A vagal reflex evoked by airway closure.

Airway integrity must be continuously maintained throughout life. Sensory neurons guard against airway obstruction and, on a moment-by-moment basis, enact vital reflexes to maintain respiratory function1,2. Decreased lung capacity is common and life-threatening across many respiratory diseases, and lung collapse can be acutely evoked by chest wall trauma, pneumothorax or airway compression. Here we characterize a neuronal reflex of the vagus nerve evoked by airway closure that leads to gasping. In vivo vagal ganglion imaging revealed dedicated sensory neurons that detect airway compression but not airway stretch. Vagal neurons expressing PVALB mediate airway closure responses and innervate clusters of lung epithelial cells called neuroepithelial bodies (NEBs). Stimulating NEBs or vagal PVALB neurons evoked gasping in the absence of airway threats, whereas ablating NEBs or vagal PVALB neurons eliminated gasping in response to airway closure. Single-cell RNA sequencing revealed that NEBs uniformly express the mechanoreceptor PIEZO2, and targeted knockout of Piezo2 in NEBs eliminated responses to airway closure. NEBs were dispensable for the Hering-Breuer inspiratory reflex, which indicated that discrete terminal structures detect airway closure and inflation. Similar to the involvement of Merkel cells in touch sensation3,4, NEBs are PIEZO2-expressing epithelial cells and, moreover, are crucial for an aspect of lung mechanosensation. These findings expand our understanding of neuronal diversity in the airways and reveal a dedicated vagal pathway that detects airway closure to help preserve respiratory function.

Production enhancement of human adipose-derived mesenchymal stem cells by low-intensity ultrasound stimulation.

Low-intensity ultrasound (LIUS) enhances the proliferation rate of various mammalian stem cells through mechanical stimulation. This study quantitively finds suitable LIUS stimulation parameters for increasing the proliferation rate of human adipose-derived mesenchymal stem cells (hAdMSCs) for mass production. Various stimulation conditions of LIUS were assessed based on the beam pattern of the ultrasonic transducer and the attenuation of the sound waves. Using optimal LIUS stimulation parameters for enhancing proliferation of hAdMSCs taken from bromodeoxyuridine (BrdU) incorporation assay, long-term culture of hAdMSCs was performed for 16 days. The resultant hAdMSCs were characterized for various biomarkers such as CD34-, CD45-, CD73+, CD95+, CD105+ and cytological staining and a cytokine array assay. LIUS stimulation parameters found for enhancing the hAdMSCs proliferation were the frequency of 5 MHz, an intensity of 300 mWcm-2, a duration of 10 min per day, and continuous waves with a 100% duty cycle. The LIUS stimulated hAdMSCs group showed a 3.25-fold increase in the cell number compared to the control group after 16 days of culture. By confirming the effects of quantitatively measured LIUS stimulation on the enhancement of hAdMSCs proliferation, this study may be a foundation for the applications of LIUS stimulation in the industrial-scale production of hAdMSCs.

Discovery of levodopa-induced dyskinesia-associated genes using genomic studies in patients and Drosophila behavioral analyses.

Although levodopa is the most effective medication for Parkinson's disease, long-term levodopa treatment is largely compromised due to late motor complications, including levodopa-induced dyskinesia (LID). However, the genetic basis of LID pathogenesis has not been fully understood. Here, we discover genes pathogenic for LID using Drosophila genetics and behavioral analyses combined with genome-wide association studies on 578 patients clinically diagnosed with LID. Similar to the therapeutic effect of levodopa in patients, acute levodopa treatments restore the motor defect of Parkinson's disease model flies, while prolonged treatments cause LID-related symptoms, such as increased yawing, freezing and abrupt acceleration of locomotion. These symptoms require dopamine 1-like receptor 1 and are induced by neuronal overexpression of the receptor. Among genes selected from our analyses in the patient genome, neuronal knockdown of adenylyl cyclase 2 suppresses the levodopa-induced phenotypes and the receptor overexpression-induced symptoms in Drosophila. Together, our study provides genetic insights for LID pathogenesis through the D1-like receptor-adenylyl cyclase 2 signaling axis.

Loss of UCHL1 rescues the defects related to Parkinson's disease by suppressing glycolysis.

The role of ubiquitin carboxyl-terminal hydrolase L1 (UCHL1; also called PARK5) in the pathogenesis of Parkinson's disease (PD) has been controversial. Here, we find that the loss of UCHL1 destabilizes pyruvate kinase (PKM) and mitigates the PD-related phenotypes induced by PTEN-induced kinase 1 (PINK1) or Parkin loss-of-function mutations in Drosophila and mammalian cells. In UCHL1 knockout cells, cellular pyruvate production and ATP levels are diminished, and the activity of AMP-activated protein kinase (AMPK) is highly induced. Consequently, the activated AMPK promotes the mitophagy mediated by Unc-51-like kinase 1 (ULK1) and FUN14 domain-containing 1 (FUNDC1), which underlies the effects of UCHL1 deficiency in rescuing PD-related defects. Furthermore, we identify tripartite motif-containing 63 (TRIM63) as a previously unknown E3 ligase of PKM and demonstrate its antagonistic interaction with UCHL1 to regulate PD-related pathologies. These results suggest that UCHL1 is an integrative factor for connecting glycolysis and PD pathology.

Interoperable Nanoparticle Sensor Capable of Strain and Vibration Measurement for Rotor Blade Monitoring.

Recent advances in nanomaterials technology create the new possibility to fabricate high performance sensors. However, there has been limitations in terms of multivariate measurable and interoperable sensors. In this study, we fabricated an interoperable silver nanoparticle sensor fabricated by an aerodynamically focused nanomaterial (AFN) printing system which is a direct printing technique for inorganic nanomaterials onto a flexible substrate. The printed sensor exhibited the maximum measurable frequency of 850 Hz, and a gauge factor of 290.62. Using a fabricated sensor, we evaluated the sensing performance and demonstrated the measurement independency of strain and vibration sensing. Furthermore, using the proposed signal separation algorithm based on the Kalman filter, strain and vibration were each measured in real time. Finally, we applied the printed sensor to quadrotor condition monitoring to predict the motion of a quadrotor.

Periphery signals generated by Piezo-mediated stomach stretch and Neuromedin-mediated glucose load regulate the Drosophila brain nutrient sensor.

Nutrient sensors allow animals to identify foods rich in specific nutrients. The Drosophila nutrient sensor, diuretic hormone 44 (DH44) neurons, helps the fly to detect nutritive sugar. This sensor becomes operational during starvation; however, the mechanisms by which DH44 neurons or other nutrient sensors are regulated remain unclear. Here, we identified two satiety signals that inhibit DH44 neurons: (1) Piezo-mediated stomach/crop stretch after food ingestion and (2) Neuromedin/Hugin neurosecretory neurons in the ventral nerve cord (VNC) activated by an increase in the internal glucose level. A subset of Piezo+ neurons that express DH44 neuropeptide project to the crop. We found that DH44 neuronal activity and food intake were stimulated following a knockdown of piezo in DH44 neurons or silencing of Hugin neurons in the VNC, even in fed flies. Together, we propose that these two qualitatively distinct peripheral signals work in concert to regulate the DH44 nutrient sensor during the fed state.

Control of feeding by Piezo-mediated gut mechanosensation in Drosophila.

Across animal species, meals are terminated after ingestion of large food volumes, yet underlying mechanosensory receptors have so far remained elusive. Here, we identify an essential role for Drosophila Piezo in volume-based control of meal size. We discover a rare population of fly neurons that express Piezo, innervate the anterior gut and crop (a food reservoir organ), and respond to tissue distension in a Piezo-dependent manner. Activating Piezo neurons decreases appetite, while Piezo knockout and Piezo neuron silencing cause gut bloating and increase both food consumption and body weight. These studies reveal that disrupting gut distension receptors changes feeding patterns and identify a key role for Drosophila Piezo in internal organ mechanosensation.

The reasons why fractional flow reserve and instantaneous wave-free ratio are similar using wave separation analysis.

BACKGROUND AND OBJECTIVES: Fractional flow reserve (FFR) and instantaneous wave-free ratio (iFR) are the two most commonly used coronary indices of physiological stenosis severity based on pressure. To minimize the effect of wedge pressure (Pwedge), FFR is measured during hyperemia conditions, and iFR is calculated as the ratio of distal and aortic pressures (Pd/Pa) in the wave-free period. The goal of this study was to predict Pwedge using the backward wave (Pback) through wave separation analysis (WSA) and to reflect the effect of Pwedge on FFR and iFR to identify the relationship between the two indices. METHODS: An in vitro circulation system was constructed to calculate Pwedge. The measurements were performed in cases with stenosis percentages of 48, 71, and 88% and with hydrostatic pressures of 10 and 30 mmHg. Then, the correlation between Pback by WSA and Pwedge was calculated. In vivo coronary flow and pressure were simultaneously measured for 11 vessels in all patients. The FFR and iFR values were reconstructed as the ratios of forward wave at distal and proximal sites during hyperemia and at rest, respectively. RESULTS: Based on the in vitro results, the correlation between Pback and Pwedge was high (r = 0.990, p < 0.0001). In vivo results showed high correlations between FFR and reconstructed FFR (r = 0.992, p < 0.001) and between iFR and reconstructed iFR (r = 0.930, p < 0.001). CONCLUSIONS: Reconstructed FFR and iFR were in good agreement with conventional FFR and iFR. FFR and iFR can be expressed as the variation of trans-stenotic forward pressure, indicating that the two values are inferred from the same formula under different conditions.

Directly Printed Low-Cost Nanoparticle Sensor for Vibration Measurement during Milling Process.

A real-time, accurate, and reliable process monitoring is a basic and crucial enabler of intelligent manufacturing operation and digital twin applications. In this study, we represent a novel vibration measurement method for workpiece during the milling process using a low-cost nanoparticle vibration sensor. We directly printed the vibration sensor based on silver nanoparticles positioned onto a polyimide substrate using an aerodynamically-focused nanomaterials printing system, which is a direct printing technique for inorganic nanomaterials positioned onto a flexible substrate. Since it does not require any post-process such as chemical etching and heat treatment, a highly sensitive vibration sensor composed of a microscale porous structure was fabricated at a cost of several cents each. Furthermore, accurate and reliable vibration data was obtained by simple and direct attachment to a workpiece. In this study, we discussed the performance of vibration measurement of a fabricated sensor in comparison to a commercial vibration sensor. Using frequency and power spectrum analysis of obtained data, we directly measured the vibration of workpiece during the milling process, according to a process parameter. Lastly, we applied a fabricated sensor for the digital twins of turbine blade manufacturing in which vibration greatly affects the quality of the product to predict the process defects in real-time.

Stretchable Biaxial and Shear Strain Sensors Using Diffractive Structural Colors.

Structural colors that can be changed dynamically, using either plasmonic nanostructures or photonic crystals, are rapidly emerging research areas for stretchable sensors. Despite the wide applications of various techniques to achieve strain-responsive structural colors, important factors in the feasibility of strain sensors-such as their sensing mechanism, stability, and reproducibility-have not yet been explored. Here, we introduce a stretchable, diffractive, color-based wireless strain sensor that can measure strain using the entire visible spectrum, based on an array of cone-shaped nanostructures on the surface of an elastomeric substrate. By stretching or compressing the substrate, the diffractive color can be tuned according to the changing grating pitch. Using the proposed method, we designed three types of strain-sensing modes: large-deformation (maximum 100%) tensile strain, biaxial 2D strain, and shear strain (maximum 78%). The strain sensors were fabricated, and applicability to strain-sensing was evaluated.

Arterial Baroreceptors Sense Blood Pressure through Decorated Aortic Claws.

Mechanosensory neurons across physiological systems sense force using diverse terminal morphologies. Arterial baroreceptors are sensory neurons that monitor blood pressure for real-time stabilization of cardiovascular output. Various aortic sensory terminals have been described, but those that sense blood pressure are unclear because of a lack of selective genetic tools. Here, we find that all baroreceptor neurons are marked in Piezo2-ires-Cre mice and then use genetic approaches to visualize the architecture of mechanosensory endings. Cre-guided ablation of vagal and glossopharyngeal PIEZO2 neurons eliminates the baroreceptor reflex and aortic depressor nerve effects on blood pressure and heart rate. Genetic mapping reveals that PIEZO2 neurons form a distinctive mechanosensory structure: macroscopic claws that surround the aortic arch and exude fine end-net endings. Other arterial sensory neurons that form flower-spray terminals are dispensable for baroreception. Together, these findings provide structural insights into how blood pressure is sensed in the aortic vessel wall.

Highly Sensitive Solvent-free Silver Nanoparticle Strain Sensors with Tunable Sensitivity Created Using an Aerodynamically Focused Nanoparticle Printer.

We developed and presented highly sensitive solvent-free silver nanoparticle strain sensors fabricated using the aerodynamically focused nanoparticle (AFN) printer. The nanoparticles were printed in various conductive patterns. We explored how printer scan velocity affected pattern geometry and sensor sensitivity. The strain sensors were highly sensitive; the scan velocity afforded tunable sensitivity; and an analytical model predicted the behavior well under low-strain (<0.4%) conditions. We describe a prototype sensor that reliably measured composite beam tensile strain. We further enhanced the sensitivity by creating mechanical cracks, facilitating small dynamic signal measurements. The linear sensitivity of the sensor could be tuned from 18.60 to 290.62 by varying the scan velocity from 2 to 40 µm/s. The cracked sensor afforded the greatest sensitivity (1056) and captured small vibrations from a stringed instrument. We report highly sensitive and reliable measurements of dynamic behavior with simple tunability.

PIEZOs mediate neuronal sensing of blood pressure and the baroreceptor reflex.

Activation of stretch-sensitive baroreceptor neurons exerts acute control over heart rate and blood pressure. Although this homeostatic baroreflex has been described for more than 80 years, the molecular identity of baroreceptor mechanosensitivity remains unknown. We discovered that mechanically activated ion channels PIEZO1 and PIEZO2 are together required for baroreception. Genetic ablation of both Piezo1 and Piezo2 in the nodose and petrosal sensory ganglia of mice abolished drug-induced baroreflex and aortic depressor nerve activity. Awake, behaving animals that lack Piezos had labile hypertension and increased blood pressure variability, consistent with phenotypes in baroreceptor-denervated animals and humans with baroreflex failure. Optogenetic activation of Piezo2-positive sensory afferents was sufficient to initiate baroreflex in mice. These findings suggest that PIEZO1 and PIEZO2 are the long-sought baroreceptor mechanosensors critical for acute blood pressure control.

NEDD4-induced degradative ubiquitination of phosphatidylinositol 4-phosphate 5-kinase α and its implication in breast cancer cell proliferation.

Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) family members generate phosphatidylinositol 4,5-bisphosphate (PIP2), a critical lipid regulator of diverse physiological processes. The PIP5K-dependent PIP2 generation can also act upstream of the oncogenic phosphatidylinositol 3-kinase (PI3K)/Akt pathway. Many studies have demonstrated various mechanisms of spatiotemporal regulation of PIP5K catalytic activity. However, there are few studies on regulation of PIP5K protein stability. Here, we examined potential regulation of PIP5Kα, a PIP5K isoform, via ubiquitin-proteasome system, and its implication for breast cancer. Our results showed that the ubiquitin ligase NEDD4 (neural precursor cell expressed, developmentally down-regulated gene 4) mediated ubiquitination and proteasomal degradation of PIP5Kα, consequently reducing plasma membrane PIP2 level. NEDD4 interacted with the C-terminal region and ubiquitinated the N-terminal lysine 88 in PIP5Kα. In addition, PIP5Kα gene disruption inhibited epidermal growth factor (EGF)-induced Akt activation and caused significant proliferation defect in breast cancer cells. Notably, PIP5Kα K88R mutant that was resistant to NEDD4-mediated ubiquitination and degradation showed more potentiating effects on Akt activation by EGF and cell proliferation than wild-type PIP5Kα. Collectively, these results suggest that PIP5Kα is a novel degradative substrate of NEDD4 and that the PIP5Kα-dependent PIP2 pool contributing to breast cancer cell proliferation through PI3K/Akt activation is negatively controlled by NEDD4.

Misato underlies visceral myopathy in Drosophila.

Genetic mechanisms for the pathogenesis of visceral myopathy (VM) have been rarely demonstrated. Here we report the visceral role of misato (mst) in Drosophila and its implications for the pathogenesis of VM. Depletion of mst using three independent RNAi lines expressed by a pan-muscular driver elicited characteristic symptoms of VM, such as abnormal dilation of intestinal tracts, reduced gut motility, feeding defects, and decreased life span. By contrast, exaggerated expression of mst reduced intestine diameters, but increased intestinal motilities along with thickened muscle fibers, demonstrating a critical role of mst in the visceral muscle. Mst expression was detected in the adult intestine with its prominent localization to actin filaments and was required for maintenance of intestinal tubulin and actomyosin structures. Consistent with the subcellular localization of Mst, the intestinal defects induced by mst depletion were dramatically rescued by exogenous expression of an actin member. Upon ageing the intestinal defects were deteriorative with marked increase of apoptotic responses in the visceral muscle. Taken together, we propose the impairment of actomyosin structures induced by mst depletion in the visceral muscle as a pathogenic mechanism for VM.

The Hippo-Salvador signaling pathway regulates renal tubulointerstitial fibrosis.

Renal tubulointerstitial fibrosis (TIF) is the final pathway of various renal injuries that result in chronic kidney disease. The mammalian Hippo-Salvador signaling pathway has been implicated in the regulation of cell proliferation, cell death, tissue regeneration, and tumorigenesis. Here, we report that the Hippo-Salvador pathway plays a role in disease development in patients with TIF and in a mouse model of TIF. Mice with tubular epithelial cell (TEC)-specific deletions of Sav1 (Salvador homolog 1) exhibited aggravated renal TIF, enhanced epithelial-mesenchymal transition-like phenotypic changes, apoptosis, and proliferation after unilateral ureteral obstruction (UUO). Moreover, Sav1 depletion in TECs increased transforming growth factor (TGF)-ß and activated ß-catenin expression after UUO, which likely accounts for the abovementioned enhanced TEC fibrotic phenotype. In addition, TAZ (transcriptional coactivator with PDZ-binding motif), a major downstream effector of the Hippo pathway, was significantly activated in Sav1-knockout mice in vivo. An in vitro study showed that TAZ directly regulates TGF-ß and TGF-ß receptor II expression. Collectively, our data indicate that the Hippo-Salvador pathway plays a role in the pathogenesis of TIF and that regulating this pathway may be a therapeutic strategy for reducing TIF.

Positive geotactic behaviors induced by geomagnetic field in Drosophila.

BACKGROUND: Appropriate vertical movement is critical for the survival of flying animals. Although negative geotaxis (moving away from Earth) driven by gravity has been extensively studied, much less is understood concerning a static regulatory mechanism for inducing positive geotaxis (moving toward Earth). RESULTS: Using Drosophila melanogaster as a model organism, we showed that geomagnetic field (GMF) induces positive geotaxis and antagonizes negative gravitaxis. Remarkably, GMF acts as a sensory cue for an appetite-driven associative learning behavior through the GMF-induced positive geotaxis. This GMF-induced positive geotaxis requires the three geotaxis genes, such as cry, pyx and pdf, and the corresponding neurons residing in Johnston's organ of the fly's antennae. CONCLUSIONS: These findings provide a novel concept with the neurogenetic basis on the regulation of vertical movement by GMF in the flying animals.

Identification of a Peptidergic Pathway Critical to Satiety Responses in Drosophila.

Although several neural pathways have been implicated in feeding behaviors in mammals [1-7], it remains unclear how the brain coordinates feeding motivations to maintain a constant body weight (BW). Here, we identified a neuropeptide pathway important for the satiety and BW control in Drosophila. Silencing of myoinhibitory peptide (MIP) neurons significantly increased BW through augmented food intake and fat storage. Likewise, the loss-of-function mutation of mip also increased feeding and BW. Suppressing the MIP pathway induced satiated flies to behave like starved ones, with elevated sensitivity toward food. Conversely, activating MIP neurons greatly decreased food intake and BW and markedly blunted the sensitivity of starved flies toward food. Upon terminating the activation protocol of MIP neurons, the decreased BW reverts rapidly to the normal level through a strong feeding rebound, indicating the switch-like role of MIP pathway in feeding. Surprisingly, the MIP-mediated BW decrease occurred independently of sex peptide receptor (SPR), the only known receptor for MIP, suggesting the presence of a yet-unknown MIP receptor. Together, our results reveal a novel anorexigenic pathway that controls satiety in Drosophila and provide a new avenue to study how the brain actively maintains a constant BW.

Identification of a neural pathway governing satiety in Drosophila.

Satiety cues a feeding animal to cease further ingestion of food, thus protecting it from excessive energy gain. Impaired control of satiety is often associated with feeding-related disorders such as obesity. In our recent study, we reported the identification of a neural pathway that expresses the myoinhibitory peptide (MIP), critical for satiety responses in Drosophila. Targeted silencing of MIP neuron activity strikingly increased the body weight (BW) through elevated food intake. Similarly, genetic disruption of the gene encoding MIP also elevated feeding and BW. Suppressing the MIP pathway behaviorally transformed the satiated flies to feed similar to the starved ones, with augmented sensitivity to food. Conversely, temporal activation of MIP neuron markedly reduced the food intake and BW, and blunted the sensitivity of the starved flies to food as if they have been satiated. Shortly after termination of MIP neuron activation, the reduced BW reverted to the normal level along with a strong feeding rebound. Together our results reveal the switch-like role of the MIP pathway in feeding regulation by controlling satiety. [BMB Reports 2016; 49(3): 137-138].

Nutrient Sensor in the Brain Directs the Action of the Brain-Gut Axis in Drosophila.

Animals can detect and consume nutritive sugars without the influence of taste. However, the identity of the taste-independent nutrient sensor and the mechanism by which animals respond to the nutritional value of sugar are unclear. Here, we report that six neurosecretory cells in the Drosophila brain that produce Diuretic hormone 44 (Dh44), a homolog of the mammalian corticotropin-releasing hormone (CRH), were specifically activated by nutritive sugars. Flies in which the activity of these neurons or the expression of Dh44 was disrupted failed to select nutritive sugars. Manipulation of the function of Dh44 receptors had a similar effect. Notably, artificial activation of Dh44 receptor-1 neurons resulted in proboscis extensions and frequent episodes of excretion. Conversely, reduced Dh44 activity led to decreased excretion. Together, these actions facilitate ingestion and digestion of nutritive foods. We propose that the Dh44 system directs the detection and consumption of nutritive sugars through a positive feedback loop.