Improvement of infrared imaging video bolometer for application to deuterium experiment on the large helical device.

An infrared imaging video bolometer was improved for application to a neutron environment in fusion plasma devices, i.e., the Large Helical Device (LHD). In order to calibrate the thermal characteristics of the activated foil absorber inside the plasma vacuum vessel, the remote-controlled in situ calibration system was improved with high-surface-flatness mirrors. Furthermore, the carbon coating method was improved by introducing a vacuum evaporation technique instead of the conventional spray technique to realize the coating on both sides of the absorber with reproducibility and uniformity. The optimal thickness of the coating was also determined. Owing to these coating improvements, the reproducibility of the effective emissivity on both sides especially was improved. Finally, the variation with the neutron irradiation of the thermal characteristics of the foil absorber was investigated. It was found that the effect was not significant for the total neutron emission of 3.6 × 1018 on LHD.

New insights into mammalian signaling pathways using microfluidic pulsatile inputs and mathematical modeling.

Temporally modulated input mimics physiology. This chemical communication strategy filters the biochemical noise through entrainment and phase-locking. Under laboratory conditions, it also expands the observability space for downstream responses. A combined approach involving microfluidic pulsatile stimulation and mathematical modeling has led to deciphering of hidden/unknown temporal motifs in several mammalian signaling pathways and has provided mechanistic insights, including how these motifs combine to form distinct band-pass filters and govern fate regulation under dynamic microenvironment. This approach can be utilized to understand signaling circuit architectures and to gain mechanistic insights for several other signaling systems. Potential applications include synthetic biology and biotechnology, in developing pharmaceutical interventions, and in developing lab-on-chip models.

In situ calibration of the foil detector for an infrared imaging video bolometer using a carbon evaporation technique.

The InfraRed imaging Video Bolometer (IRVB) is a useful diagnostic for the multi-dimensional measurement of plasma radiation profiles. For the application of IRVB measurement to the neutron environment in fusion plasma devices such as the Large Helical Device (LHD), in situ calibration of the thermal characteristics of the foil detector is required. Laser irradiation tests of sample foils show that the reproducibility and uniformity of the carbon coating for the foil were improved using a vacuum evaporation method. Also, the principle of the in situ calibration system was justified.

Re-dislocation after corrective osteotomy for chronic dislocation of the radial head in children.

This retrospective study was designed to evaluate the outcomes of re-dislocation of the radial head after corrective osteotomy for chronic dislocation. A total of 12 children with a mean age of 11 years (5 to 16), with further dislocation of the radial head after corrective osteotomy of the forearm, were followed for a mean of five years (2 to 10). Re-operations were performed for radial head re-dislocation in six children, while the other six did not undergo re-operation ('non-re-operation group'). The active range of movement (ROM) of their elbows was evaluated before and after the first operation, and at the most recent follow-up. In the re-operation group, there were significant decreases in extension, pronation, and supination when comparing the ROM following the corrective osteotomy and following re-operation (p < 0.05). The children who had not undergone re-operation achieved a better ROM than those who had undergone re-operation. There was a significant difference in mean pronation (76° vs 0°) between the non- re-operation and the re-operation group (p = 0.002), and a trend towards increases in mean flexion (133° vs 111°), extension (0° vs 23°), and supination (62° vs 29°). We did not find a clear benefit for re-operation in children with a re-dislocation following corrective osteotomy for chronic dislocation of the radial head.

Band-pass processing in a GPCR signaling pathway selects for NFAT transcription factor activation.

Many biological processes are rhythmic and proper timing is increasingly appreciated as being critical for development and maintenance of physiological functions. To understand how temporal modulation of an input signal influences downstream responses, we employ microfluidic pulsatile stimulation of a G-protein coupled receptor, the muscarinic M3 receptor, in single cells with simultaneous real-time imaging of both intracellular calcium and NFAT nuclear localization. Interestingly, we find that reduced stimulation with pulses of ligand can give more efficient transcription factor activation, if stimuli are timed appropriately. Our experiments and computational analyses show that M3 receptor-induced calcium oscillations form a low pass filter while calcium-induced NFAT translocation forms a high pass filter. The combination acts as a band-pass filter optimized for intermediate frequencies of stimulation. We demonstrate that receptor desensitization and NFAT translocation rates determine critical features of the band-pass filter and that the band-pass may be shifted for different receptors or NFAT dynamics. As an example, we show that the two NFAT isoforms (NFAT4 and NFAT1) have shifted band-pass windows for the same receptor. While we focus specifically on the M3 muscarinic receptor and NFAT translocation, band-pass processing is expected to be a general theme that applies to multiple signaling pathways.

Evaluation of a novel immunochromatographic device for rapid and accurate clinical detection of Porphyromonas gingivalis in subgingival plaque.

UNLABELLED: An important goal for the improved diagnosis and management of infectious and inflammatory diseases, such as periodontitis, is the development of rapid and accurate technologies for the decentralized detection of bacterial pathogens. The aim of this prospective multicenter study was to evaluate the clinical use of a novel immunochromatographic device with monoclonal antibodies for the rapid point-of-care detection and semi-quantification of Porphyromonas gingivalis in subgingival plaque. Sixty-three patients with chronic periodontitis and 28 periodontally healthy volunteers were subjected to clinical and microbiological examinations. Subgingival plaque samples were analyzed for the presence of P. gingivalis using a novel immunochromatography based device DK13-PG-001, designed to detect the 40k-outer membrane protein of P. gingivalis, and compared with a PCR-Invader method. In the periodontitis group, a significant strong positive correlation in detection results was found between the test device score and the PCR-Invader method (Spearman rank correlation, r=0.737, p<0.0001). The sensitivity, specificity, and positive and negative predictive values of the test device were 96.2%, 91.8%, 90.4% and 96.7%, respectively. The detection threshold of the test device was determined to be approximately 10(4) (per two paper points). There were significant differences in the bacterial counts by the PCR-Invader method among groups with different ranges of device scores. With a cut-off value of ≥0.25 in device score, none of periodontally healthy volunteers were tested positive for the subgingival presence of P. gingivalis, whereas 76% (n=48) of periodontitis subjects were tested positive. There was a significant positive correlation between device scores for P. gingivalis and periodontal parameters including probing pocket depth and clinical attachment level (r=0.317 and 0.281, respectively, p<0.01). The results suggested that the DK13-PG-001 device kit can be effectively used for rapid, chair-side detection and semi-quantification of P. gingivalis in subgingival plaque. TRIAL REGISTRATION: UMIN Clinical Trials Registry (UMIN-CTR) UMIN000011943.

Slow and steady cell shrinkage reduces osmotic stress in bovine and murine oocyte and zygote vitrification.

STUDY QUESTION: Does the use of a new cryoprotectant agent (CPA) exchange protocol designed to minimize osmotic stress improve oocyte or zygote vitrification by reducing sublethal cryodamage? SUMMARY ANSWER: The use of a new CPA exchange protocol made possible by automated microfluidics improved oocyte and zygote vitrification with superior morphology as indicated by a smoother cell surface, higher sphericity, higher cytoplasmic lipid retention, less cytoplasmic leakage and higher developmental competence compared with conventional methods. WHAT IS KNOWN ALREADY: The use of more 'steps' of CPA exposure during the vitrification protocol increases cryosurvival and development in the bovine model. However, such an attempt to eliminate osmotic stress is limited by the practicality of performing numerous precise pipetting steps in a short amount of time. STUDY DESIGN, SIZE, DURATION: Murine meiotically competent germinal vesicle intact oocytes and zygotes were harvested from the antral follicles in ovaries and ampulla, respectively. Bovine ovaries were obtained from a local abattoir at random stages of the estrous cycle. A total of 110 murine oocytes, 802 murine zygotes and 52 bovine oocytes were used in this study. PARTICIPANTS/MATERIALS, SETTING, METHODS: Microfluidic devices were fabricated using conventional photo- and soft-lithography. CPAs used were 7.5% ethylene glycol (EG) and 7.5% dimethyl sulfoxide (DMSO) for equilibration solution and 15% EG, 15% DMSO and 0.5 M sucrose for vitrification solution. End-point analyses include mathematical modeling using Kedem-Katchalsky equations, morphometrics assessed by conventional and confocal microscopy, cytoplasmic lipid quantification by nile red staining, cytoplasmic leakage quantification by fluorescent dextran intercalation and developmental competence analysis by 96 h embryo culture and blastomere quantification. MAIN RESULTS AND THE ROLE OF CHANCE: The automated microfluidics protocol decreased the shrinkage rate of the oocyte and zygote by 13.8 times over its manual pipetting alternative. Oocytes and zygotes with a lower shrinkage rate during CPA exposure experienced less osmotic stress resulting in better morphology, higher cell quality and improved developmental competence. This microfluidic procedure resulted in murine zygotes with a significantly smoother cell surface (P < 0.001), more spherical cellular morphology (P < 0.001), increased cytoplasmic lipid retention in vitrified and warmed bovine oocytes (P < 0.01), decreased membrane perforations and cytoplasmic leakage in CPA-exposed murine zygotes (P < 0.05) and improved developmental competence of vitrified and warmed murine zygotes (P < 0.05) than CPA exposure using the current clinically used manual pipetting method. LIMITATIONS, REASONS FOR CAUTION: It is necessary to design the microfluidic device to be more user-friendly for widespread use. WIDER IMPLICATIONS OF THE FINDINGS: The theory and approach of eliminating osmotic stress by decreasing shrinkage rate is complementary to the prevalent osmotic stress theory in cryobiology which focuses on a minimum cell volume at which the cells shrink. The auto-microfluidic protocol described here has immediate applications for improving animal and human oocyte, zygote and embryo cryopreservation. On a fundamental level, the clear demonstration that at the same minimum cell volume, cell shrinkage rate affects sublethal damage should be broadly useful for cryobiology. STUDY FUNDING/COMPETING INTERESTS: This project was funded by the National Institutes of Health and the University of Michigan Reproductive Sciences Program. The authors declare no conflicts of interest.

Microfluidic source-sink model reveals effects of biophysically distinct CXCL12 isoforms in breast cancer chemotaxis.

Chemokines critically regulate chemotaxis in normal and pathologic states, but there is limited understanding of how multicellular interactions generate gradients needed for cell migration. Previous studies of chemotaxis of CXCR4+ cells toward chemokine CXCL12 suggest the requirement of cells expressing scavenger receptor CXCR7 in a source-sink system. We leveraged an established microfluidic device to discover that chemotaxis of CXCR4 cells toward distinct isoforms of CXCL12 required CXCR7 scavenging only under conditions with higher than optimal levels of CXCL12. Chemotaxis toward CXCL12-ß and -γ isoforms, which have greater binding to extracellular molecules and have been largely overlooked, was less dependent on CXCR7 than the more commonly studied CXCL12-α. Chemotaxis of CXCR4+ cells toward even low levels of CXCL12-γ and CXCL12-ß still occurred during treatment with a FDA-approved inhibitor of CXCR4. We also detected CXCL12-γ only in breast cancers from patients with advanced disease. Physiological gradient formation within the device facilitated interrogation of key differences in chemotaxis among CXCL12 isoforms and suggests CXCL12-γ as a biomarker for metastatic cancer.

Thinking big by thinking small: application of microfluidic technology to improve ART.

In Vitro Fertilization (IVF) laboratories often carry a penchant to resist change while in the pursuit of maintaining consistency in laboratory conditions. However, implementation of new technology is often critical to expand scientific discoveries and to improve upon prior successes to advance the field. Microfluidic platforms represent a technology that has the potential to revolutionize the fundamental processes of IVF. While the focus of microfluidic application in IVF has centered on embryo culture, the innovative platforms carry tremendous potential to improve other procedural steps and represents a possible paradigm shift in how we handle gametes and embryos. The following review will highlight application of various microfluidic platforms in IVF for use in maturation, manipulation, culture, cryopreservation and non-invasive quality assessment; pointing out new insights gained into functions of sperm, oocytes and embryos. Platform design and function will also be discussed, focusing on limitations, advancements and future refinements that can further aid in their clinical implementation.

A new model of gastric bleeding induced in rats by aspirin plus clopidogrel under stimulation of acid secretion. Prophylactic effects of antiulcer drugs.

We set up a new model of gastric bleeding induced by the luminal perfusion of aspirin (ASA) in rats pretreated with clopidogrel under conditions where acid secretion is stimulated, and examined the effect of antiulcer drugs on the bleeding. Under urethane anesthesia, acid secretion was stimulated by i.v. infusion of histamine (8 mg/kg/h), and two catheters were inserted into the stomach, one from the esophagus and another from the duodenum. The stomach was perfused with 25 mM ASA at a rate of 0.1 ml/min using an infusion pump, and gastric bleeding was measured as hemoglobin concentration in the perfusate collected every 15 min. Clopidogrel (30 mg/kg) was given orally 24 h before the perfusion. Various antiulcer drugs were given intraduodenally 30 min before the ASA treatment. Perfusion of the stomach with ASA provoked little gastric bleeding or damage even when acid secretion was stimulated. Pretreatment with clopidogrel significantly increased the bleeding and damage caused by ASA. The bleeding and lesions produced by ASA plus clopidogrel were significantly prevented by pretreatment with famotidine and omeprazole. Mucosal protective drugs such as rebamipide, irsogladine and teprenone also prevented gastric bleeding response to ASA/clopidogrel under conditions of acid secretion, although the effect was less pronounced than that of the antisecretory drugs. We conclude that clopidogrel increases gastric bleeding induced by ASA when acid secretion is stimulated. Both antisecretory and mucosal protective drugs are effective in reducing gastric bleeding under such conditions. This model is useful for the screening of drugs that protect against gastric bleeding.

Periodic cracking of films supported on compliant substrates.

When a tensile strain is applied to a film supported on a compliant substrate, a pattern of parallel cracks can channel through both the film and substrate. A linear-elastic fracture-mechanics model for the phenomenon is presented to extend earlier analyses in which cracking was limited to the film. It is shown how failure of the substrate reduces the critical strain required to initiate fracture of the film. This effect is more pronounced for relatively tough films. However, there is a critical ratio of the film to substrate toughness above which stable cracks do not form in response to an applied load. Instead, catastrophic failure of the substrate occurs simultaneously with the propagation of a single channel crack. This critical toughness ratio increases with the modulus mismatch between the film and substrate, so that periodic crack patterns are more likely to be observed with relatively stiff films. With relatively low values of modulus mismatch, even a film that is more brittle than the substrate can cause catastrophic failure of the substrate. Below the critical toughness ratio, there is a regime in which stable crack arrays can be formed in the film and substrate. The depth of these arrays increases, while the spacing decreases, as the strain is increased. Eventually, the crack array can become deep enough to cause substrate failure.

Yokukansan, a traditional Japanese medicine, ameliorates memory disturbance and abnormal social interaction with anti-aggregation effect of cerebral amyloid ß proteins in amyloid precursor protein transgenic mice.

The deposition of amyloid ß protein (Aß) is a consistent pathological hallmark of Alzheimer's disease (AD) brains. Therefore, inhibition of Aß aggregation in the brain is an attractive therapeutic and preventive strategy in the development of disease-modifying drugs for AD. An in vitro study demonstrated that yokukansan (YKS), a traditional Japanese medicine, inhibited Aß aggregation in a concentration-dependent manner. An in vivo study demonstrated that YKS and Uncaria hook (UH), a constituent of YKS, prevented the accumulation of cerebral Aß. YKS also improved the memory disturbance and abnormal social interaction such as increased aggressive behavior and decreased social behavior in amyloid precursor protein transgenic mice. These results suggest that YKS is likely to be a potent and novel therapeutic agent to prevent and/or treat AD, and that this may be attributed to UH.

Surgical treatment of spontaneous posterior interosseous nerve palsy: a retrospective study of 50 cases.

We have reviewed 38 surgically treated cases of spontaneous posterior interosseous nerve palsy in 38 patients with a mean age of 43 years (13 to 68) in order to identify clinical factors associated with its prognosis. Interfascicular neurolysis was performed at a mean of 13 months (1 to 187) after the onset of symptoms. The mean follow-up was 21 months (5.5 to 221). Medical Research Council muscle power of more than grade 4 was considered to be a good result. A further 12 cases in ten patients were treated conservatively and assessed similarly. Of the 30 cases treated surgically with available outcome data, the result of interfascicular neurolysis was significantly better in patients < 50 years old (younger group (18 nerves); good: 13 nerves (72%), poor: five nerves (28%)) than in cases > 50 years old (older group (12 nerves); good: one nerve (8%), poor: 11 nerves (92%)) (p < 0.001). A pre-operative period of less than seven months was also associated with a good result in the younger group (p = 0.01). The older group had a poor result regardless of the pre-operative delay. Our recommended therapeutic approach therefore is to perform interfascicular neurolysis if the patient is < 50 years of age, and the pre-operative delay is < seven months. If the patient is > 50 years of age with no sign of recovery for seven months, or in the younger group with a pre-operative delay of more than a year, we advise interfascicular neurolysis together with tendon transfer as the primary surgical procedure.

Co-chaperone BAG3 and adenovirus penton base protein partnership.

The BAG family of Hsp70/Hsc70 co-chaperones is characterised by the presence of a conserved BAG domain at the carboxyl-terminus. BAG3 protein is the only member of this family containing also the N-terminally located WW domain. We describe here the identification of adenovirus (Ad) penton base protein as the first BAG3 partner recognising BAG3 WW domain. Ad penton base is the viral capsid constituent responsible for virus internalisation. It contains in the N-terminal part two conserved PPxY motifs, known ligands of WW domains. In cells producing Ad penton base protein, cytoplasmic endogenous BAG3 interacts with it and co-migrates to the nucleus. Preincubation of BAG3 with Ad base protein results in only slight modulation of BAG3 co-chaperone activity, suggesting that this interaction is not related to the classical BAG3 co-chaperone function. However, depletion of BAG3 impairs the cell entry of the virus and viral progeny production in Ad-infected cells, suggesting that the interaction between virus penton base protein and cellular co-chaperone BAG3 positively influences virus life cycle. These results thus demonstrate a novel host-pathogen interaction, which contributes to the successful infectious life cycle of adenoviruses. In addition, these data enrich our knowledge about the multifunctionality of the BAG3 co-chaperone.

Dynamic microfunnel culture enhances mouse embryo development and pregnancy rates.

BACKGROUND: Despite advances in in vitro manipulation of preimplantation embryos, there is still a reduction in the quality of embryos produced leading to lower pregnancy rates compared with embryos produced in vivo. We hypothesized that a dynamic microfunnel embryo culture system would enhance outcomes by better mimicking the fluid-mechanical and biochemical stimulation embryos experience in vivo from ciliary currents and oviductal contractions. METHODS AND RESULTS: Mouse embryos were cultured in microdrop-static control, microfunnel-static control or microfunnel-dynamic conditions with microfluidics. All groups tested had greater than 90% total blastocyst development from zygotes after 96 h culture. Blastocyst developmental stage was significantly enhanced (P < 0.01) under dynamic microfunnel culture conditions as evidenced by an increased percentage of hatching or hatched blastocysts (Microdrop-control 31%; Microfunnel-control 23%; Microfunnel-pulsatile 71%) and significantly higher (P < 0.01) average number of cells per blastocyst (Microdrop-control 67 +/- 3; Microfunnel-control 60 +/- 3; Microfunnel-pulsatile 109 +/- 5). Blastocyst cell numbers in dynamic microfunnel cultures (109 +/- 5) more closely matched numbers obtained from in vivo grown blastocysts (144 +/- 9). Importantly, dynamic microfunnel culture significantly improved embryo implantation and ongoing pregnancy rates over static culture to levels approaching that of in utero derived preimplantation embryos. CONCLUSIONS: The improved pregnancy outcomes along with the simple and user-friendly design of the microfluidic/microfunnel system has potential to alleviate many inefficiencies in embryo production for biomedical research, genetic gain in domestic species and assisted reproductive technologies in humans.

Nanolitre liquid patterning in aqueous environments for spatially defined reagent delivery to mammalian cells.

Microscale biopatterning enables regulation of cell-material interactions and cell shape, and enables multiplexed high-throughput studies in a cell- and reagent-efficient manner. The majority of available techniques rely on physical contact of a stamp, pin, or mask with mainly a dry surface. Inkjet and piezoelectric printing is carried out in a non-contact manner but still requires a substantially dry substrate to ensure fidelity of printed patterns. These existing methods, therefore, are limited for patterning onto delicate surfaces of living cells because physical contact or substantially dry conditions are damaging to them. Microfluidic patterning with laminar streams does enable non-contact patterning in fully aqueous environments but with limited throughput and reagent diffusion across interfacial flows. Here, we describe a polymeric aqueous two-phase system that enables patterning nanolitres of a reagent-containing aqueous phase, in arbitrary shapes, within a second aqueous phase covering a cell monolayer. With the appropriate medium formulation, reagents of interest remain confined to the patterned phase without significant diffusion. The fully aqueous environment ensures high reagent activity and cell viability. The utility of this strategy is demonstrated with patterned delivery of genetic materials to mammalian cells for phenotypic screening of gene expression and gene silencing.

Liquid plug propagation in flexible microchannels: A small airway model.

In the present study, we investigate the effect of wall flexibility on the plug propagation and the resulting wall stresses in small airway models with experimental measurements and numerical simulations. Experimentally, a flexible microchannel was fabricated to mimic the flexible small airways using soft lithography. Liquid plugs were generated and propagated through the microchannels. The local wall deformation is observed instantaneously during plug propagation with the maximum increasing with plug speed. The pressure drop across the plug is measured and observed to increase with plug speed, and is slightly smaller in a flexible channel compared to that in a rigid channel. A computational model is then presented to model the steady plug propagation through a flexible channel corresponding to the middle plane in the experimental device. The results show qualitative agreements with experiments on wall shapes and pressure drops and the discrepancies bring up interesting questions on current field of modeling. The flexible wall deforms inward near the plug core region, the deformation and pressure drop across the plug increase with the plug speed. The wall deformation and resulting stresses vary with different longitudinal tensions, i.e., for large wall longitudinal tension, the wall deforms slightly, which causes decreased fluid stress and stress gradients on the flexible wall comparing to that on rigid walls; however, the wall stress gradients are found to be much larger on highly deformable walls with small longitudinal tensions. Therefore, in diseases such as emphysema, with more deformable airways, there is a high possibility of induced injuries on lining cells along the airways because of larger wall stresses and stress gradients.

Gas-liquid two-phase flow patterns in rectangular polymeric microchannels: effect of surface wetting properties.

Here we map gas-liquid two-phase flow regimes observed in polymeric microchannels with different wetting properties. We utilized video and confocal microscopy to examine two-phase flow patterns produced by parallel injection of air and water through a Y-shaped junction into a rectangular microchannel made of poly(dimethylsiloxane) (PDMS). We observed seven flow regimes in microchannels with hydrophobic walls, whereas only two flow patterns were identified in hydrophilic microchannels. Our study demonstrates that surface wettability has a profound influence on the spatial distribution of air and water moving in microchannels.