A sublethal ATP11A mutation associated with neurological deterioration causes aberrant phosphatidylcholine flipping in plasma membranes.

ATP11A translocates phosphatidylserine (PtdSer), but not phosphatidylcholine (PtdCho), from the outer to the inner leaflet of plasma membranes, thereby maintaining the asymmetric distribution of PtdSer. Here, we detected a de novo heterozygous point mutation of ATP11A in a patient with developmental delays and neurological deterioration. Mice carrying the corresponding mutation died perinatally of neurological disorders. This mutation caused an amino acid substitution (Q84E) in the first transmembrane segment of ATP11A, and mutant ATP11A flipped PtdCho. Molecular dynamics simulations revealed that the mutation allowed PtdCho binding at the substrate entry site. Aberrant PtdCho flipping markedly decreased the concentration of PtdCho in the outer leaflet of plasma membranes, whereas sphingomyelin (SM) concentrations in the outer leaflet increased. This change in the distribution of phospholipids altered cell characteristics, including cell growth, cholesterol homeostasis, and sensitivity to sphingomyelinase. Matrix-assisted laser desorption ionization-imaging mass spectrometry (MALDI-IMS) showed a marked increase of SM levels in the brains of Q84E-knockin mouse embryos. These results provide insights into the physiological importance of the substrate specificity of plasma membrane flippases for the proper distribution of PtdCho and SM.

Optogenetics on Zebrafish.

Optogenetics brought noninvasive neural activation in living organisms. Transparent zebrafish larva is one of the suitable animal models that receive the full benefit of this technique and provides behavioral studies based on intact individual nervous system. In this chapter, we describe methods to introduce optogenetic genes into zebrafish, and desirable apparatus for photostimulation and motion analysis with an example from our studies.

Trajectories for Post-traumatic Stress Disorder Symptoms Among Local Disaster Recovery Workers Following the Great East Japan Earthquake: Group-based Trajectory Modeling.

BACKGROUND: As many local municipality and medical workers were involved in disaster recovery duties following the Great East Japan Earthquake (GEJE) on March 11, 2011, the aim of this work was to elucidate the distinct trajectories for post-traumatic stress disorder (PTSD) symptoms and associated factors among these personnel. They confronted a diverse range of stressors both as survivors and as relief workers; however, little is known about their longitudinal PTSD symptoms. METHODS: The participants were 745 local municipality and hospital medical workers [average age: 43.6 ± 9.5 years, range: 20 - 66 years; 306 (59%) women] involved in disaster recovery duties following the GEJE. PTSD symptoms were measured using the Japanese version of the PTSD Checklist Specific Version (PCL-S) at four time points: 14, 30, 43, and 54 months after the GEJE. Using group-based trajectory modeling, distinct trajectories were elucidated. RESULTS: We identified five distinct PTSD symptoms profiles: resistance (n = 467, 62.7%), subsyndromal (n = 181, 24.3%), recovery (n = 47, 6.3%), fluctuating (n = 26, 3.5%), and chronic (n = 24, 3.2%). The trajectories differed according to the post-disaster working conditions and personal disaster experiences. LIMITATIONS: Potential selection bias resulting from the limited number of participants who completed all waves. The survey was conducted in one region of the disaster area. CONCLUSIONS: The majority of participants remained stable, with a relatively small group classified as chronic and fluctuating. Our results highlight the importance of improved working conditions and sustained monitoring of workers responding to natural disasters.

Dataset on mechanical, thermal and structural characterization of plant fiber-based biopolymers prepared by hot-pressing raw coconut coir, and milled powders of cotton, waste bagasse, wood, and bamboo.

This article presents experimental data on visual, mechanical, thermal, and structural characterization by hot-pressing four sources of milled plant powders and coconut fibers. It correlates chemical composition obtained by (FTIR), particle size, and reports bending strength, water resistance morphological (SEM) and thermal stability, structural properties (FTIR and XRD). It further supplements findings of the influence of microfibrillation and chemical composition on hot-pressing plant fibers as presented in the research article "Effects of Chemical Composition, Mild Alkaline Pretreatment and Particle Size on Mechanical, Thermal, and Structural Properties of Binderless Lignocellulosic Biopolymers Prepared by Hot-Pressing Raw Microfibrillated Phoenix Dactylifera and Cocos Nucifera Fibers and Leaves" [1]. For more insights into the difference among non-lignin-, lignin-, and semi lignin-based adhesion refer to the research article [1]. This dataset is made publicly available for potential reuse in recycling agricultural waste fibers for value-added materials.

Tissue regeneration during lower jaw restoration in zebrafish shows some features of epimorphic regeneration.

Zebrafish have the ability to regenerate skeletal structures, including the fin, skull roof, and jaw. Although fin regeneration proceeds by epimorphic regeneration, it remains unclear whether this process is involved in other skeletal regeneration in zebrafish. Initially in epimorphic regeneration, the wound epidermis covers the wound surface. Subsequently, the blastema, an undifferentiated mesenchymal mass, forms beneath the epidermis. In the present study, we re-examined the regeneration of the zebrafish lower jaw in detail, and investigated whether epimorphic regeneration is involved in this process. We performed amputation of the lower jaw at two different positions; the proximal level (presence of Meckel's cartilage) and the distal level (absence of Meckel's cartilage). In both manipulations, a blastema-like cellular mass was initially formed. Subsequently, cartilaginous aggregates were formed in this mass. In the proximal amputation, the cartilaginous aggregates were then fused with Meckel's cartilage and remained as a skeletal component of the regenerated jaw, whereas in the distal amputation, the cartilaginous aggregates disappeared as regeneration progressed. Two molecules that were observed during epimorphic regeneration, Laminin and msxb, were expressed in the regenerating lower jaw, although the domain of msxb expression was out of the main plain of the aggregate formation. Administration of an inhibitor of Wnt/ß-catenin signaling, a pathway associated with epimorphic regeneration, showed few effects on lower jaw regeneration. Our finding suggests that skeletal regeneration of the lower jaw mainly progresses through tissue regeneration that is dependent on the position in the jaw, and epimorphic regeneration plays an adjunctive role in this regeneration.

Diabetes care providers' manual for disaster diabetes care.

To ensure that experiences and lessons learned from the unprecedented 2011 Great East Japan Earthquake are used to improve future disaster planning, the Japan Diabetes Society (JDS) launched the "Research and Survey Committee for Establishing Disaster Diabetes Care Systems Based on Relevant Findings from the Great East Japan Earthquake" under the supervision of the Chairman of the JDS. The Committee conducted a questionnaire survey among patients with diabetes, physicians, disaster medical assistance teams (DMATs), nurses, pharmacists, and nutritionists in disaster areas about the events they saw happening, the situations they found difficult to handle, and the needs that they felt required to be met during the 2011 Great East Japan Earthquake. A total of 3,481 completed questionnaires were received. Based on these and other experiences and lessons reported following the 2011 Great East Japan Earthquake and the 2004 Niigata-Chuetsu Earthquakes, the current "Manual for Disaster Diabetes Care" has been developed by the members of the Committee and other invited authors from relevant specialties. To our knowledge, the current Manual is the world's first to focus on emergency diabetes care, with this digest English version translated from the Japanese original. It is sincerely hoped that patients with diabetes and healthcare providers around the world will find this manual helpful in promoting disaster preparedness and implementing disaster relief.

Diabetes Care Providers' Manual for Disaster Diabetes Care.

To ensure that experiences and lessons learned from the unprecedented 2011 Great East Japan Earthquake are used to improve future disaster planning, the Japan Diabetes Society (JDS) launched the "Research and Survey Committee for Establishing Disaster Diabetes Care Systems Based on Relevant Findings from the Great East Japan Earthquake" under the supervision of the Chairman of the JDS. The Committee conducted a questionnaire survey among patients with diabetes, physicians, disaster medical assistance teams (DMATs), nurses, pharmacists, and nutritionists in disaster areas about the events they saw happening, the situations they found difficult to handle, and the needs that they felt required to be met during the 2011 Great East Japan Earthquake. A total of 3,481 completed questionnaires were received. Based on these and other experiences and lessons reported following the 2011 Great East Japan Earthquake and the 2004 Niigata-Chuetsu Earthquakes, the current "Manual for Disaster Diabetes Care" has been developed by the members of the Committee and other invited authors from relevant specialties. To our knowledge, the current Manual is the world's first to focus on emergency diabetes care, with this digest English version translated from the Japanese original. It is sincerely hoped that patients with diabetes and healthcare providers around the world will find this manual helpful in promoting disaster preparedness and implementing disaster relief.

Recurrent de novo MAPK8IP3 variants cause neurological phenotypes.

c-Jun-amino-terminal kinase-interacting protein 3 (JIP3), encoded by MAPK8IP3, is an adaptor protein of the kinesin-1 complex and essential for axonal transport in neurons. However, an association between MAPK8IP3 variants and human disease has not been established. We identified 5 individuals from four families with recurrent de novo variants c.1732C>T (p.Arg578Cys) and c.3436C>T (p.Arg1146Cys) in MAPK8IP3. The core phenotype includes spastic diplegia, intellectual disability, cerebral atrophy, and corpus callosum hypoplasia. Zebrafish embryos overexpressing human mutant JIP3 showed axon varicosities of the posterior lateral line nerve, suggesting an adverse effect on the developing axons. Our results suggest that MAPK8IP3 variants cause a neurodevelopmental disease. ANN NEUROL 2019;85:927-933.

The reliability and validity for Japanese type 2 diabetes patients of the Japanese version of the acceptance and action diabetes questionnaire.

BACKGROUND: The purpose of this study was to determing which psychological traits of Japanese type 2 diabetes patients would provide reliability and validity to the Japanese version of the Acceptance and Action Diabetes Questionnaire (AADQ-J). METHODS: Various questionnaires were administered to type 2 diabetes patients who were registered on the database of the research service provider; data from a total of 600 patients (mean ± SD age was 57.50 ± 9.87 years, female 21.83%) were analyzed. RESULTS: Three items were excluded because of psychometric concerns related to the original 11-item AADQ. Confirmation factor analyses revealed that the eight-item version demonstrated the best indicators of a goodness of fit. The questionnaire showed adequate internal consistency. The questionnaire demonstrated high measurement accuracy in broad trait values by the test information function of Item Response Theory. The questionnaire showed stronger positive correlations with self-care activities and HbA1c than with diabetes distress and depressive mood. CONCLUSIONS: The eight-item Japanese version of AADQ has reliability and validity for type 2 diabetes patients.

Criticism by community people and poor workplace communication as risk factors for the mental health of local welfare workers after the Great East Japan Earthquake: A cross-sectional study.

After a large-scale natural disaster, demand for social welfare services increases, and the mental health of local social welfare workers becomes a matter of great concern because of their dual role as support providers and disaster survivors. We examined whether work-related social stressors, including criticism by community people and poor workplace communication, were associated with increased risk of post-traumatic stress disorder (PTSD), depression, or psychological distress 20-22 months after the Great East Japan Earthquake (GEJE; March 11, 2011) in local social welfare workers. Demographic characteristics, disaster-related risk factors (near-death experience, dead/missing family members, loss of housing), and work-related social risk factors (criticism, lack of communication) were obtained 20-22 months after the GEJE from 822 local workers. Questionnaires measured PTSD, depression, and psychological stress. Bivariate and multivariate regression analyses were applied. More local social welfare workers suffered from mental health problems than would be expected. Criticism by community people was significantly associated with probable PTSD and high psychological distress (adjusted odds ratio = 2.31 and 2.55, respectively). Furthermore, lack of workplace communication was associated with probable PTSD, depression, and high psychological distress (adjusted odds ratio = 3.97, 4.27, and 4.65, respectively). Almost 2 years after the disaster, local relief workers still suffered from mental health problems. Because post-disaster work-related social stressors constitute risk factors for these mental health problems, measures to improve working conditions and prevent and treat mental disorders should be a priority.

Morphologies of Bacillus subtilis communities responding to environmental variation.

Bacterial communities exhibit a variety of growth morphologies in constructing robust systems under different environmental conditions. We review the diverse morphologies of Bacillus subtilis communities and their mechanisms of self-organization. B. subtilis uses different cell types to suit environmental conditions and cell density. The subpopulation of each cell type exhibits various environment-sensitive properties. Furthermore, division of labor among the subpopulations results in flexible development for the community as a whole. We review how B. subtilis community morphologies and growth strategies respond to environmental perturbations.

Self-organization of bacterial communities against environmental pH variation: Controlled chemotactic motility arranges cell population structures in biofilms.

As with many living organisms, bacteria often live on the surface of solids, such as foods, organisms, buildings and soil. Compared with dispersive behavior in liquid, bacteria on surface environment exhibit significantly restricted mobility. They have access to only limited resources and cannot be liberated from the changing environment. Accordingly, appropriate collective strategies are necessarily required for long-term growth and survival. However, in spite of our deepening knowledge of the structure and characteristics of individual cells, strategic self-organizing dynamics of their community is poorly understood and therefore not yet predictable. Here, we report a morphological change in Bacillus subtilis biofilms due to environmental pH variations, and present a mathematical model for the macroscopic spatio-temporal dynamics. We show that an environmental pH shift transforms colony morphology on hard agar media from notched 'volcano-like' to round and front-elevated 'crater-like'. We discover that a pH-dependent dose-response relationship between nutritional resource level and quantitative bacterial motility at the population level plays a central role in the mechanism of the spatio-temporal cell population structure design in biofilms.

From neuron to behavior: Sensory-motor coordination of zebrafish turning behavior.

Recent development of optogenetics brought non-invasive neural activation in living organisms. Transparent zebrafish larva is one of the suitable animal models for this technique, which enables us to investigate neural circuits for behaviors based on a whole individual nervous system. In this article we review our recent finding that suggests sensory-motor coordination in larval zebrafish escape behavior. When water vibration stimulates mechanosensory Rohon-Beard (RB) neurons, intra-spinal reflex circuit launches contralateral trunk muscle contraction that makes rapid body curvature for turning. In addition, positional information of the stimulus is conveyed to supra-spinal circuits, and then regulates the curvature strength for appropriate escape pathway from the threat. Sensory-motor coordination is a fundamental feature to adapt behaviors to environment, and zebrafish larvae would be an excellent model for elucidating its neural backbones.

Position- and quantity-dependent responses in zebrafish turning behavior.

Neural reflexes are stereotypical automatic responses often modulated by both intrinsic and environmental factors. We report herein that zebrafish larval C-shaped turning is modulated by the stimulated position of Rohon-Beard (RB) neurons. Targeted stimulation of more anterior RB neurons produces larger trunk flexion, which anticipates adult escape behavior by coordinated turning toward the appropriate direction. We also demonstrated that turning laterality varies with the numbers of stimulated neurons. Multi-cell stimulation of RB neurons elicits contralateral turning, as seen in the touch response to physical contact, while minimum input from single-cell stimulation induces ipsilateral turning, a phenomenon not previously reported. This ipsilateral response, but not the contralateral one, is impaired by transecting the ascending neural tract known as the dorsolateral fascicule (DLF), indicating that two, distinct neural circuits trigger these two responses. Our results suggest that RB neurons transmit the position and quantity of sensory information, which are then processed separately to modulate behavioral strength and to select turning laterality.

Membrane Translocation and Organelle-Selective Delivery Steered by Polymeric Zwitterionic Nanospheres.

The majority of nanoparticles designed for cellular delivery of drugs and imaging agents enter the cell via endocytotic pathways leading to their entrapment in endosomes that present a robust barrier to further trafficking of the nanoparticles within the cells. A few materials, such as the cell penetrating peptides (CPPs), are known to enter cells not only via endocytosis, but also via translocation through the cell membrane into the cytoplasm, successfully bypassing the endosomes. We report here that random copolymers of 3-dimethyl(methacryloyloxyethyl)ammonium propanesulfonate and poly(ethylene glycol) methacrylate, p(DMAPS-ran-PEGMA), are internalized in cells primarily via translocation through the cell membrane rather than endocytosis. The properties of the polymers and their modes of uptake were investigated systematically by dynamic light scattering, confocal fluorescence microscopy, and flow cytometry. Using specific inhibitors of the cellular uptake machinery in a human cervical carcinoma cell line (HeLa), we show that these nontoxic synthetic polyzwitterions exist in cell media as self-assembled nanospheres that unravel as they adsorb on the plasma membrane and translocate through it. Conjugates of p(DMAPS-ran-PEGMA) with rhodamine B were delivered selectively to the mitochondria, whereas doxorubicin (Dox)-p(DMAPS-ran-PEGMA) conjugates were accumulated in both the nucleus and the mitochondria, effectively inducing apoptosis in HeLa cells. These findings suggest that the noncytotoxic and readily synthesized p(DMAPS-ran-PEGMA) can find applications as bioimaging tools and drug nanocarriers.

NCYM promotes calpain-mediated Myc-nick production in human MYCN-amplified neuroblastoma cells.

NCYM is a cis-antisense gene of MYCN and is amplified in human neuroblastomas. High NCYM expression is associated with poor prognoses, and the NCYM protein stabilizes MYCN to promote proliferation of neuroblastoma cells. However, the molecular mechanisms of NCYM in the regulation of cell survival have remained poorly characterized. Here we show that NCYM promotes cleavage of MYCN to produce the anti-apoptotic protein, Myc-nick, both in vitro and in vivo. NCYM and Myc-nick were induced at G2/M phase, and NCYM knockdown induced apoptotic cell death accompanied by Myc-nick downregulation. These results reveal a novel function of NCYM as a regulator of Myc-nick production in human neuroblastomas.

NCYM, a Cis-antisense gene of MYCN, encodes a de novo evolved protein that inhibits GSK3ß resulting in the stabilization of MYCN in human neuroblastomas.

The rearrangement of pre-existing genes has long been thought of as the major mode of new gene generation. Recently, de novo gene birth from non-genic DNA was found to be an alternative mechanism to generate novel protein-coding genes. However, its functional role in human disease remains largely unknown. Here we show that NCYM, a cis-antisense gene of the MYCN oncogene, initially thought to be a large non-coding RNA, encodes a de novo evolved protein regulating the pathogenesis of human cancers, particularly neuroblastoma. The NCYM gene is evolutionally conserved only in the taxonomic group containing humans and chimpanzees. In primary human neuroblastomas, NCYM is 100% co-amplified and co-expressed with MYCN, and NCYM mRNA expression is associated with poor clinical outcome. MYCN directly transactivates both NCYM and MYCN mRNA, whereas NCYM stabilizes MYCN protein by inhibiting the activity of GSK3ß, a kinase that promotes MYCN degradation. In contrast to MYCN transgenic mice, neuroblastomas in MYCN/NCYM double transgenic mice were frequently accompanied by distant metastases, behavior reminiscent of human neuroblastomas with MYCN amplification. The NCYM protein also interacts with GSK3ß, thereby stabilizing the MYCN protein in the tumors of the MYCN/NCYM double transgenic mice. Thus, these results suggest that GSK3ß inhibition by NCYM stabilizes the MYCN protein both in vitro and in vivo. Furthermore, the survival of MYCN transgenic mice bearing neuroblastoma was improved by treatment with NVP-BEZ235, a dual PI3K/mTOR inhibitor shown to destabilize MYCN via GSK3ß activation. In contrast, tumors caused in MYCN/NCYM double transgenic mice showed chemo-resistance to the drug. Collectively, our results show that NCYM is the first de novo evolved protein known to act as an oncopromoting factor in human cancer, and suggest that de novo evolved proteins may functionally characterize human disease.

Hindbrain V2a neurons in the excitation of spinal locomotor circuits during zebrafish swimming.

BACKGROUND: During locomotion in vertebrates, reticulospinal neurons in the hindbrain play critical roles in providing descending excitation to the spinal cord locomotor systems. However, despite the fact that many genes that are used to classify the neuronal identities of neurons in the hindbrain have been identified, the molecular identity of the reticulospinal neurons that are critically involved in locomotor drive is not well understood. Chx10-expressing neurons (V2a neurons) are ipsilaterally projecting glutamatergic neurons in the spinal cord and the hindbrain. Many of the V2a neurons in the hindbrain are known to project to the spinal cord in zebrafish, making hindbrain V2a neurons a prime candidate in descending locomotor drive. RESULTS: We investigated the roles of hindbrain V2a neurons using optogenetic and electrophysiological approaches. The forced activation of hindbrain V2a neurons using channelrhodopsin efficiently evoked swimming, whereas the forced inactivation of them using Archearhodopsin3 or Halorhodpsin reliably stopped ongoing swimming. Electrophysiological recordings of two populations of hindbrain reticulospinal V2a neurons showed that they were active during swimming. One population of neurons, small V2a neurons in the caudal hindbrain, fired with low rhythmicity, whereas the other population of neurons, large reticulospinal V2a neurons, called MiV1 neurons, fired more rhythmically. CONCLUSIONS: These results indicated that hindbrain reticulospinal V2a neurons play critical roles in providing excitation to the spinal locomotor circuits during swimming by providing both tonic and phasic inputs to the circuits.

Application of infrared laser to the zebrafish vascular system: gene induction, tracing, and ablation of single endothelial cells.

OBJECTIVE: Infrared laser-evoked gene operator is a new microscopic method optimized to heat cells in living organisms without causing photochemical damage. By combining the promoter system for the heat shock response, infrared laser-evoked gene operator enables laser-mediated gene induction in targeted cells. We applied this method to the vascular system in zebrafish embryos and demonstrated its usability to investigate mechanisms of vascular morphogenesis in vivo. APPROACH AND RESULTS: We used double-transgenic zebrafish with fli1:nEGFP to identify the endothelial cells, and with hsp:mCherry to carry out single-cell labeling. Optimizing the irradiation conditions, we finally succeeded in inducing the expression of the mCherry gene in single targeted endothelial cells, at a maximum efficiency rate of 60%. In addition, we indicated that this system could be used for laser ablation under certain conditions. To evaluate infrared laser-evoked gene operator, we applied this system to the endothelial cells of the first intersegmental arteries, and captured images of the connection between the vascular systems of the brain and spinal cord. CONCLUSIONS: Our results suggest that the infrared laser-evoked gene operator system will contribute to the elucidation of the mechanisms underlying vascular morphogenesis by controlling spatiotemporal gene activation in single endothelial cells, by labeling or deleting individual vessels in living embryos.

Targeted expression of a chimeric channelrhodopsin in zebrafish under regulation of Gal4-UAS system.

Channelrhodopsin (ChR)-wide receiver (ChRWR), one of the chimeric molecule of ChR1 and ChR2, has several advantages over ChR2 such as improved expression in the plasma membrane and enhanced photocurrent with small desensitization. Here we generated transgenic zebrafish (Danio rerio) expressing ChRWR as a conjugate of EGFP under the regulation of UAS promoter (UAS:ChRWR-EGFP). When crossed with a Gal4 line, SAGFF36B, ChRWR-EGFP was selectively expressed in primary mechanosensory Rohon-Beard (RB) neurons. The direct photoactivation of RB neurons was sufficient to trigger the escape behavior. The UAS:ChRWR-EGFP line could facilitate a variety of investigations of neural networks and behaviors of zebrafish in vivo.