Biomedical subjects

Yuko Yoshikawa

Publications and source records attributed to Yuko Yoshikawa.

6 recordsLinked to original sources

PubMed2006

High vaccine efficacy against shigellosis of recombinant noninvasive Shigella mutant that expresses Yersinia invasin.

Live attenuated Shigella vaccines elicit protective immune responses, but involve a potential risk of inducing a strong inflammatory reaction. The bacterial invasiveness that is crucial for Ag delivery causes inflammatory destruction of infected epithelial cells and proinflammatory cell death of infected macrophages. In this study, the noninvasive Shigella mutant DeltaipaB was equipped with Yersinia invasin protein, which has been shown to mediate bacterial invasion and targeting to M cells located in follicle-associated epithelium. Invasin-expressing DeltaipaB (DeltaipaB/inv) was internalized into epithelial cells and retained in the intraphagosomal space. DeltaipaB/inv did not induce necrotic cell death of infected macrophages nor cause symptomatic damage after intranasal vaccination of mice. DeltaipaB/inv was safer and more effective than the conventional live vaccine, DeltavirG. Infection by DeltaipaB/inv caused polymorphonuclear neutrophil infiltration in the lung, but did not induce production of large amounts of proinflammatory cytokines. We concluded that the low experimental morbidity and high vaccine efficacy of DeltaipaB/inv are primarily based on high protective immune responses, which may be enhanced by the polymorphonuclear neutrophil infiltration unaccompanied by tissue injury.

Adhesins, Bacterial↗

PubMed2005

Protective effect of vitamin C against double-strand breaks in reconstituted chromatin visualized by single-molecule observation.

Direct attack to genomic DNA by reactive oxygen species causes various types of lesions, including base modifications and strand breaks. The most significant lesion is considered to be an unrepaired double-strand break that can lead to fatal cell damage. We directly observed double-strand breaks of DNA in reconstituted chromatin stained by a fluorescent cyanine dye, YOYO (quinolinium, 1,1'-[1,3- propanediylbis[(dimethyliminio)-3,1- propanediyl]]bis[4-[(3-methyl-2(3H)-benzoxazolylidene)methyl]]-, tetraiodide), in solution, where YOYO is known to have the ability to photo-cleave DNAs by generating reactive oxygen species. Reconstituted chromatin was assembled from large circular DNA (106 kbp) with core histone proteins. We also investigated the effect of vitamin C (ascorbic acid) on preventing photo-induced double-strand breaks in a quantitative manner. We found that DNA is protected against double-strand breaks by the addition of ascorbic acid, and this protective effect is dose dependent. The effective kinetic constant of the breakage reaction in the presence of 5 mM ascorbic acid is 20 times lower than that in the absence of ascorbic acid. This protective effect of ascorbic acid in reconstituted chromatin is discussed in relation to the highly compacted polynucleosomal structure. The results highlight the fact that single-molecule observation is a useful tool for studying double-strand breaks in giant DNA and chromatin.

Ascorbic Acid↗

PubMed2004

Double-strand break of giant DNA: protection by glucosyl-hesperidin as evidenced through direct observation on individual DNA molecules.

We have performed a real-time observation of photo-induced breakage on individual long DNA molecules stained by a cyanine dye, YOYO, under the intense illumination (lambda=450-490 nm) in solution. It was shown that the double-strand breakage is accelerated by Fenton's reagent (Fe2+/H2O2). In addition, it was found that the breakage reaction is protected in the presence of a water-soluble flavonoid, glucosyl-hesperidin. To interpret the kinetic process of the double-strand breakage reaction, a two-step mechanism is proposed: under light illumination on giant DNA molecules, single-strand breaks are generated to induce nicks in a stochastic manner, and then reactive oxygen attacks these nicks causing the double-strand break.

Bacteriophage T4↗

PubMed2004

Folding transition of DNA depending on ionic environments.

We examined the effect of monovalent salts, NaCl and KCl, on spermidine-induced DNA compaction using fluorescence microscopy. By increasing concentrations of salts, unfolding process of compacted DNAs were observed. It was also shown that NaCl is more effective for unfolding compacted DNAs than KCl.

Bacteriophage T4↗

PubMed2003

Ascorbic acid induces a marked conformational change in long duplex DNA.

Ascorbic acid is often regarded as an antioxidant in vivo, where it protects against cancer by scavenging DNA-damaging reactive oxygen species. However, the detailed mechanism of the action of ascorbic acid on genetic DNA is still unclear. We examined the effect of ascorbic acid on the higher-order structure of DNA through real-time observation by fluorescence microscopy. We found that ascorbic acid generates a pearling structure in single giant DNA molecules, with elongated and compact regions coexisting along a molecular chain. Results from electron microscopy and atomic force microscopy indicate that the compact regions assume a loosely packed conformation. A possible mechanism for the induction of this conformational change is discussed in relation to the interplay between the higher-order and second-order structures of DNA.

Ascorbic Acid↗

PubMed2002

Folding transition of large DNA completely inhibits the action of a restriction endonuclease as revealed by single-chain observation.

The biochemical characteristics of lambda DNA chains in folded/unfolded states upon cleavage by the restriction enzyme ApaLI were investigated in the presence of spermine. These characteristics of DNA chains depending on their higher-order structure were studied at the single-molecule level using fluorescence microscopy. With a low concentration of spermine, lambda DNA takes a random coiled conformation and allows digestion by the enzyme, while under a high concentration of spermine, lambda DNA takes a compact folded structure and inhibits such attack. Together with comparative experiments on short oligomeric DNA, our results suggest that the transition in the higher-order structure causes on/off-type switching of sensitivity to the enzyme.

Base Sequence↗