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Daisuke Sasaki

Publications and source records attributed to Daisuke Sasaki.

At least 19 recordsLinked to original sources
PubMed2006

Myocardial sarcomeres spontaneously oscillate with the period of heartbeat under physiological conditions.

During heartbeat, the repeated contractions of myocardium are induced by the oscillation of intracellular Ca(2+) concentration. On the other hand, when intermediately activated at a certain Ca(2+) concentration, cardiac myofibrils exhibit the spontaneous sarcomeric oscillation (Ca-SPOC) under steady ionic conditions. In the present study, we found that Ca-SPOC occurred over a wide range of Ca(2+) concentrations, including physiological contractile conditions, in skinned myocardium prepared from various animal species (rat, rabbit, pig, and cow). The period of sarcomeric oscillation fell within the same range as the period of heartbeat of each animal species. On the basis of these results we propose that the intrinsic auto-oscillatory property of sarcomeres (myofibrils) significantly contributes to myocardial beating in vivo.

Animals↗
PubMed2006

New method for the quantitative determination of major protein carbonyls, alpha-aminoadipic and gamma-glutamic semialdehydes: investigation of the formation mechanism and chemical nature in vitro and in vivo.

Alpha-aminoadipic semialdehyde (AAS) and gamma-glutamic semialdehyde (GGS) are identified as the major carbonyl products in oxidized proteins. To elucidate the formation pathway of AAS and GGS in vivo, we developed and validated a new quantification method. AAS and GGS in proteins were derivatized by reductive amination with NaCNBH(3) and p-aminobenzoic acid, a fluorescent reagent, followed by acid hydrolysis. It is noteworthy that the fluorescent derivatives were completely stable during acid hydrolysis. The present method permitted the specific, accurate, and sensitive quantification of both semialdehydes by fluorometric high-performance liquid chromatography. Analysis of proteins oxidized by various oxidation systems revealed that AAS and GGS are notably generated by the reaction of proteins with (*)OH, which is produced by metal-catalyzed oxidation (MCO). Furthermore, exposure of transferrin and human plasma to ascorbic acid and H(2)O(2) significantly promoted the formation of AAS and GGS in vitro, suggesting that both semialdehydes can be generated by MCO in vivo. We also demonstrated their generation through oxidative stress induced by acute iron overload in vivo. In this paper, we describe this analytical technique for simple and precise measurement of AAS and GGS and discuss their formation mechanism in vivo.

2-Aminoadipic Acid↗
PubMed2006

[Irritable bowel syndrome in the elderly].

Irritable bowel syndrome (IBS) is a very common gastrointestinal disorder. The prevalence of IBS is about 10-15% of the general population. Epidemiological studies suggested that the prevalence of IBS decreased with age, but IBS remains an important gastrointestinal illness in the aged. But there has been very few research examining on IBS in elderly. Whether advancing age impacts on IBS is largely unknown and how the disorder manifest in the elderly remains unclear. Aging is connected with an increasing prevalence of many chronic neurological difficulties, cardiovascular diseases and mental disabilities. The management of the IBS needs to take the age-related issues into account in elderly. Clinical therapeutic trials should be undertaken in elderly people to ascertain treatment.

Adult↗
PubMed2005

Overexpression of Rad23 confers resistance to methylmercury in saccharomyces cerevisiae via inhibition of the degradation of ubiquitinated proteins.

We report here that overexpression of Rad23, a protein related to the ubiquitin-proteasome system, renders yeast cells resistant to methylmercury. Rad23 has three domains: two ubiquitin-associated (UBA) domains that bind to the multiubiquitin chain of ubiquitinated proteins and a single ubiquitin-like (UbL) domain that binds to proteasomes. To examine the mechanism of acquisition of methylmercury resistance that is induced by overexpression of Rad23, we expressed variants of Rad23 in which one or the other of the two types of domain was defective in yeast cells. In cells that overexpressed full-length intact Rad23, we detected elevated levels of intracellular ubiquitinated proteins, and the cells were resistant to methylmercury. In contrast, cells that overexpressed Rad23 with a defective UBA domain were not resistant to methylmercury and contained control levels of ubiquitinated proteins. Yeast cells that overexpressed Rad23 with a defective UbL domain exhibited enhanced resistance to methylmercury and contained even higher levels of ubiquitinated proteins than cells that overexpressed intact full-length Rad23. Rad23 is known to have two mutually contradictory functions. It suppresses the degradation of ubiquitinated proteins by proteasomes via a mechanism mediated by the UBA domains, and it enhances the degradation of ubiquitinated proteins via a mechanism that is mediated by the UbL domain. Therefore, our findings suggest that Rad23 might induce resistance to methylmercury in yeast cells by suppressing the degradation of proteins that reduce the toxicity of methylmercury via a UBA domain-mediated mechanism.

Base Sequence↗
PubMed2005

Auto-oscillations of skinned myocardium correlating with heartbeat.

Skinned myocardium (or myofibrils) exhibits auto-oscillations of sarcomere length and developed force called SPOC (SPontaneousOscillatoryContraction) under partial activation conditions. In SPOC, each sarcomere repeats the cycle of slow shortening and rapid lengthening, and the lengthening phase propagates sequentially to the adjacent sarcomeres in waves (SPOC wave). In this study, we analyzed the sarcomeric oscillation in SPOC in skinned myocardium of various animal species (rat, rabbit, dog, pig, and cow) with different heart rates. The period of oscillation, the sarcomere shortening velocity, and the velocity of SPOC wave, strongly correlated with the resting heart rate of the animal species. The shortening velocity in particular was proportional to the resting heart rate. We then examined the motile activity of each cardiac myosin by an in vitro motility assay. The sliding velocity of actin filaments, which is an index of the motile activity of myosin, also correlated with the resting heart rate but the relationship was not proportional. As a result, the ratio of sarcomere shortening velocity in SPOC to the sliding velocity of actin filaments was not constant but became higher with a higher heart rate. This suggests that the sarcomere shortening velocity in SPOC is modulated by some additional factors besides the motile activity of myosin, resulting in the proportional relationship between the shortening velocity of the sarcomere and the resting heart rate.

Actin Cytoskeleton↗
PubMed2005

Identification of a novel conjugate in human urine: bile acid acyl galactosides.

We report a novel conjugate, bile acid acyl galactosides, which exist in the urine of healthy volunteers. To identify the two unknown peaks obtained in urine specimens from healthy subjects, the specimens were subjected to solid phase extraction and then to liquid chromatographic separation. The eluate corresponding to the unknown peaks on the chromatogram was collected. Following alkaline hydrolysis and liquid chromatography (LC)/electrospray ionization (ESI)-mass spectrometric (MS) analysis, cholic acid (CA) and deoxycholic acid (DCA) were identified as liberated bile acids. When a portion of the alkaline hydrolyzate was subjected to a derivatization reaction with 1-phenyl-3-methyl-5-pyrazolone, a derivative of galactose was detected by LC/ESI-MS. Finally, the liquid chromatographic and mass spectrometric properties of these unknown compounds in urine specimens were compared to those of authentic specimens and the structures were confirmed as CA 24-galactoside and DCA 24-galactoside. These results strongly imply that bile acid 24-galactosides, a novel conjugate, were synthesized in the human body.

Adult↗
PubMed2005

Formation of alpha-aminoadipic and gamma-glutamic semialdehydes in proteins by the maillard reaction.

Recent research has demonstrated that nonenzymatic glycation (the Maillard reaction) lead to the formation of carbonyl groups and advanced glycation end products (AGEs) in proteins. Such oxidative modifications are a major contributing factor to diabetic complications and aging. alpha-Aminoadipic semialdehyde (AAS) and gamma-glutamic semialdehyde (GGS) have been identified as the major carbonyl products in oxidized proteins both in vitro and in vivo. AAS is an oxidative deamination product of lysine residue, while GGS originates from arginine and proline residues. To evaluate oxidative damage to proteins by the Maillard reaction, we developed a method of detecting AAS and GGS by high-performance liquid chromatography (HPLC). The aldehydic residues in proteins were derivatized by reductive amination with NaCNBH3 and p-aminobenzoic acid (ABA), a fluorescence regent. After acid hydrolysis of the ABA-derivatized protein, ABA-AAS and ABA-GGS were measured by fluorometric HPLC. Thus, AAS and GGS could be detected in various proteins such as human plasma protein using the present method. Accumulation of both aldehydic residues was observed in oxidized proteins by reactive oxygen species. Furthermore, AAS and GGS were markedly formed in the incubation of BSA with ascorbic acid. The formation of both aldehydic residues was also observed in the incubation of BSA with 100 mM glucose or 1.0 mM methylglyoxal in the absence and presence of 100 microM Fe3+ for 2 weeks. These results suggest that the Maillard reaction can contribute to the formation of AAS and GGS in vivo.

2-Aminoadipic Acid↗
PubMed2004

Asymmetric total syntheses of (+)-mycoepoxydiene and related natural product (-)-1893A: application of one-pot ring-opening/cross/ring-closing metathesis to construct their 9-oxabicyclo[4.2.1]nona-2,4-diene skeleton.

The total syntheses of (+)-mycoepoxydiene and (-)-1893A have been completed. The present synthetic strategy features the use of one-pot ring-opening/cross metathesis (ROM/CM) followed by a ring-closing metathesis (RCM) reaction, allowing for the concise construction of the 9-oxabicyclo[4.2.1]nona-2,4-diene framework from a 7-oxabicyclo[2.2.1]hept-2-ene derivative and 1,3-butadiene. The sequential metathesis product was converted into (+)-mycoepoxydiene through the oxidative rearrangement of a furfuryl alcohol to a pyranone, thereby establishing its absolute stereochemistry. From the common intermediate, a structurally related natural product (-)-1893A was also synthesized via the vinylogous aldol reaction.

Bridged-Ring Compounds↗
PubMed2004

Libraries enriched for alternatively spliced exons reveal splicing patterns in melanocytes and melanomas.

It is becoming increasingly clear that alternative splicing enables the complex development and homeostasis of higher organisms. To gain a better understanding of how splicing contributes to regulatory pathways, we have developed an alternative splicing library approach for the identification of alternatively spliced exons and their flanking regions by alternative splicing sequence enriched tags sequencing. Here, we have applied our approach to mouse melan-c melanocyte and B16-F10Y melanoma cell lines, in which 5,401 genes were found to be alternatively spliced. These genes include those encoding important regulatory factors such as cyclin D2, Ilk, MAPK12, MAPK14, RAB4, melastatin 1 and previously unidentified splicing events for 436 genes. Real-time PCR further identified cell line-specific exons for Tmc6, Abi1, Sorbs1, Ndel1 and Snx16. Thus, the ASL approach proved effective in identifying splicing events, which suggest that alternative splicing is important in melanoma development.

Alternative Splicing↗
PubMed2004

Absolute expression values for mouse transcripts: re-annotation of the READ expression database by the use of CAGE and EST sequence tags.

The RIKEN expression array database (READ) provides comprehensive gene expression data for the mouse, which were obtained as relative values from microarray double-staining experiments with E17.5 mRNA as common reference. To assign absolute expression values for mouse transcripts within READ, we applied the E17.5 reference sample to CAGE (cap analysis of gene expression) and expressed sequence tag (EST) high-throughput tag sequencing. Newly assigned values within the READ database were validated by comparison to expression data from serial analysis of gene expression, CAGE and EST experiments. These experiments confirmed the great significance of the absolute expression values within the improved READ database. The new Absolute READ database on absolute expression data is available under.

Animals↗
PubMed2004

Minimization of photooxidative insult to calf lens protein irradiated with near UV-light in the presence of pigmented glucosides derived from human lens protein.

An aqueous solution of a pigmented glucoside associated with human lens protein, 2-amino-3-hydroxyacetophenone-O-beta-D-glucoside (AHA-Glc), was irradiated with near UV-light. The near UV-irradiated glucoside was shown to generate a much lower level of active species of molecular oxygen as compared to the level of the active species generated from the irradiated aglycon, 2-amino-3-hydroxyacetophenone (AHA). This result suggests that the glycon of the glucoside is functioning as a scavenger for active oxygen generated from the aglycon of the irradiated glucoside. Superoxide dismutase (SOD) was shown to remove a large portion of the active oxygen generated from the irradiated AHA, so the bulk of the active species generated is assumed to be superoxide anion. The small portion of active oxygen remains after removal of superoxide anion may include singlet oxygen. The photooxidation of tryptophan residues of calf alpha-crystallin irradiated with near UV-light in the presence of AHA-Glc or AHA was investigated to confirm the role that the glycon plays in diminution of the active species of oxygen generated through the photosensitized aglycon of the glucoside. A decrease with time in the fluorescence intensity of the tryptophan residues irradiated with AHA-Glc was shown to be much slower as compared to the time-dependent decrease with AHA, indicating that the photooxidation proceeds with an increase in accumulation of active oxygen generated through the aglycon and that the glycon of the glucoside deactivates the active species as it is formed in the photodynamic process. Similar effects have also been observed in calf lens crystallin irradiated with either 3-hydroxykynurenine-O-beta-D-glucoside (HKN-Glc) or 3-hydroxykynurenine (HKN). Furthermore, effects of near UV-irradiation on calf lens soluble protein in the presence of AHA-Glc or AHA were studied by monitoring changes in the SDS-PAGE profile of the irradiated protein. Near UV-irradiation with AHA-Glc was shown to bring about a slight change in cross-linking of the polypeptides, while irradiation with AHA was shown to give rise to a significant increase in cross-linking of the polypeptides. In conclusion, pigmented glucoside associated with human lens protein is not only a photosensitizer for near UV-light but also an anti-photooxidant to deactivate active oxygen formed through the in situ photosensitizer, in order that photooxidative insults to lens proteins may be minimized during aging.

Animals↗
PubMed2004

Stimulation of in vitro angiogenesis by nitric oxide through the induction of transcription factor ETS-1.

The aim of this study was to examine whether transcription factor ETS-1, which is responsible for the expression of metalloproteinases and integrin beta(3), is implicated in the induction of nitric oxide (NO)-induced angiogenesis. Bovine aortic endothelial cells were cultured on type I collagen gel to measure the length of the formed tube-like structure, which is a marker for in vitro angiogenesis. The addition of S-nitroso-N-acetylpenicillamine (SNAP), an NO-donor, to confluent endothelial cells stimulated the formation of the tube-like structure, with disappearance of covered endothelial cell monolayers. Another NO-donor 2,2'-(hydroxynitrosohydrazono)bis-ethanamine (NOC 18) also induced the formation of the tube-like structure. In contrast to the induction of the formation of the tube-like structure by SNAP, it reduced cell proliferation. SNAP and NOC 18 also increased the expression of the ets-1 mRNA level in a concentration-dependent manner. The maximum expression was observed at 2h after their addition. Moreover, the SNAP-induced in vitro angiogenesis, ets-1 mRNA expression and ETS-1 protein expression were strongly reduced by the treatment with ets-1 antisense oligonucleotide. These results strongly suggest that NO stimulates in vitro angiogenesis through the induction of ETS-1 expression. NO appears to stimulate endothelial cell differentiation to the angiogenic phenotype via the induction of ETS-1 transcription factor.

Animals↗
PubMed2003

Cap analysis gene expression for high-throughput analysis of transcriptional starting point and identification of promoter usage.

We introduce cap analysis gene expression (CAGE), which is based on preparation and sequencing of concatamers of DNA tags deriving from the initial 20 nucleotides from 5' end mRNAs. CAGE allows high-throughout gene expression analysis and the profiling of transcriptional start points (TSP), including promoter usage analysis. By analyzing four libraries (brain, cortex, hippocampus, and cerebellum), we redefined more accurately the TSPs of 11-27% of the analyzed transcriptional units that were hit. The frequency of CAGE tags correlates well with results from other analyses, such as serial analysis of gene expression, and furthermore maps the TSPs more accurately, including in tissue-specific cases. The high-throughput nature of this technology paves the way for understanding gene networks via correlation of promoter usage and gene transcriptional factor expression.

Animals↗
PubMed2003

Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice.

We collected and completely sequenced 28,469 full-length complementary DNA clones from Oryza sativa L. ssp. japonica cv. Nipponbare. Through homology searches of publicly available sequence data, we assigned tentative protein functions to 21,596 clones (75.86%). Mapping of the cDNA clones to genomic DNA revealed that there are 19,000 to 20,500 transcription units in the rice genome. Protein informatics analysis against the InterPro database revealed the existence of proteins presented in rice but not in Arabidopsis. Sixty-four percent of our cDNAs are homologous to Arabidopsis proteins.

Alternative Splicing↗
PubMed2003

DNA cleavage characteristics of non-protein enediyne antibiotic N1999A2.

N1999A2 (NA2) is a new non-protein antitumor antibiotic that contains a stable 9-membered ring enediyne chromophore similar to a neocarzinostatin chromophore (NCS-chr). We have compared DNA cleavage reactions between NA2 and NCS-chr, and also clarified some characteristics of DNA strand scission by NA2. It was found that: (1) NA2 is considerably stable in nature, (2) the compound intercalates into base pairs of a DNA minor groove and decreases its base-attacking frequency in the order of T>A>> C>G, (3) the base-sequence specificity 5(')-GGT/3(')-CCA presented by NA2 is significantly related to recognition of the base pair with the naphthoate moiety, and (4) the different cleavage property between NCS-chr and NA2 is associated with the presence or absence of an aminoglycoside residue. Based on the results of the site-specific cleavage by NA2 for certain bulged DNAs and a fluorescence study of NA2-DNA oligomer complexes, the DNA interaction mode of NA2 has also been examined. These results provide important information to design a new enediyne molecule for a DNA target.

Anti-Bacterial Agents↗