Field-Induced Rocking-Curve Effects in Attosecond Electron Diffraction.

Recent advances in electron microscopy trigger the question of whether attosecond electron diffraction can resolve atomic-scale electron dynamics in crystalline materials in space and time. Here, we explore the ultrafast dynamics of the relevant electron-lattice scattering process. We drive a single-crystalline silicon membrane with the optical cycles of near-infrared laser light and use phase-locked attosecond electron pulses to produce electron diffraction patterns as a function of delay. For all Bragg spots, we observe time-dependent intensity changes and position shifts that are correlated with a time shift of 0.5-1.2 fs. For single-cycle excitation pulses with strong peak intensity, the correlations become nonlinear. The origins of these effects are local and integrated beam deflections by the optical electric and magnetic fields at the crystal membrane. Those deflections modify the diffraction intensities in addition to the atomic structure factor dynamics by time-dependent rocking-curve effects. However, the measured time delays and symmetries allow one to disentangle both effects. Future attosecond electron diffraction and microscopy experiments need to be based on these results.

Cell-Based Biohybrid Sensor Device for Chemical Source Direction Estimation.

This paper describes a method to estimate the direction from which the signal molecule reaches the sensor by using living cells. In this context, biohybrid sensors that utilize a sophisticated sensing system of cells can potentially offer high levels of chemical-detection sensitivity and selectivity. However, biohybrid-sensor-based chemical-source-direction estimation has not received research attention because the cellular response to chemicals has not been examined in the context of directional information. In our approach, we fabricated a device that can limit the interface between the cell-laden hydrogel and the chemical solution of interest to enhance the time difference over which the chemical solution reaches the cells. Chemical detection by cells that express specific receptors is reflected as the fluorescence of the calcium indicator within the cells. Our device has eight chambers that each house 3D cell-laden collagen hydrogels facing circularly outward. The device also works as a cover to prevent chemicals from permeating the hydrogel from above. In our study, by observing the time course of the fluorescence emission of each chamber, we were able to successfully estimate the chemical-source direction within an error range of 7-13°. Our results suggest that a combination of microstructure devices embedded with living cells can be used to exploit cell functionalities to yield chemical-source directional information.

Evaluation of velopharyngeal function using high-speed cine-magnetic resonance imaging based on T2-weighted sequences: a preliminary study.

The objective was to introduce a new technique for visualizing the three-dimensional (3D) movements of velopharyngeal-related muscles using high-speed cine-magnetic resonance imaging (MRI) based on T2-weighted sequences. The evaluation of phonation- and water swallowing-related events was performed in 11 healthy subjects. Specifically, whether cine-MRI could precisely visualize normal velopharyngeal function during these two events was examined. The 3D movements of the soft palate, superior pharyngeal constrictor muscles, and levator veli palatini muscles were visualized in all 11 subjects. A noteworthy finding was that the magnetic resonance signals of the superior constrictor pharyngeal muscles and the levator veli palatini muscles were significantly higher during phonation and during water swallowing than at rest. This initial study suggests that the 3D movements of velopharyngeal-related muscles can be successfully and precisely visualized without side effects. The magnetic resonance signal changes seen in the superior pharyngeal constrictor and levator veli palatini muscles using the technique described here should be useful to develop better methods of evaluation of velopharyngeal function.

Predicting the therapeutic effect of carbamazepine in trigeminal neuralgia by analysis of neurovascular compression utilizing magnetic resonance cisternography.

The aim of this study was to determine whether the evaluation of neurovascular compression (NVC) using new criteria on magnetic resonance (MR) cisternography improves the prediction of the curative effects of carbamazepine (CBZ) in trigeminal neuralgia (TN). In this study, a new analysis of NVC using MR cisternography was performed retrospectively for 280 patients with clinical signs and symptoms suggesting TN. The new analysis examined whether the site of the NVC was less than 3mm from the edge of the exit point and within the first third of the root entry zone on MR cisternography. Prediction of the curative effects of CBZ using the new evaluation of distance to the NVC improved on the predictive ability of the previous method of calculating the NVC volume (Mantel-Haenszel coefficient, P<0.01). In particular, initial treatment with CBZ 100mg/day for 2 weeks appeared more effective for patients with NVC volumes of ≤5mm3 plus a distance to the NVC of ≤3mm, than for those with NVC volumes >5mm3. The evaluation of NVC on MR cisternography using this new approach appears to be more useful than the previous method for predicting the initial treatment response in patients with TN.

Identification of specifically reduced Th2 cell subsets in allergic rhinitis patients after sublingual immunotherapy.

BACKGROUND: Although Th2 cells are well known to play important roles in allergic diseases including allergic rhinitis (AR), the factors that induce and sustain the pathogenesis of AR remain unclear. The recent development of sublingual immunotherapy (SLIT) is expected to allow changes to the underlying pathogenesis of AR. However, which Th2 cell subsets are important in house dust mite-induced AR (HDM-AR), the influence of SLIT on the pathogenic Th2 cells, and the association of Th2 cell subsets with SLIT efficacy have not been clarified. METHODS: The cytokine production and frequency of HDM-reactive T-cell subsets in peripheral blood mononuclear cells (PBMCs) were evaluated using flow cytometry in 89 HDM-AR patients (placebo [n = 43] and HDM 300 IR [n = 46]) who participated in a placebo-controlled study of SLIT with HDM tablets. All patients provided samples both before treatment as a baseline and at the end of the 52-week study. The PBMCs were stained with CellTrace™ Violet (CTV) before culture with HDM extract, and HDM-reactive T cells were detected as the proliferated cells with diminished CTV. RESULTS: HDM-reactive IL-5+ IL-13+ CD27- CD161+ CD4+ cells and ST2+ CD45RO+ CD4+ cells were observed in the peripheral blood from each patient with HDM-AR; these cells significantly decreased after SLIT in the group treated with active tablets. HDM-reactive ST2+ CD45RO+ CD4+ cells were significantly lower in active-responders. CONCLUSION: Allergen-reactive ST2+ CD45RO+ CD4+ cells or those combined with IL-5+ IL-13+ CD27- CD161+ CD4+ cells may be useful as markers indicating the successful treatment of SLIT. These cells may play a crucial role in the pathogenesis of AR as pathogenic memory Th2 cells.

Pathogenicity of memory Th2 cells is linked to stage of allergic rhinitis.

BACKGROUND: Allergic rhinitis (AR) consists of three developmental stages that are based on the presence/absence of antigen-specific IgE and symptoms. The pathogenic Th2 (Tpath2) cells constitute a population of Th2 cells with additional potentially pathogenic characteristics. We examined the relationship between Tpath2 cells and the stages of allergic rhinitis by focusing on ST2, which is an IL-33 receptor. METHODS: Patients with Japanese cedar pollen-induced AR (JCP-AR) and healthy volunteers were divided into "nonsensitized," "asymptomatic sensitized (AS)," and "JCP-AR" groups. We analyzed the ST2 expression and the Th2 function of cultured CD4+ T cells. Next, we observed the progress of patients in the AS stage around the time of seasonal pollen dispersal, with the characteristics of Th2 cells. RESULTS: The ST2 expression of T cells was only upregulated in the AR group. The production of IL-4 and IL-13 was found in CD4+ T cells obtained from AS by stimulation with JCP, but reactivity to IL-33 was not observed. Although IL-33 did not induce the elevation of IL-4 production in the JCP-AR group, IL-33 substantially increased the production of IL-5 and IL-13 in comparison with antigen stimulation alone. In newly afflicted patients, the increased expression of ST2 and elevated reactivity to IL-33 was observed, even before the pollen dispersal season. CONCLUSIONS: Our study demonstrated that the pathogenicity of memory Th2 cells is linked to sensitization and the stage of allergic rhinitis. Therefore, Tpath2 cells may provide useful insights into the mechanism of the onset and progression of allergic rhinitis.

Importance of absorbable surgical sutures for the prevention of stitch abscess after surgery in patients with oral squamous cell carcinoma.

BACKGROUND: To elucidate the significance of absorbable surgical sutures in the occurrence of stitch abscess after surgery in patients with oral squamous cell carcinoma (SCC). MATERIAL AND METHODS: The subjects were 251 patients who underwent excision and/or reconstruction and/or neck dissection for oral SCC using absorbable surgical sutures. Detection rates and characteristics of patients with stitch abscess were retrospectively evaluated by comparing between our present and previous data. RESULTS: There was only one stitch abscess among the 251 patients. A significant difference in the incidence of stitch abscess was found between the present data and our previous data. Of course, no significant correlations were found between the occurrence of stitch abscess using absorbable surgical sutures and the various factors seen in our previous analysis. CONCLUSIONS: A complete switch of surgical sutures from silk to absorbable surgical sutures is needed for surgery in patients with oral SCC.

Expression of hOvol2 in the XY body of human spermatocytes.

Male infertility is common at infertile clinics, and 10-20% of infertile males are azoospermic. Non-obstructive azoospermia is a complex multifactorial disease, and the process of spermatogenesis remains largely unknown. Ovol1 and Ovol2, a family of zinc finger transcription factors, are expressed in spermatocytes at the pachytene stage and are suggested to be critical regulators of pachytene progression in male germ cells. In this study, we examined the expression of human Ovol2 (hOvol2) in the seminiferous tubes of patients subjected to testicular sperm extraction. We first cloned hOvol1 and hOvol2 from the testis of one of the patients and found no alteration in these nucleotide sequences of this patient. While hOvol1 and hOvol2 were detected by RT-PCR in the testis of patients capable of spermatogenesis, they were not detected in those with Sertoli cell-only syndrome. We recently succeeded in preparing anti-Ovol2 antibody by immunising rats with recombinant mouse Ovol2 (mOvol2) and confirmed the specificity and cross-reactivity of this antibody with hOvol2 in cells transfected with hOvol1 or hOvol2 cDNA. hOvol2 expression was restricted to the XY body of spermatocytes at the pachytene stage. This study demonstrates that hOvol2 is expressed in germ cells and may be involved in spermatogenesis.

Spatial and Temporal Brain Responses to Noxious Heat Thermal Stimuli in Burning Mouth Syndrome.

Burning mouth syndrome (BMS) is an idiopathic orofacial pain condition. Although the pathophysiology of BMS is not clearly understood, central and peripheral neuropathic mechanisms are thought to be involved. The authors compared brain response to noxious heat stimuli in 16 right-handed women with primary BMS and 15 sex- and age-matched right-handed healthy female controls. A thermal stimulus sequence of 32 °C to 40 °C to 32 °C to 49 °C was repeated 4 times in a cycle. Warm and noxious heat stimuli were delivered with a Peltier thermode placed on the right palm or right lower lip for 32 s each in a session. Functional magnetic resonance imaging data were obtained by recording echoplanar images with a block design. Statistical Parametric Mapping 8 software was used to analyze the data. Patients and controls both reported feeling more pain during palm stimulation than during lip stimulation. Repetition of noxious heat stimulus on the lower lip but not on the palm induced habituation in brain activity in the cingulate cortex without reduction in pain perception. Multiple regression analysis revealed a correlation between perceived pain intensity and suppression of brain activity in the anterior cingulate cortex when the repeated thermal sequence was applied at the lower lip. Furthermore, the response of the parahippocampal area differed in BMS patients and controls when the same repeated thermal sequence was applied at the palm. The authors' findings indicate that BMS patients show specific brain responses due to impaired function of the central and peripheral nervous systems (clinical trial registration: UMIN000015002).

Human induced pluripotent stem cell-derived fiber-shaped cardiac tissue on a chip.

We propose a method for the production of a fiber-shaped three-dimensional (3D) cellular construct of human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) for the quantification of the contractile force. By culturing the cardiomyocytes in a patterned hydrogel structure with fixed edges, we succeeded in fabricating hiPS-CM fibers with aligned cardiomyocytes. The fiber generated contractile force along the fiber direction due to the hiPS-CM alignment, and we were able to measure its contractile force accurately. Furthermore, to demonstrate the drug reactivity of hiPS-CM fibers, the changes in the contractile frequency and force following treatment with isoproterenol and propranolol were observed. We believe that hiPS-CM fibers will be a useful tool for pharmacokinetic analyses during drug development.

Alterations in 18F-FDG accumulation into neck-related muscles after neck dissection for patients with oral cancers.

BACKGROUND: 18F-fluoro-2-deoxy-D-glucose (18F-FDG) accumulations are commonly seen in the neck-related muscles of the surgical and non-surgical sides after surgery with neck dissection (ND) for oral cancers, which leads to radiologists having difficulty in diagnosing the lesions. To examine the alterations in 18F-FDG accumulation in neck-related muscles of patients after ND for oral cancer. MATERIAL AND METHODS: 18F-FDG accumulations on positron emission tomography (PET)-computed tomography (CT) in neck-related muscles were retrospectively analyzed after surgical dissection of cervical lymph nodes in oral cancers. RESULTS: According to the extent of ND of cervical lymph nodes, the rate of patients with 18F-FDG-PET-positive areas increased in the trapezius, sternocleidomastoid, and posterior neck muscles of the surgical and/or non-surgical sides. In addition, SUVmax of 18F-FDG-PET-positive areas in the trapezius and sternocleidomastoid muscles were increased according to the extent of the ND. CONCLUSIONS: In evaluating 18F-FDG accumulations after ND for oral cancers, we should pay attention to the 18F-FDG distributions in neck-related muscles including the non-surgical side as false-positive findings.

Sublingual immunotherapy for allergic rhinitis: subjective versus objective tools to evaluate its success.

BACKGROUND: Biomarkers that enable objective evaluation of the clinical effects of immunotherapy for allergic rhinitis have yet to be identified. METHODS: This study included 40 patients who were enrolled in a large randomized, double-blind, placebo-controlled, multicenter study examining the efficacy of sublingual immunotherapy (SLIT) using Japanese cedar (JC) pollen extract during two consecutive pollen seasons from 2010 to 2012. Based on changes in total nasal symptom medication score, patients in the SLIT and placebo groups were subdivided into two subgroups: good responders and poor responders. The levels of JC pollen-specific IL-10+Foxp3+ cells and specific Th2 cytokine-producing cells were measured and the association with the efficacy of SLIT was analysed. RESULTS: The total nasal symptom medication score was significantly lower in the SLIT group compared with the placebo group. The number of JC pollen-specific Th2 cytokine-producing cells increased during the pollen season in the placebo group and in poor responders in the SLIT group; however, the increases were inhibited in the good responders in the SLIT group. The number of JC pollen-specific IL-10+Foxp3+ cells increased only in these good responders. CONCLUSIONS: Changes in levels of allergen-specific Th2 cytokine-producing cells and IL-10+Foxp3+ cells could be objective biomarkers for SLIT.

Ethnic admixture affects diabetes risk in native Hawaiians: the Multiethnic Cohort.

BACKGROUND/OBJECTIVES: Obesity and diabetes rates are high in Native Hawaiians (NHs) who commonly have mixed ancestries. People of Asian ancestry experience a high risk of type 2 diabetes despite the relatively low body weight. We evaluated the impact of ethnic admixture on diabetes risk among NHs in the Multiethnic Cohort (MEC). SUBJECTS/METHODS: On the basis of self-reports, 11 521 eligible men and women were categorized into NH/white, NH/other, NH alone, NH/Asian and the most common three ancestry admixture, NH/Chinese/white. Cox proportional hazard ratios (HRs) and 95% confidence intervals (CIs) were calculated with the NH/white category as the reference group; covariates included known confounders-that is, body mass index (BMI), dietary and other lifestyle factors. RESULTS: The NH alone category had the highest proportion of overweight and obese individuals and the NH/Asian category the lowest proportion. During 12 years of follow-up after cohort entry at 56 years, 2072 incident cases were ascertained through questionnaires and health plan linkages. All NH categories had higher HRs than the NH/white category before and after adjustment for BMI. In the fully adjusted models, the NH/Asian category showed the highest risk (HR=1.45; 95% CI: 1.27-1.65), followed by NH/other (HR=1.20; 95% CI: 1.03-1.39), NH/Chinese/white (HR=1.19; 95% CI: 1.04-1.37) and NH alone (HR=1.19; 95% CI: 1.03-1.37). The elevated risk by Asian admixture was more pronounced in normal weight than overweight/obese individuals. CONCLUSIONS: These findings indicate that Asian admixture in NHs is associated with a higher risk for type 2 diabetes independent of known risk factors and suggest a role for ethnicity-related genetic factors in the development of this disease.

Mitochondrial dynamics and their intracellular traffic in porcine oocytes.

Meiotic maturation of oocytes requires a variety of ATP-dependent reactions, such as germinal vesicle breakdown, spindle formation, and rearrangement of plasma membrane structure, which is required for fertilization. Mitochondria are accordingly expected be localized to subcellular sites of energy utilization. Although microtubule-dependent cellular traffic for mitochondria has been studied extensively in cultured neuronal (and some other somatic) cells, the molecular mechanism of their dynamics in mammalian oocytes at different stages of maturation remains obscure. The present work describes dynamic aspects of mitochondria in porcine oocytes at the germinal vesicle stage. After incubation of oocytes with MitoTracker Orange followed by centrifugation, mitochondria-enriched ooplasm was obtained using a glass needle and transferred into a recipient oocyte. The intracellular distribution of the fluorescent mitochondria was then observed over time using a laser scanning confocal microscopy equipped with an incubator. Kinetic analysis revealed that fluorescent mitochondria moved from central to subcortical areas of oocytes and were dispersed along plasma membranes. Such movement of mitochondria was inhibited by either cytochalasin B or cytochalasin D but not by colcemid, suggesting the involvement of microfilaments. This method of visualizing mitochondrial dynamics in live cells permits study of the pathophysiology of cytoskeleton-dependent intracellular traffic of mitochondria and associated energy metabolism during meiotic maturation of oocytes.

Heat treatment inhibits skeletal muscle atrophy of glucocorticoid-induced myopathy in rats.

The purpose of this study was to investigate the influence of heat treatment on glucocorticoid (GC)-induced myopathy. Eight-week-old Wistar rats were randomly assigned to the control, Dex, and Dex + Heat groups. Dexamethasone (2 mg/kg) was injected subcutaneously 6 days per week for 2 weeks in the Dex and Dex + Heat group. In the Dex + Heat group, heat treatment was performed by immersing hindlimbs in water at 42 °C for 60 min, once every 3 days for 2 weeks. The extensor digitorum longus muscle was extracted following 2 weeks of experimentation. In the Dex + Heat group, muscle fiber diameter, capillary/muscle fiber ratio, and level of heat shock protein 72 were significantly higher and atrogene expression levels were significantly lower than in the Dex group. Our results suggest that heat treatment inhibits the development of GC-induced myopathy by decreasing atrogene expression and increasing angiogenesis.

Intraoperative detection of viable bone with fluorescence imaging using Visually Enhanced Lesion Scope in patients with bisphosphonate-related osteonecrosis of the jaw: clinical and pathological evaluation.

UNLABELLED: There is no standard surgical protocol of bisphosphonate-related osteonecrosis of the jaws (BRONJ), because of the impossibility to visualize this feature intraoperatively. The aim of this study was to introduce how to provide preoperative labeling of the viable bone with minocycline bone fluorescence technique (MBFT) by using VELscope® and investigate histopathologically. INTRODUCTION: The American Association of Oral and Maxillofacial Surgeons (AAOMS) and the Japanese Society of Oral and Maxillofacial Surgeons (JSOMS) now recommend a more conservative treatment strategy. There is no standard surgical protocol of bisphosphonate-related osteonecrosis of the jaws (BRONJ) because of the impossibility to visualize this feature intraoperatively. The aim of this study was to introduce a mechanism providing preoperative labeling of a viable bone using minocycline bone fluorescence technique (MBFT) with VELscope® and to histopathologically investigate. METHODS: This report describes a surgical technique used in six patients with BRONJ who underwent jawbone resection under minocycline bone fluorescence imaging using VELscope®. Subsequently, we investigated and compared the clinical findings using VELscope® and histopathological findings. RESULTS: Histopathological examinations showed that the non-fluorescent moiety was consistent with the BRONJ lesions. CONCLUSIONS: The surgical treatments that were exactly performed using MBFT with VELscope® offered successful management of BRONJ. This bone fluorescence helped to define the margins of resection, thus improving surgical therapy for extended osteonecrosis.

Photodynamic therapy mediates innate immune responses via fibroblast-macrophage interactions.

Antibacterial photodynamic therapy (PDT) has come to attract attention as an alternative therapy for drug-resistant bacteria. Recent reports revealed that antibacterial PDT induces innate immune response and stimulates abundant cytokine secretion as a part of inflammatory responses. However, the underlying mechanism how antibacterial PDT interacts with immune cells responsible for cytokine secretion has not been well outlined. In this study, we aimed to clarify the difference in gene expression and cytokine secretion between combined culture of fibroblasts and macrophages and their independent cultures. SCRC-1008, mouse fibroblast cell line and J774, mouse macrophage-like cell line were co-cultured and PDT treatments with different parameters were carried out. After various incubation periods (1-24 h), cells and culture medium were collected, and mRNA and protein levels for cytokines were measured using real-time PCR and ELISA, respectively. Our results showed that fibroblasts and macrophages interact with each other to mediate the immune response. We propose that fibroblasts initially respond to PDT by expressing Hspa1b, which regulates the NF-κB pathway via Tlr2 and Tlr4. Activation of the NF-κB pathway then results in an enhanced secretion of pro-inflammatory cytokines (TNF-α, IL-6 and IL-1ß) and neutrophil chemoattractant MIP-2 and KC from macrophages.

Droplet digital PCR measurement of HER2 in patients with gastric cancer.

BACKGROUND: Although there are some new criteria for human epidermal growth factor receptor 2 (HER2) expression with immunohistochemistry/fluorescence in situ hybridisation (IHC/FISH) in gastric cancer, the method is still ambiguous and is somewhat dependent on the subjective qualities of the evaluator. METHODS: We used droplet digital polymerase chain reaction (ddPCR) to evaluate HER2 amplification in formalin-fixed and paraffin-embedded (FFPE) samples and cell-free serum circulating tumour DNA (ctDNA) in 25 patients with gastric cancer. RESULTS: The concordance rate of HER2 amplification examined in FFPE samples with ddPCR and IHC/FISH was 92% (23 out of 25). The concordance rate of FFPE with ctDNA was not high (62.5%); however, patients who were HER2-positive by ctDNA had significantly shorter survival compared with HER2-negative patients. CONCLUSIONS: Our results demonstrated that this ddPCR method was as effective as IHC/FISH and therefore might become a standard method for analysing not only FFPE but also ctDNA.

Multi-drug-resistant Staphylococcus aureus and future chemotherapy.

Staphylococcus (S.) aureus silently stays as our natural flora, and yet sometimes threatens our life as a tenacious pathogen. In addition to its ability to outwit our immune system, its multi-drug resistance phenotype makes it one of the most intractable pathogenic bacteria in the history of antibiotic chemotherapy. It conquered practically all the antibiotics that have been developed since 1940s. In 1961, the first MRSA was found among S. aureus clinical isolates. Then MRSA prevailed throughout the world as a multi-resistant hospital pathogen. In 1997, MRSA strain Mu50 with reduced susceptibility to vancomycin was isolated. Vancomycin-intermediate S. aureus (VISA), so named according to the CLSI criteria, was the product of adaptive mutation of S. aureus against vancomycin that had long been the last resort to MRSA infection. Here, we describe the genetic basis for the remarkable ability of S. aureus to acquire multi-antibiotic resistance, and propose a novel paradigm for future chemotherapy against the multi-resistant pathogens.