodontal ligament of the homozygous mice: A majority of the periodontal Ruffini endings continued to show smooth outlines at postnatal 12 weeks. Quantitative analysis on neural density demonstrated a reduction in the homozygous mice with a significant difference by postnatal 8 weeks. Enzymatic histochemistry for non-specific ChE did not exhibit a distinct difference in the distribution and density of terminal Schwann cells between wild-type and homozygous mice. Furthermore, TrkB and p75-NGFR mRNA and proteins did not differ in the trigeminal ganglion between the two types. The periodontal Ruffini endings also displayed immunoreactivities for TrkB and p75- NGFR in both phenotypes. These findings suggest that the nt-4/5 gene depletion caused a delay in the formation and maturation of the periodontal Ruffini endings in the mice by inhibiting the growth of the periodontal nerves at an early stage, and indicate that multiple neurotrophins such as NT- 4/5 and BDNF might play roles in the development and/or maturation of the periodontal Ruffini endings.

Animals↗
PubMed2004

Photostimulated hole transport through a DNA duplex immobilized on a gold electrode.

Photostimulated hole transport through DNA duplexes immobilized on gold electrodes has been investigated. By modifying a gold electrode with a DNA duplex containing a photosensitizer, we have observed a sequence-dependent cathodic photocurrent. DNA acts as a good mediator for cathodic photocurrent when appropriate sequences are selected.

Anthraquinones↗
PubMed2004

Synthesis of an artificial hole-transporting nucleoside triphosphate, dMDATP, and its enzymatic incorporation into DNA.

We report on the enzymatic synthesis of DNA wires by successive incorporation of artificial nucleotides. We synthesized an efficient hole-transporting nucleoside triphosphate, d(MD)ATP, and have examined the screening of polymerases and reverse transcriptases for enzymatic incorporation of d(MD)ATP into DNA. We found that KOD Dash is an effective enzyme for the enzymatic synthesis of (MD)A-containing DNA. Furthermore, the efficiency of enzymatic incorporation was remarkably improved by the addition of manganese chloride. By applying the PCR method to the synthesis of DNA containing a run of (MD)A, the (MD)A run was amplified up to 6 x 10(6) times. (MD)A runs prepared by the enzymatic method exhibited a high hole-transporting ability.

Adenosine Triphosphate↗
PubMed2004

DNA logic gates.

A conceptually new logic gate based on DNA has been devised. Methoxybenzodeazaadenine ((MD)A), an artificial nucleobase which we recently developed for efficient hole transport through DNA, formed stable base pairs with T and C. However, a reasonable hole-transport efficiency was observed in the reaction for the duplex containing an (MD)A/T base pair, whereas the hole transport was strongly suppressed in the reaction using a duplex where the base opposite (MD)A was replaced by C. The influence of complementary pyrimidines on the efficiency of hole transport through (MD)A was quite contrary to the selectivity observed for hole transport through G. The orthogonality of the modulation of these hole-transport properties by complementary pyrimidine bases is promising for the design of a new molecular logic gate. The logic gate system was executed by hole transport through short DNA duplexes, which consisted of the "logic gate strand", containing hole-transporting nucleobases, and the "input strand", containing pyrimidines which modulate the hole-transport efficiency of logic bases. A logic gate strand containing multiple (MD)A bases in series provided the basis for a sharp AND logic action. On the other hand, for OR logic and combinational logic, conversion of Boolean expressions to standard sum-of-product (SOP) expressions was indispensable. Three logic gate strands were designed for OR logic according to each product term in the standard SOP expression of OR logic. The hole-transport efficiency observed for the mixed sample of logic gate strands exhibited an OR logic behavior. This approach is generally applicable to the design of other complicated combinational logic circuits such as the full-adder.

Base Sequence↗
PubMed2004

Cytosine detection by a fluorescein-labeled probe containing base-discriminating fluorescent nucleobase.

We report on a new method for the detection of a base at a specific site in a DNA sequence by monitoring the fluorescence emission of fluorescein. To achieve this goal, we developed a new base-discriminating fluorescent (BDF) nucleobase, naphthodeazaadenine ((ND)A). The fluorescence spectrum of the duplex possessing a cytosine base as a complementary base of (ND)A showed a fluorescence peak at 383 nm when using an excitation wavelength of 350 nm. When the complementary base of (ND)A was one of the other bases, the fluorescence intensity was very low. The fluorescence emission spectrum of (ND)A overlapped with the fluorescence excitation spectrum of fluorescein in the wavelength range of 400-500 nm. Thus, we designed FRET-BDF probes containing (ND)A as the FRET donor and fluorescein as the acceptor. The interaction of these two fluorophores, which are separated by defined base pairs, allowed an efficient energy transfer that resulted in a dominant fluorescence emission of fluorescein at 520 nm when using an excitation wavelength of 350 nm. Fluorescence emission from FRET-BDF probes was observed only when the complementary base of (ND)A is C, thus achieving a clear distinction of a C base on the complementary DNA strand. However, the general utility of our method is limited due to the quenching of the (ND)A fluorescence by a G/C base pair flanking (ND)A.

Base Pairing↗
PubMed2004

2-Ureidoquinoline: a useful molecular element for stabilizing single cytosine and thymine bulges.

We have demonstrated that aromatic heterocycles having hydrogen-bonding surfaces complementary to those of nucleotide bases are effective molecular elements for the binding to single nucleotide bulges and base mismatches. We here report that a new molecule, 2-ureidoquinoline having an alignment of hydrogen-bonding groups in the order of acceptor-donor-donor stabilizes single cytosine and thymine bulges in duplex DNAs. Furthermore, a dimeric form of 2-ureidoquinoline stabilizes cytosine-cytosine and cytosine-thymine mismatches.

Base Pair Mismatch↗
PubMed2004

The binding of guanine-guanine mismatched DNA to naphthyridine dimer immobilized sensor surfaces: kinetic aspects.

Naphthyridine dimer composed of two naphthyridine chromophores and a linker connecting them strongly, and selectively, binds to the guanine-guanine mismatch in duplex DNA. The kinetics for the binding of the G-G mismatch to the naphthyridine dimer was investigated by surface plasmon resonance assay. The sensor surface was prepared by immobilizing naphthyridine dimer through a long poly(ethylene oxide) linker with the ligand density of 9.1 x 10(-12) fmolnm(-2). The kinetic analyses revealed that the binding of the G-G mismatch was sequence dependent on the flanking base pairs, and the G-G mismatches flanking at least one G-C base pair bound to the surface via a two-step process with a 1:1 DNA-ligand stoichiometry. The first association rate constant for the binding of the G-G mismatch in the 5'-CGG-3'/3'-GGC-5' sequence to the naphthyridine dimer-immobilized sensor surface was 3.2 x 10(3)M(-1)s(-1) and the first dissociation rate constant was 1.4 x 10(-2)s(-1). The association and dissociation rate constants for the second step were insensitive to the flanking sequences, and were almost of the same order of magnitude as the first dissociation rate constant. This indicates that the second step had only a small energetic contribution to the binding. The association constant calculated from kinetic parameters was 2.7 x 10(5)M(-1), which is significantly smaller than the apparent association constants obtained from experiments in solution. Electrospray ionization time-of-flight (ESI-TOF) mass spectrometry on the complex produced from the G-G mismatch and naphthyridine dimer showed the formation of the 1:1 complex and a 1:2 DNA-ligand complex in solution. The latter complex became the dominant complex when a six-fold excess of naphthyridine dimer was added to DNA.

Base Pair Mismatch↗
PubMed2004

Pyrene-labeled base-discriminating fluorescent DNA probes for homogeneous SNP typing.

This paper describes the design of novel base-discriminating fluorescent (BDF) nucleobases and their application to single nucleotide polymorphism (SNP) typing. We devised novel BDF nucleosides, (Py)U and (Py)C, which contain a pyrenecarboxamide chromophore connected by a propargyl linker. The fluorescence spectrum of the duplex containing a (Py)U/A base pair showed a strong emission at 397 nm on 327 nm excitation. In contrast, the fluorescence of duplexes containing (Py)U/N base pairs (N = C, G, or T) was considerably weaker. The proposed structure of the duplex containing a matched (Py)U/A base pair suggests that the high polarity near the pyrenecarboxamide group is responsible for the strong A-selective fluorescence emission. Moreover, the fluorescence of the duplex containing a (Py)U/A base pair was not quenched by a flanking C/G base pair. The fluorescence properties are quite different from previous BDF nucleobases, where fluorescence is quenchable by flanking C/G base pairs. The duplex containing the C derivative, (Py)C, selectively emitted fluorescence when the base opposite (Py)C was G. The drastic change of fluorescence intensity by the nature of the complementary base is extremely useful for SNP typing. (Py)U- and (Py)C-containing oligodeoxynucleotides acted as effective reporter probes for homogeneous SNP typing of DNA samples containing c-Ha-ras and BRCA2 SNP sites.

Amides↗
PubMed2004

Highly sensitive detection of GG mismatched DNA by surfaces immobilized naphthyridine dimer through poly(ethylene oxide) linkers.

Naphthyridine dimer is a unique molecule that strongly, and selectively, binds to the guanine-guanine mismatch in duplex DNA. We have synthesized naphthyridine dimers possessing a different length of poly(ethylene oxide) (PEO) linker, and immobilized them to CM5 sensor chip to carry out a surface plasmon resonance (SPR) assay of DNA duplexes containing a single base mismatch. The sensitivity of the sensor remarkably increased with increasing numbers of PEO units incorporated into the linker. With the sensor surface immobilized naphthyridine dimer for 1.5 x 10(3) response unit (RU) through three PEO units, the distinct SPR signal was observed at a concentration of 1 nM of the 27-mer G-G mismatch.

Base Pair Mismatch↗
PubMed2004

The SPR sensor detecting cytosine[bond]cytosine mismatches.

We have synthesized the first surface plasmon resonance (SPR) sensor that detects cytosine-cytosine (C[bond]C) mismatches in duplex DNA by immobilizing aminonaphthyridine dimer on the gold surface. The ligand consisting of two 2-aminonaphthyridine chromophores and an alkyl linker connecting them strongly stabilized the C[bond]C mismatches regardless of the flanking sequences. The fully matched duplexes were not stabilized at all under the same conditions. The C[bond]T, C[bond]A, and T[bond]T mismatches were also stabilized with a reduced efficiency. SPR analyses of mismatch-containing 27-mer duplexes were performed with the sensor surface on which the aminonaphthyridine dimer was immobilized. The response for the C[bond]C mismatch in 5'-GCC-3'/3'-CCG-5' was about 83 times stronger than that obtained for the fully matched duplex. The sensor successfully detects the C[bond]C mismatch at the concentration of 10 nM. SPR responses are proportional to the concentration of the C[bond]C mismatch in a range up to 200 nM. Aminonaphthyridine dimer could bind strongly to the C[bond]C mismatches having 10 possible flanking sequences with association constants in the order of 10(6) M(-1). The facile protonation of 2-aminonaphthyridine chromophore at pH 7 producing the hydrogen-bonding surface complementary to that of cytosine was most likely due to the remarkably high selectivity of 1 to the C[bond]C mismatch.

Base Pair Mismatch↗
PubMed2004

A nucleobase that releases reporter tags upon DNA oxidation.

We have developed a novel nucleosbase, edaG, that efficiently releases various reporter units upon one-electron oxidation. The edaG-selective degradation of ODNs was achieved by various mild oxidizing agents. The oxidant-dependent molecular releasing technique is quite useful not only for DNA-based drug releasing systems but also for the detection of long-range hole transport through DNA without time-consuming analysis.

Biosensing Techniques↗
PubMed2004

Detection of guanine-adenine mismatches by surface plasmon resonance sensor carrying naphthyridine-azaquinolone hybrid on the surface.

We have discovered a new molecule naphthyridine-azaquinolone hybrid (Npt-Azq) that strongly stabilized the guanine-adenine (G-A) mismatch in duplex DNA. In the presence of Npt-Azq, the melting temperature (T(m)) of 5'-d(CTA ACG GAA TG)-3'/3'-d(GAT TGA CTT AC)-5' containing a single G-A mismatch increased by 15.4 degrees C, whereas fully matched duplex increased its T(m) only by 2.2 degrees C. Npt-Azq was immobilized on the sensor surface for the surface plasmon resonance (SPR) assay to examine SPR detection of duplexes containing a G-A mismatch. Distinct SPR signals were observed when 27mer DNA containing a G-A mismatch was analyzed by the Npt-Azq immobilized sensor surfaces, whereas the signal of the fully matched duplex was approximately 6-fold weaker in intensity. The SPR signals for the G-A mismatch were proportional to the concentration of DNA in a range up to 1 microM, confirming that the SPR signal is in fact due to the binding of the G-A mismatch to Npt-Azq immobilized on the surface. Examination of all 16 G-A mismatches regarding the flanking sequence revealed that the sensor surface reported here is applicable to eight flanking sequences, covering 50% of all possible G-A mismatches.

Adenine↗
PubMed2004

The semi-quantitative comparison of oxidative stress mediated DNA single and double strand breaks using terminal deoxynucleotidyl transferase mediated end labeling combined with a slot blot technique.

The accumulation of DNA damages by environmental stresses is represented by the steady state level of single strand breaks (SSBs) and double strand breaks (DSBs). Terminal deoxynucleotidyl transferase (TdT) mediated end labeling is suitable in detecting DSBs, but is unsuitable for SSBs due to its catalyzing characteristics. However, the sensitivity of TdT to detect SSBs may be significantly improved by first denaturing the double strands and expose all the DNA nicks as potential substrates for TdT. By coupling DNA denaturation to slot blot southern hybridization, the authors demonstrate the sensitive detection of SSBs as well as DSBs in 20 ng DNA samples derived from a retinal pigment epithelial cell line treated with tert-butyl hydroperoxide. The signal intensity of denatured and TdT-treated DNA in slot blot hybridization correlated to the amount of SSBs calculated in an S1 nuclease digestion assay. The signal ratio between denatured and non-denatured DNA likely approximates the SSBs/DSBs ratio in genomic DNA. The combination of DNA denaturing, TdT treatment and slot blot hybridization could be a useful method to assess oxidative stress-induced DNA strand damages.

DNA↗