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Success of tumor photothermal immunotherapy requires a system that induces heat stress in cancer cells and enhances strong anti-tumor immune responses. Here, we designed a composite-type immunostimulatory DNA hydrogel consisting of a hexapod-like structured DNA (hexapodna) with CpG sequences and gold nanoparticles. Mixing of the properly designed hexapodna and oligodeoxynucleotide-modified gold nanoparticles resulted in the formation of composite-type gold nanoparticle-DNA hydrogels. Laser irradiation of the hydrogel resulted in the release of hexapodna, which efficiently stimulated immune cells to release proinflammatory cytokines. Then, EG7-OVA tumor-bearing mice received an intratumoral injection of a gold nanoparticle-DNA hydrogel, followed by laser irradiation at 780 nm. This treatment increased the local temperature and the mRNA expression of heat shock protein 70 in the tumor tissue, increased tumor-associated antigen-specific IgG levels in the serum, and induced tumor-associated antigen-specific interferon-γ production from splenocytes. Moreover, the treatment significantly retarded the tumor growth and extended the survival of the tumor-bearing mice.

We previously demonstrated that polypod-like structured DNA, or polypodna, constructed with three or more oligodeoxynucleotides (ODNs), is efficiently taken up by immune cells such as dendritic cells and macrophages, depending on its structural complexity. The ODNs comprising the polypodna should bend to form the polypod-like structure, and may do so by adopting either a bendtype conformation or a cross-type conformation. Here, we tried to elucidate the orientation and bending of ODNs in polypodnas using atomic force microscopy (AFM). We designed two types of pentapodnas (i.e., a polypodna with five pods) using 60- to 88-base ODNs, which were then immobilized on DNA origami frames. AFM imaging showed that the ODNs in the pentapodna adopted bend-type conformations. Tetrapodna and hexapodna also adopted bend-type conformations when they were immobilized on frames under unconstrained conditions. These findings provide useful information toward the coherent design of, and the structure–activity relationships for, a variety of DNA nanostructures.

Recent advances in 3D technology have been accompanied by increasing complaints of visual fatigue. The usual explanation for such fatigue is that accommodation and convergence are mismatched during stereoscopic vision. The aim of this study was to measure fixation distances between lens accommodation and convergence in young subjects while they viewed real objects and 3D video clips. Measurements were made using an original instrument. The 3D video clips were presented to subjects using a liquid crystal shutter glass system. The results showed that when viewing real objects, the diopter values of subjects' accommodation and convergence were similar and changed periodically. This measurement method was thus considered to be appropriate for the measurement of stereoscopic vision. We also investigated lens accommodation and convergence when subjects viewed 3D video clips. Both accommodation and convergence were found to move along with the virtual position of 3D video clips. Therefore, there was little discrepancy between accommodation and convergence during the viewing of 3D images.

The immunostimulatory activity of phosphodiester DNA containing unmethylated cytosine-guanine (CpG) dinucleotides can be increased by converting it into branched structures. These structures could be stabilized by ligating the 5′- and 3′-ends to form a closed loop with no terminal ends. To further increase the ability of branched DNA assemblies to induce cytokines, a series of tetrapod-like structured DNA, or tetrapodna, were designed using four 48-base oligodeoxynucleotides (ODNs). All these preparations were designed to have the same sequence except for the nick sites, and all the ODNs of one of the tetrapodna preparations were ligated to obtain circular tetrapodna. The nick site significantly influenced the formation of the structure and melting temperature (Tm), but hardly affected the enzymatic stability of the tetrapodna preparations. Circular tetrapodna exhibited a significantly higher Tm and was more stable in mouse serum than its non-ligated counterparts. The amounts of cytokines released from macrophage-like RAW264.7 cells or dendritic DC2.4 cells after addition of circular tetrapodna were not significantly higher than those after addition of other tetrapodna preparations under conditions when no serum was present. However, when serum was present, circular tetrapodna induced the greatest amount of tumor necrosis factor-α, indicating that circular tetrapodna is effective in inducing cytokines under conditions where DNA-degrading enzymes are present. The cellular association of tetrapodna preparations was almost unaffected by ligation of the terminal ends. These results indicate that circular tetrapodna with no terminal ends is more effective than its non-ligated counterparts in the presence of serum.

In this study, we invented a method to simultaneously measure lens accommodation and convergence used for natural and 3D vision. We found that there is almost never a difference between natural vision and 3D vision in terms of accommodation and convergence.

We explored in detail the relationship between the structure in aqueous solution and immunostimulatory activity of polypod-shaped DNAs, called polypodnas. The polypodnas were constructed using 3-6 oligodeoxynucleotides (ODNs) to obtain tri-, tetra-, penta-, and hexapodna, each of which had 3, 4, 5, and 6 arms made of double-stranded DNA, respectively. A highly potent immunostimulatory CpG sequence was included into each of the polypodnas. Synchrotron X-ray scattering analysis showed that the double-stranded DNA arms of all of the polypodnas adopted a B-form DNA conformation. The analysis also suggested that some nucleotides in the central parts of pentapodna and hexapodna did not form base pairs, whereas those of tripodna and tetrapodna all formed base pairs. This difference would occur because of an increase in steric hindrance and electrical repulsion with increasing number of arms. The pentapodna and hexapodna induced a large amount of tumor necrosis factor α-release from macrophage-like cells compared with the tripodna and tetrapodna, suggesting that the partly loosened DNA in polypodna with many arms is advantageous for exposing the immunostimulatory sequences of the polypodna.

We carried out synchrotron X-ray scattering experiments from four DNA supermolecules designed to form tetrapod shapes; these supermolecules had different sequences but identical numbers of total base pairs, and each contained an immunostimulatory CpG motif. We confirmed that the supermolecules did indeed form the expected tetrapod shape. The sample that had the largest radius of gyration (Rg) induced the most cytokine secretion from cultured immune cells. Structural analysis in combination with a rigid tetrapod model and an atomic scale DNA model revealed that the larger Rg can be ascribed to dissociation of the DNA double strands in the central connecting portion of the DNA tetrapod. This finding suggests that the biological activity is related to the ease with which single DNA strands can be formed.

1Department of Information Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8603, Japan 2Department of Information and Media Studies, Nagoya Bunri University, 365 Maeda, Inazawa-cho, Inazawa, Aichi 492-8520, Japan 3Graduate School of Engineering Human and Artificial Intelligent Systems, University of Fukui, 3-9-1 Bunkyo, Fukui 910-8507, Japan 4Facility of Home Ergonomics, Kobe Women’s University, 2-1 Aoyama, Higashisuma, Suma-ku, Kobe 654-8585, Japan ∗E-mail address: miyao@nagoya-u.jp

Equilibrium function in the cerebellum (vestibulo-cerebellar system) can deteriorate under the influence of alcohol. In the Romberg posture, the center of gravity, which was measured every 50 ms by stabilometry, appeared to shift with alcohol ingestion. In the previous study, a locus in the center of gravity (stabilogram) was converted to values of statistical indices such as area of sway, total locus length, and locus length per unit area, although these indices could not always distinguish between the statokinesigrams sampled from seven healthy young males in sober and intoxicated states. This measurement was made with an AMTI force plate. In this study, “translation error” was estimated in a d-dimensional embedding space in order to compare statokinesigrams recorded before and after the ingestion of doubly diluted brandy in 30 s (1 ≤ d ≤ 10). We succeeded in validating a stochastic differential equation as a mathematical model of the body sway. The randomness in the model was preserved after alcohol intake and significantly increased in the medial/lateral direction. Visual information referred by the postural control system when standing might be interfered by the effects of intoxication, which was regarded as disturbance. This method is considered to be useful to diagnose the disorders of the vestibulocerebellar system.

The immunostimulatory activity of phosphodiester DNA containing unmethylated cytosine-phosphate-guanine (CpG) dinucleotides, or CpG motifs, was significantly increased by the formation of Y-, X-, or dendrimer-like multibranched shape. These results suggest the possibility that the activity of CpG DNA is a function of the structural properties of branched DNA assemblies. To elucidate the relationship between them, we have designed and developed nanosized DNA assemblies in polypod-like structures (polypod-like structured DNA, or polypodna for short) using oligodeoxynucleotides (ODNs) containing CpG motifs and investigated their structural and immunological properties. Those assemblies consisting of three (tripodna) to eight (octapodna) ODNs were successfully obtained, but one consisting of 12 ODNs was not when 36-mer ODNs were annealed under physiological sodium chloride concentration. High-speed atomic force microscopy revealed that these assemblies were in polypod-like structures. The apparent size of the products was about 10 nm in diameter, and there was an increasing trend with an increase in ODN length or with the pod number. Circular dichroism spectral data showed that DNA in polypodna preparations were in the B-form. The melting temperature of polypodna decreased with increasing pod number. Each polypodna induced the secretion of tumor necrosis factor-α and interleukin-6 from macrophage-like RAW264.7 cells, with the greatest induction by those with hexa- and octapodna. Increasing the pod number increased the uptake by RAW264.7 cells but reduced the stability in serum. These results indicate that CpG DNA-containing polypodna preparations with six or more pods are a promising nanosized device with biodegradability and high immunostimulatory activity.

A few studies have evaluated swaying motion of the body in the seated posture; however, these studies did not evaluate motion sickness caused by watching movies, traveling, etc. This study aimed to analyze stabilograms recorded while the individuals were seated. Stabilometry was performed on 7 healthy male subjects (age, 21–22 years) in the seated position 5 min before, and at 5-min intervals from 5 to 60 min after alcohol load. Alcohol caused a decrease in the cerebellar equilibrium function that controls the vestibulospinal reflex. We adjusted the degree of simulative effect on the human equilibrium function. The stabilograms of the test subjects were analyzed according to the area of sway, total locus length, and total locus length per unit area. We found that significant alcohol-induced changes in the sway values in the seated posture were similar to those in the upright posture.