Animal-borne soundscape logger as a system for edge classification of sound sources and data transmission for monitoring near-real-time underwater soundscape.

The underwater environment is filled with various sounds, with its soundscape composed of biological, geographical, and anthropological sounds. Our work focused on developing a novel method to observe and classify these sounds, enriching our understanding of the underwater ecosystem. We constructed a biologging system allowing near-real-time observation of underwater soundscapes. Utilizing deep-learning-based edge processing, this system classifies the sources of sounds, and upon the tagged animal surfacing, it transmits positional data, results of sound source classification, and sensor readings such as depth and temperature. To test the system, we attached the logger to sea turtles (Chelonia mydas) and collected data through a cellular network. The data provided information on the location-specific sounds detected by the sea turtles, suggesting the possibility to infer the distribution of specific species of organisms over time. The data showed that not only biological sounds but also geographical and anthropological sounds can be classified, highlighting the potential for conducting multi-point and long-term observations to monitor the distribution patterns of various sound sources. This system, which can be considered an autonomous mobile platform for oceanographic observations, including soundscapes, has significant potential to enhance our understanding of acoustic diversity.

Sensitivity assessment of diphacinone by pharmacokinetic analysis in invasive black rats in the Bonin (Ogasawara) Archipelago, Japan.

The Bonin Archipelago is a United Nations Educational, Scientific and Cultural Organization's World Natural Heritage Site in Japan with a unique ecosystem; however, the invasive rodents preying on endemic species have been a significant concern. The anticoagulant rodenticide, diphacinone, sprayed by the Ministry of the Environment, has succeeded; however, its repeated use leads to rodenticide resistance. This study evaluated the sensitivity by in vivo pharmacokinetics/pharmacodynamics (PK/PD) analysis and physiologically-based pharmacokinetic modeling to diphacinone in black rats (Rattus rattus) captured on the Bonin Archipelago in February 2022. The Bonin rats exhibited prolonged coagulation time after diphacinone administration. They recovered earlier than susceptible black rats, indicating that Bonin rats were less susceptible, though there were no genetic mutations in Vkorc1, the target enzyme of diphacinone. After the administration of diphacinone, hepatic expression levels of Fsp1, identified as the vitamin K reductase, was decreased, however, the Bonin rats exhibited the most minor suppression. The PK analysis showed that the excretion capacity of the Bonin rats was lower than that of the resistant black rats. In the PBPK modeling, the resistant black rats showed higher clearance than the Bonin and susceptible black rats due to high hepatic metabolic capacity. The Bonin rats demonstrated slow absorption and relatively low clearance. This study highlighted the reduced rodenticide-sensitive tendency of wild black rats in the Bonin Archipelago at an in vivo phenotype level. At the same time, they do not have known rodenticide resistance mechanisms, such as hepatic metabolic enhancement or Vkorc1 mutations. It is crucial to monitor the biological levels to evaluate rodenticide sensitivity accurately.

Structural basis of hydroxycarboxylic acid receptor signaling mechanisms through ligand binding.

Hydroxycarboxylic acid receptors (HCA) are expressed in various tissues and immune cells. HCA2 and its agonist are thus important targets for treating inflammatory and metabolic disorders. Only limited information is available, however, on the active-state binding of HCAs with agonists. Here, we present cryo-EM structures of human HCA2-Gi and HCA3-Gi signaling complexes binding with multiple compounds bound. Agonists were revealed to form a salt bridge with arginine, which is conserved in the HCA family, to activate these receptors. Extracellular regions of the receptors form a lid-like structure that covers the ligand-binding pocket. Although transmembrane (TM) 6 in HCAs undergoes dynamic conformational changes, ligands do not directly interact with amino acids in TM6, suggesting that indirect signaling induces a slight shift in TM6 to activate Gi proteins. Structural analyses of agonist-bound HCA2 and HCA3 together with mutagenesis and molecular dynamics simulation provide molecular insights into HCA ligand recognition and activation mechanisms.

Structures and mechanisms of actin ATP hydrolysis.

The major cytoskeleton protein actin undergoes cyclic transitions between the monomeric G-form and the filamentous F-form, which drive organelle transport and cell motility. This mechanical work is driven by the ATPase activity at the catalytic site in the F-form. For deeper understanding of the actin cellular functions, the reaction mechanism must be elucidated. Here, we show that a single actin molecule is trapped in the F-form by fragmin domain-1 binding and present their crystal structures in the ATP analog-, ADP-Pi-, and ADP-bound forms, at 1.15-Å resolutions. The G-to-F conformational transition shifts the side chains of Gln137 and His161, which relocate four water molecules including W1 (attacking water) and W2 (helping water) to facilitate the hydrolysis. By applying quantum mechanics/molecular mechanics calculations to the structures, we have revealed a consistent and comprehensive reaction path of ATP hydrolysis by the F-form actin. The reaction path consists of four steps: 1) W1 and W2 rotations; 2) PG-O3B bond cleavage; 3) four concomitant events: W1-PO3- formation, OH- and proton cleavage, nucleophilic attack by the OH- against PG, and the abstracted proton transfer; and 4) proton relocation that stabilizes the ADP-Pi-bound F-form actin. The mechanism explains the slow rate of ATP hydrolysis by actin and the irreversibility of the hydrolysis reaction. While the catalytic strategy of actin ATP hydrolysis is essentially the same as those of motor proteins like myosin, the process after the hydrolysis is distinct and discussed in terms of Pi release, F-form destabilization, and global conformational changes.

Structural insight into the activation mechanism of MrgD with heterotrimeric Gi-protein revealed by cryo-EM.

MrgD, a member of the Mas-related G protein-coupled receptor (MRGPR) family, has high basal activity for Gi activation. It recognizes endogenous ligands, such as ß-alanine, and is involved in pain and itch signaling. The lack of a high-resolution structure for MrgD hinders our understanding of whether its activation is ligand-dependent or constitutive. Here, we report two cryo-EM structures of the MrgD-Gi complex in the ß-alanine-bound and apo states at 3.1 Å and 2.8 Å resolution, respectively. These structures show that ß-alanine is bound to a shallow pocket at the extracellular domains. The extracellular half of the sixth transmembrane helix undergoes a significant movement and is tightly packed into the third transmembrane helix through hydrophobic residues, creating the active form. Our structures demonstrate a structural basis for the characteristic ligand recognition of MrgD. These findings provide a framework to guide drug designs targeting the MrgD receptor.

Role and Function of Mesenchymal Stem Cells on Fibroblast in Cutaneous Wound Healing.

Skin wounds often repair themselves completely over time; however, this is true only for healthy individuals. Although various studies are being conducted to improve wound-healing therapy outcomes, the mechanisms of wound healing and regeneration are not completely understood yet. In recent years, mesenchymal stem cells (MSCs) have been reported to contribute significantly to wound healing and regeneration. Understanding the function of MSCs will help to elucidate the fundamentals of wound healing. MSCs are multipotent stem cells that are used in regenerative medicine for their ability to self-renew and differentiate into bone, fat, and cartilage, with few ethical problems associated with cell harvesting. Additionally, they have anti-inflammatory and immunomodulatory properties and antifibrotic effects via paracrine signaling, and many studies have been conducted to use them to treat graft-versus-host disease, inflammatory bowel disease, and intractable cutaneous wounds. Many substances derived from MSCs are involved in the wound-healing process, and specific cascades and pathways have been elucidated. This review aims to explain the fundamental role of MSCs in wound healing and the effects of MSCs on fibroblasts.

Quartz crystal microbalance sensor for the detection of collagen model peptides based on the formation of triple helical structure.

Collagen is a major structural protein, and abnormalities in collagen structure can lead to several connective tissue diseases such as osteoporosis. We report the preparation of a collagen sensor using a synthetic peptide as proof of concept for detecting the collagen like peptides. The synthetic peptide 9-fluorenylmethyloxycarbonyl (Fmoc)-(prolyl-prolyl-glycine)7-OH was coupled to thiazolidine, which gets adsorbed on metal surfaces. Fmoc-(prolyl-prolyl-glycine)7-thiazolidine was immobilized on the surface of a quartz crystal microbalance (QCM) electrode used as a sensor probe. The collagen model peptide (prolyl-prolyl-glycine)10 could be detected, and the model peptide was directly adsorbed onto the surface of the electrode and was not removed by washing with hot water. Additionally, it was proved that the sensitivity of the probe could be enhanced to nanogram order by immobilizing the blocking reagent, Fmoc-prolyl-prolyl-glycine, within the gap of sensor probes on the electrode. The detectable mass of the model peptide decreased as the probe gap became narrower because of self-association of the probes. Moreover, the sensitivity of sensor probes also decreases as the gap between the probes becomes wider. Therefore, the optimum distance between the immobilized probes was determined from the simulation based on the experimental values. The association rate of the model peptide with sensor probes could be quantitatively determined when the distance between the probes was optimum, and this result suggested that most sensor probes could form a triple helical structure with the model peptide.

Description of the first acoustic recording of spinner dolphins (Stenella longirostris) from the northern straits of Malacca, Malaysia (L).

Relatively little is known about spinner dolphins in Malaysian waters and the wider Southeast Asian region. This note represents the first known acoustic recording of the species sighted opportunistically in the northern Straits of Malacca. Over a brief 20 min sighting, 46 whistles were recorded and four tonal types were detected, with 54.4% being upsweep whistles. The whistle duration ranged from 36 to 977 ms and the frequency ranged from 6.6 to 23.8 kHz. Fifty-seven click trains with a mean interclick interval of 41.5 ± 19.3 ms were detected. These findings provide a baseline for future regional acoustic research on this species.

Deep neural networks based automated extraction of dugong feeding trails from UAV images in the intertidal seagrass beds.

Dugongs (Dugong dugon) are seagrass specialists distributed in shallow coastal waters in tropical and subtropical seas. The area and distribution of the dugongs' feeding trails, which are unvegetated winding tracks left after feeding, have been used as an indicator of their feeding ground utilization. However, current ground-based measurements of these trails require a large amount of time and effort. Here, we developed effective methods to observe the dugongs' feeding trails using unmanned aerial vehicle (UAV) images (1) by extracting the dugong feeding trails using deep neural networks. Furthermore, we demonstrated two applications as follows; (2) extraction of the daily new feeding trails with deep neural networks and (3) estimation the direction of the feeding trails. We obtained aerial photographs from the intertidal seagrass bed at Talibong Island, Trang Province, Thailand. The F1 scores, which are a measure of binary classification model's accuracy taking false positives and false negatives into account, for the method (1) were 89.5% and 87.7% for the images with ground sampling resolutions of 1 cm/pixel and 0.5 cm/pixel, respectively, while the F1 score for the method (2) was 61.9%. The F1 score for the method (1) was high enough to perform scientific studies on the dugong. However, the method (2) should be improved, and there remains a need for manual correction. The mean area of the extracted daily new feeding trails from September 12-27, 2019, was 187.8 m2 per day (n = 9). Total 63.9% of the feeding trails was estimated to have direction within a range of 112.5° and 157.5°. These proposed new methods will reduce the time and efforts required for future feeding trail observations and contribute to future assessments of the dugongs' seagrass habitat use.

Formation of polycyclic aromatic hydrocarbons, benzofuran, and dibenzofuran in fuel-rich oxidation of toluene using a flow reactor.

Recently, polycyclic aromatic hydrocarbons (PAHs) and oxygenated PAHs (OPAHs) have been attracting considerable attention owing to their high toxicity. Understanding their formation mechanism during combustion processes is important to control their emission. However, there are few studies that have quantitatively investigated OPAH formation in the fuel-rich oxidation of hydrocarbons, despite the availability of several studies on PAH formation. In this study, benzofuran and dibenzofuran as OPAHs were quantified in the fuel-rich oxidation of toluene using a flow reactor at atmospheric pressure in a temperature range of 1050-1350 K at equivalence ratios from 3.0 to 12.0 and residence times from 0.2 to 1.5 s. In addition to benzofuran and dibenzofuran, 4 types of monocyclic aromatic hydrocarbons and 19 types of PAHs were also evaluated. The experimental data obtained in this study were compared with those of the ethylene oxidation performed in our previous study. The existing kinetic model for PAH growth was modified based on several theoretical studies to predict the behavior of OPAHs with furan structures. The modified model showed significant improvements in the prediction of benzofuran and dibenzofuran formation. Based on the rate of production and sensitivity analysis using the modified model, the dominant reaction pathways of benzofuran and dibenzofuran were investigated.

The structure of a 15-stranded actin-like filament from Clostridium botulinum.

Microfilaments (actin) and microtubules represent the extremes in eukaryotic cytoskeleton cross-sectional dimensions, raising the question of whether filament architectures are limited by protein fold. Here, we report the cryoelectron microscopy structure of a complex filament formed from 15 protofilaments of an actin-like protein. This actin-like ParM is encoded on the large pCBH Clostridium botulinum plasmid. In cross-section, the ~26 nm diameter filament comprises a central helical protofilament surrounded by intermediate and outer layers of six and eight twisted protofilaments, respectively. Alternating polarity of the layers allows for similar lateral contacts between each layer. This filament design is stiffer than the actin filament, and has likely been selected for during evolution to move large cargos. The comparable sizes of microtubule and pCBH ParM filaments indicate that larger filament architectures are not limited by the protomer fold. Instead, function appears to have been the evolutionary driving force to produce broad, complex filaments.

Structural basis for cofilin binding and actin filament disassembly.

Actin depolymerizing factor (ADF) and cofilin accelerate actin dynamics by severing and disassembling actin filaments. Here, we present the 3.8 Å resolution cryo-EM structure of cofilactin (cofilin-decorated actin filament). The actin subunit structure of cofilactin (C-form) is distinct from those of F-actin (F-form) and monomeric actin (G-form). During the transition between these three conformations, the inner domain of actin (subdomains 3 and 4) and the majority of subdomain 1 move as two separate rigid bodies. The cofilin-actin interface consists of three distinct parts. Based on the rigid body movements of actin and the three cofilin-actin interfaces, we propose models for the cooperative binding of cofilin to actin, preferential binding of cofilin to ADP-bound actin filaments and cofilin-mediated severing of actin filaments.

Helical rotation of the diaphanous-related formin mDia1 generates actin filaments resistant to cofilin.

The complex interplay between actin regulatory proteins facilitates the formation of diverse cellular actin structures. Formin homology proteins (formins) play an essential role in the formation of actin stress fibers and yeast actin cables, to which the major actin depolymerizing factor cofilin barely associates. In vitro, F-actin decorated with cofilin exhibits a marked increase in the filament twist. On the other hand, a mammalian formin mDia1 rotates along the long-pitch actin helix during processive actin elongation (helical rotation). Helical rotation may impose torsional force on F-actin in the opposite direction of the cofilin-induced twisting. Here, we show that helical rotation of mDia1 converts F-actin resistant to cofilin both in vivo and in vitro. F-actin assembled by mDia1 without rotational freedom became more resistant to the severing and binding activities of cofilin than freely rotatable F-actin. Electron micrographic analysis revealed untwisting of the long-pitch helix of F-actin elongating from mDia1 on tethering of both mDia1 and the pointed end side of the filament. In cells, single molecules of mDia1ΔC63, an activated mutant containing N-terminal regulatory domains, showed tethering to cell structures more frequently than autoinhibited wild-type mDia1 and mDia1 devoid of N-terminal domains. Overexpression of mDia1ΔC63 induced the formation of F-actin, which has prolonged lifetime and accelerates dissociation of cofilin. Helical rotation of formins may thus serve as an F-actin stabilizing mechanism by which a barbed end-bound molecule can enhance the stability of a filament over a long range.

Successful vitrification of whole juvenile testis in the critically endangered cyprinid honmoroko (Gnathopogon caerulescens).

Sperm cryopreservation is a valuable conservation method for endangered fish species. Here we report an easy and efficient cryopreservation method for juvenile whole testis by vitrification and successful sperm production from the vitrified whole testis via in vitro spermatogenesis in the critically endangered cyprinid honmoroko (Gnathopogon caerulescens). Juvenile testis (approximately 10 mm in length and 1 mm in width), consisting predominantly of spermatogonia, were aseptically dissected out and adherent fatty and non-testicular tissues were subsequently removed. Then, the testes were rapidly cooled on a nylon mesh by direct immersion in liquid nitrogen after serial exposures to pretreatment solution (PS), containing 2 M ethylene glycol (EG) and 1 M dimethyl sulfoxide (DMSO), for 20 or 30 min and vitrification solution (VS), containing 3 M EG, 2 M DMSO, and 0.5 M sucrose, for 5, 10, or 20 min. The highest survival rate of testicular cells (84.0%) was obtained from testes vitrified by immersion in PS for 20 min and in VS for 10 min. Spermatogonia were recovered from the vitrified testis by dissociation and cell culture produced many haploid sperm. Fertility and developmental competence were confirmed by in vitro fertilization assays. These results indicate that the vitrification of juvenile whole testis provides a new strategy to preserve the genetic resources of endangered fishes without affecting their reproductive population.

Long-term inhibition of cyclophilin D results in intracellular translocation of calcein AM from mitochondria to lysosomes.

Cyclophilin D is a peptidyl-prolyl cis-trans isomerase localized in the mitochondrial matrix. Although its effects on mitochondrial characteristics have been well studied, its relation to the uptake of molecules by mitochondria remains unknown. Here, we demonstrated the effects of cyclophilin D on the intracellular translocation of calcein AM. Following addition of calcein AM to control cells or cells overexpressing wild-type cyclophilin D, calcein fluorescence was observed in mitochondria. However, long-term inhibition of cyclophilin D in these cells altered the localization of calcein fluorescence from mitochondria to lysosomes without changing mitochondrial esterase activity. In addition, depletion of glucose from the medium recovered calcein localization from lysosomes to mitochondria. This is the first demonstration of the effects of cyclophilin D on the intracellular translocation of molecules other than proteins and suggests that cyclophilin D may modify mitochondrial features by inducing the translocation of molecules to the mitochondria through the mechanism associated with cellular energy metabolism.

[Case of continuous trans-arterial calcium gluconate infusion using a direct arterial sphygmomanometry line that exhibited dramatic improvement of chemical burns on the fingers caused by hydrofluoric acid].

Hydrofluoric acid (HFA) is commonly used and many injuries occur on the upper extremities following exposure to HFA. The use of calcium gluconate (CG) -containing gel or local injections of CG are widely used for the initial treatment of HFA exposure. However, severe pain continues in some cases despite the treatment. There was a report that trans-arterial CG infusion could improve HFA burns, however, such treatment is not an established clinical procedure. A 30-year-old male presented at our hospital with severe pain in his left thumb. He had been cleaning tiles with an HFA-containing detergent. We diagnosed him with a chemical burn due to HFA exposure. Local CG injections were tried several times, but his terrible pain continued. Therefore, a direct arterial sphygmomanometry line was inserted from the left radial artery, and continuous transarterial CG injection was performed. His terrible pain dramatically improved. Direct arterial sphygmomanometry systems are widely used in the critical care field to monitor the hemodynamics and ICU staffs are used to dealing with it. Moreover, continuous saline infusion prevents the tube obstruction. Continuous CG infusion from a direct arterial sphygmomanometry line is simple and safe way to administer CG in HFA burns.

An isolated vestibulocochlear symptom preceding brainstem infarction due to basilar artery occlusion.

OBJECTIVE: To describe unilateral hearing loss preceding brainstem infarction due to basilar artery occlusion. STUDY DESIGN: Clinical capsule report. SETTING: University hospital. PATIENT: A 67-year-old woman presented with unilateral acute hearing loss with dizziness as a sole prodromal manifestation of basilar artery occlusion for 2 weeks before the appearance of motor and oculomotor deficits. INTERVENTION: Otologic examinations and magnetic resonance imaging (MRI) were performed. RESULTS: A pure-tone audiogram showed unilateral profound sensorineural hearing loss preceding brainstem occlusion. Diffusion-weighted MR image demonstrated acute infarction around the paramedian area of the upper and middle pons, although T2-weighted MR image showed a normal brain image except for occlusion of the basilar artery. CONCLUSION: Acute unilateral hearing loss could be a prodrome of basilar artery occlusion. Clinicians must consider this possibility, especially in patients at high risk of brainstem infarction.

Low-gravity experiments of lightweight flexible heat pipe panels with self-rewetting fluids.

Fluids with a unique surface tension behavior, the so-called "self-rewetting fluids," are considered to be promising working fluids not only in reduced-gravity environments but also in terrestrial applications. Ultralightweight polyimide-based wickless heat pipe panels with flexible, inflatable, and deployable functions were fabricated using self-rewetting fluids. Fundamental operation tests of these panels were conducted under conditions of reduced gravity during parabolic flight. We obtained promising experimental results on the thermal performance of the panels in reduced gravity, although the experimental conditions were not entirely satisfactory.

Restoration of FcRgamma/Fyn signaling repairs central nervous system demyelination.

Disruption of myelin causes severe neurological diseases. An understanding of the mechanisms that control myelination and remyelination is needed to develop therapeutic strategies for demyelinating diseases such as multiple sclerosis (MS). Our previous finding indicating the critical involvement of the gamma chain of immunogloblin Fc receptors (FcRgamma) and Fyn signaling in oligodendrocyte differentiaion and myelination demands a fundamental revision of the strategies used for MS therapy, because antigen-antibody complexes in MS patients may induce the direct dysregulation of myelination process as well as the inflammatory destruction of myelin sheath. Here we show that the FcRgamma/Fyn signaling cascade is critically involved in cuprizone-induced demyelination/remyelination, with no lymphocytic response. The levels of phosphorylated myelin basic proteins (p-MBPs), especially the 21.5-kDa isoform, but not the levels of total MBPs, decreased markedly during demyelination induced by aging, cuprizone treatment, and double knockout of FcRgamma/Fyn genes. We also showed that the recovery from demyelination in cuprizone-treated and aged mice is achieved after administration of the herbal medicine Ninjin'yoeito, an effective therapy targeting the FcRgamma/Fyn-Rho (Rac1)-MAPK (P38 MAPK)-p-MBPs signaling cascade. These results suggest that the restoration of FcRgamma/Fyn signaling represents a new approach for the treatment of demyelinating diseases.