FOXO-regulated DEAF1 controls muscle regeneration through autophagy.

The commonality between various muscle diseases is the loss of muscle mass, function, and regeneration, which severely restricts mobility and impairs the quality of life. With muscle stem cells (MuSCs) playing a key role in facilitating muscle repair, targeting regulators of muscle regeneration has been shown to be a promising therapeutic approach to repair muscles. However, the underlying molecular mechanisms driving muscle regeneration are complex and poorly understood. Here, we identified a new regulator of muscle regeneration, Deaf1 (Deformed epidermal autoregulatory factor-1) - a transcriptional factor downstream of foxo signaling. We showed that Deaf1 is transcriptionally repressed by FOXOs and that DEAF1 targets to Pik3c3 and Atg16l1 promoter regions and suppresses their expression. Deaf1 depletion therefore induces macroautophagy/autophagy, which in turn blocks MuSC survival and differentiation. In contrast, Deaf1 overexpression inactivates autophagy in MuSCs, leading to increased protein aggregation and cell death. The fact that Deaf1 depletion and its overexpression both lead to defects in muscle regeneration highlights the importance of fine tuning DEAF1-regulated autophagy during muscle regeneration. We further showed that Deaf1 expression is altered in aging and cachectic MuSCs. Manipulation of Deaf1 expression can attenuate muscle atrophy and restore muscle regeneration in aged mice or mice with cachectic cancers. Together, our findings unveil an evolutionarily conserved role for DEAF1 in muscle regeneration, providing insights into the development of new therapeutic strategies against muscle atrophy.

Proof-of-concept study on a water phantom-based neutron spectrometer: Experimental test with 252Cf and 241Am-Be sources.

Boron neutron capture therapy (BNCT) is a promising cancer treatment that uses energetic ions released from 10B(n, α)7Li reactions. Accurate assessment of neutron energy spectra is important for simulation-based evaluation of neutron doses during BNCT. In this study, a proof-of-concept study was conducted for a neutron spectrometry technique that involves the use of a water phantom, which is commonly used for quality assurance in BNCT, as a moderator. The technique involves applying unfolding to the count rate distribution of the thermal neutron counter measured within the phantom to derive the energy spectrum. We performed experiments using a spherical 3He proportional counter in neutron fields generated by 252Cf and 241Am-Be sources. The results demonstrated that the spectrometer reasonably reproduced neutron spectra and showed the potential of using a water phantom as a moderator for such a technique.

Intraoperative MR Imaging during Glioma Resection.

One of the major issues in the surgical treatment of gliomas is the concern about maximizing the extent of resection while minimizing neurological impairment. Thus, surgical planning by carefully observing the relationship between the glioma infiltration area and eloquent area of the connecting fibers is crucial. Neurosurgeons usually detect an eloquent area by functional MRI and identify a connecting fiber by diffusion tensor imaging. However, during surgery, the accuracy of neuronavigation can be decreased due to brain shift, but the positional information may be updated by intraoperative MRI and the next steps can be planned accordingly. In addition, various intraoperative modalities may be used to guide surgery, including neurophysiological monitoring that provides real-time information (e.g., awake surgery, motor-evoked potentials, and sensory evoked potential); photodynamic diagnosis, which can identify high-grade glioma cells; and other imaging techniques that provide anatomical information during the surgery. In this review, we present the historical and current context of the intraoperative MRI and some related approaches for an audience active in the technical, clinical, and research areas of radiology, as well as mention important aspects regarding safety and types of devices.

Simple, Quick, and Safe Dural Incision Technique for Patients with Expected Brain Bulging during Decompressive Craniectomy: "Crank-shaped Dural Incisions".

Brain bulging is an unfavorable outcome in patients with brain swelling who require decompressive craniectomy (DC) to control elevated intracranial pressure (ICP). Although several previous studies have described methods for reducing the operation time during DC in these patients, few have proposed a technique for controlling brain protrusion. Here we describe an effective and simple method for external reduction of ICP and discuss its suitability for patients at risk of brain bulging during DC. After craniectomy, crank-shaped lines extending from a central square dural canopy are all marked on the dura. As the incisions are made, pressure from the swelling brain opens the lines and the protruding cortical surface forms dural windows. The square canopy gradually rotates as it stretches, and along with the remaining dura, functions to gently support and compress the cortex. In the case of insufficient decompression, the incision lines can be extended to further reduce ICP. As the parenchyma is accessible to the surgeon, hematoma removal can be performed through the dural windows. In initial experience of four patients who underwent this technique, ICP was controlled in all cases after surgery and no adverse events occurred. The crank-shaped dural incision method is a simple, quick, and effective technique for external reduction of ICP in patients at risk of brain bulging that is intuitive in the emergency situation and thus can be performed even by relatively inexperienced neurosurgeons.

Imaging dendritic spines: molecular organization and signaling for plasticity.

The structural plasticity of dendritic spines is considered to be essential for various forms of synaptic plasticity and, ultimately, learning and memory. The process is mediated by signaling pathways that promote the reorganization of the actin cytoskeleton and subsynaptic structures, which in turn cause structural and functional changes in dendritic spines. Recent advances in optical technologies have started to reveal the fine molecular structures and dynamic signaling occurring inside spines, providing significant insights into the molecular regulation of spines. Here, we highlight recent studies to resolve the molecular mechanisms underlying the spine actin cytoskeleton and plasticity with high spatiotemporal resolution. Moreover, we discuss new genome editing-based approaches in imaging the molecular structure and plasticity of dendritic spines.

A case report of Gorham-Stout disease diagnosed during the course of recurrent meningitis and cholesteatoma.

BACKGROUND: Gorham-Stout disease is a rare bone disorder. Here, we present a case of Gorham-Stout disease diagnosed during follow-up of a patient with cholesteatoma; the disease affected the temporal bone and other sites of the skull. To the best of our knowledge, this is the first report of Gorham-Stout disease diagnosed with recurrent cerebrospinal leakage after surgery to treat cholesteatoma. CASE PRESENTATION: A 25-year-old male patient re-presented to our department for the first time in 7 years with otorrhea in the right ear and recurrent meningitis. The patient had a history of multiple surgeries for cholesteatoma and suffered from recurrent cerebrospinal fluid leakage, which initially was thought to be caused by recurrence of cholesteatoma. Therefore, skull base reconstruction was planned. However, the underlying cause was identified eventually as defects in the temporal bone caused by massive osteolysis due to Gorham-Stout disease. Skull base reconstruction was abandoned because the osteolysis was considered to be progressive. Conservative treatment with infectious control was implemented as an alternative. CONCLUSION: This case describes unusual temporal bone osteolysis after cholesteatoma surgery and the importance of considering the possibility of multiple concurrent diseases in such individuals. The distinguishing features of this case are the fact that the temporal bone had disappeared, and deconstruction was complicated by infection and inflammation caused by cholesteatoma, surgical invasion, and Gorham-Stout disease. Appropriate diagnosis saved the patient from ineffective multiple surgeries for cerebrospinal fluid leakage or cholesteatoma, and improved his quality of life.

Analysis of key clinical features for achieving complete remission in stage III and IV non-small cell lung cancer patients.

BACKGROUND: Although development of immune checkpoint inhibitors and various molecular target agents has extended overall survival time (OS) in advanced non-small cell lung cancer (NSCLC), a complete cure remains rare. We aimed to identify features and treatment modalities of complete remission (CR) cases in stages III and IV NSCLC by analyzing long-term survivors whose OS exceeded 3 years. METHODS: From our hospital database, 1,699 patients, registered as lung cancer between 1st Mar 2004 and 30th Apr 2011, were retrospectively examined. Stage III or IV histologically or cytologically confirmed NSCLC patients with chemotherapy initiated during this period were enrolled. A Cox proportion hazards regression model was used. Data collection was closed on 13th Feb 2017. RESULTS: There were 164 stage III and 279 stage IV patients, including 37 (22.6%) and 51 (18.3%) long-term survivors and 12 (7.3%) and 5 (1.8%) CR patients, respectively. The long-term survivors were divided into three groups: 3 ≤ OS < 5 years, 5 years ≤ OS with tumor, and 5 years ≤ OS without tumor (CR). The median OS of these groups were 1,405, 2,238, and 2,876 days in stage III and 1,368, 2,503, and 2,643 days in stage IV, respectively. The mean chemotherapy cycle numbers were 16, 20, and 10 in stage III and 24, 25, and 5 in stage IV, respectively. In the stage III CR group, all patients received chemoradiation, all oligometastases were controlled by radiation, and none had brain metastases. Compared with non-CR patients, the stage IV CR patients had smaller primary tumors and fewer metastases, which were independent prognostic factors for OS among long-term survivors. The 80% stage IV CR patients received radiation or surgery for controlling primary tumors, and the surgery rate for oligometastases was high. Pathological findings in the stage IV CR patients revealed that numerous inflammatory cells existed around and inside resected lung and brain tumors, indicating strong immune response. CONCLUSIONS: Multiple line chemotherapies with primary and oligometastatic controls by surgery and/or radiation might achieve cure in certain advanced NSCLC. Cure strategies must be changed according to stage III or IV. This study was retrospectively registered on 16th Jun 2019 in UMIN Clinical Trials Registry (number UMIN000037078).

Plasticity of dendritic spines: Molecular function and dysfunction in neurodevelopmental disorders.

Dendritic spines are tiny postsynaptic protrusions from a dendrite that receive most of the excitatory synaptic input in the brain. Functional and structural changes in dendritic spines are critical for synaptic plasticity, a cellular model of learning and memory. Conversely, altered spine morphology and plasticity are common hallmarks of human neurodevelopmental disorders, such as intellectual disability and autism. The advances in molecular and optical techniques have allowed for exploration of dynamic changes in structure and signal transduction at single-spine resolution, providing significant insights into the molecular regulation underlying spine structural plasticity. Here, I review recent findings on: how synaptic stimulation leads to diverse forms of spine structural plasticity; how the associated biochemical signals are initiated and transmitted into neuronal compartments; and how disruption of single genes associated with neurodevelopmental disorders can lead to abnormal spine structure in human and mouse brains. In particular, I discuss the functions of the Ras superfamily of small GTPases in spatiotemporal regulation of the actin cytoskeleton and protein synthesis in dendritic spines. Multiple lines of evidence implicate disrupted Ras signaling pathways in the spine structural abnormalities observed in neurodevelopmental disorders. Both deficient and excessive Ras activities lead to disrupted spine structure and deficits in learning and memory. Dysregulation of spine Ras signaling, therefore, may play a key role in the pathogenesis of multiple neurodevelopmental disorders with distinct etiologies.

Genome editing in the mammalian brain using the CRISPR-Cas system.

Recent advances in genome editing technologies such as the clustered regularly interspaced short palindromic repeats (CRISPR)-associated endonuclease Cas9 have enabled the rapid and efficient modification of endogenous genomes in a variety of cell types, accelerating biomedical research. In particular, precise genome editing in somatic cells in vivo allows for the rapid generation of genetically modified cells in living animals and holds great promise for the possibility of directly correcting genetic defects associated with human diseases. However, because of the limited efficiency and suitability of these technologies in the brain, especially in postmitotic neurons, the practical application of genome editing technologies has been largely limited in the field of neuroscience. Recent technological advances overcome significant challenges facing genome editing in the brain and have enabled us to precisely edit the genome in both mitotic cells and mature postmitotic neurons in vitro and in vivo, providing powerful means for studying gene function and dysfunction in the brain. This review highlights the development of genome editing technologies for the brain and discusses their applications, limitations, and future challenges.

Retrospective review of previous minor leak before major subarachnoid hemorrhage diagnosed by MRI as a predictor of occurrence of symptomatic delayed cerebral ischemia.

OBJECTIVE This study attempted to determine whether a previous minor leak correlated with the occurrence of symptomatic delayed cerebral ischemia (sDCI). METHODS The authors retrospectively evaluated sDCI-related clinical features and findings from MRI, including T1-weighted imaging (T1WI)-FLAIR mismatch at the time of admission, in 151 patients admitted with subarachnoid hemorrhage (SAH) within 48 hours of ictus. RESULTS The overall incidence of sDCI was 23% (35 of 151 patients). In all subjects, multivariate analysis revealed that World Federation of Neurosurgical Societies Grades II-V, age 70 years or older, presence of rebleeding after admission, a previous minor leak before the major SAH attack as diagnosed by T1WI-FLAIR mismatch, acute infarction on diffusion-weighted imaging, and CT SAH score were significantly associated with occurrence of sDCI. In patients with no previous minor leak before major SAH as diagnosed by T1WI-FLAIR mismatch, the incidence of sDCI was only 7% (7 of 97 patients). CONCLUSIONS A previous minor leak before major SAH as diagnosed by T1WI-FLAIR mismatch represents an important sDCI-related factor. When the analysis was restricted to patients with true acute SAH without a previous minor leak diagnosed by T1WI-FLAIR mismatch, the incidence of sDCI was extremely low.

Virus-Mediated Genome Editing via Homology-Directed Repair in Mitotic and Postmitotic Cells in Mammalian Brain.

Precise genome editing via homology-directed repair (HDR) in targeted cells, particularly in vivo, provides an invaluable tool for biomedical research. However, HDR has been considered to be largely restricted to dividing cells, making it challenging to apply the technique in postmitotic neurons. Here we show that precise genome editing via HDR is possible in mature postmitotic neurons as well as mitotic cells in mice brain by combining CRISPR-Cas9-mediated DNA cleavage and the efficient delivery of donor template with adeno-associated virus (AAV). Using this strategy, we achieved efficient tagging of endogenous proteins in primary and organotypic cultures in vitro and developing, adult, aged, and pathological brains in vivo. Thus, AAV- and CRISPR-Cas9-mediated HDR will be broadly useful for precise genome editing in basic and translational neuroscience.

Occurrence of spot signs from hypodensity areas on precontrast CT in intracerebral hemorrhage.

OBJECTIVE: Both the spot signs, which is a bright spot on computed tomography angiography (CTA) source images, and hypodensity areas within a hematoma on precontrast CT scans, which presumably represent uncoagulated blood, have been reported to be predictive of hematoma enlargement in acute spontaneous intracerebral hematoma (ICH). The aim was to investigate densities on precontrast CT scans in an area within a hematoma that matched the locations of spot signs on CTA source images. METHODS: In consecutive cases of spontaneous ICH admitted within 6 h after onset, early spot signs on CTA source images and delayed spot signs on delayed-phase CT scans 90 s after CTA were evaluated. RESULTS: Of 177 patients undergoing CTA, 41 (23.2%) showed early spot signs. Among 146 patients who underwent delayed-phase CT scans, 23 (15.8%) demonstrated delayed spot signs but not early spot signs. Spot signs originated from hypodensity areas, including densities <50 HU, in 30 of 35 patients (85.7%) with early spot signs and in 8 of 23 (34.8%) with delayed spot signs. Early spot signs arose from hypodensity areas more frequently than delayed spot signs (p < 0.05). Hematoma enlargement was observed in 10 of 24 patients (41.7%) with early spot signs, but in none with delayed spot signs. DISCUSSION: Some hypodensity areas within ICHs may indicate uncoagulated blood related to ongoing leakage, which are seen as spot signs. Minimum densities in hypodensity areas might correlate with the speed and volume of bleeding.

High-Throughput, High-Resolution Mapping of Protein Localization in Mammalian Brain by In Vivo Genome Editing.

A scalable and high-throughput method to identify precise subcellular localization of endogenous proteins is essential for integrative understanding of a cell at the molecular level. Here, we developed a simple and generalizable technique to image endogenous proteins with high specificity, resolution, and contrast in single cells in mammalian brain tissue. The technique, single-cell labeling of endogenous proteins by clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9-mediated homology-directed repair (SLENDR), uses in vivo genome editing to insert a sequence encoding an epitope tag or a fluorescent protein to a gene of interest by CRISPR-Cas9-mediated homology-directed repair (HDR). Single-cell, HDR-mediated genome editing was achieved by delivering the editing machinery to dividing neuronal progenitors through in utero electroporation. We demonstrate that SLENDR allows rapid determination of the localization and dynamics of many endogenous proteins in various cell types, regions, and ages in the brain. Thus, SLENDR provides a high-throughput platform to map the subcellular localization of endogenous proteins with the resolution of micro- to nanometers in the brain.

Chronic Subdural Hematomas Associated with Arachnoid Cysts: Significance in Young Patients with Chronic Subdural Hematomas.

Although arachnoid cysts (ACs) are associated with chronic subdural hematomas (CSDHs), especially in young patients, the detailed features of CSDHs associated with ACs remain poorly understood. The objective of this study was to clarify the relationship between the location of CSDHs and ACs and the significance of ACs in young patients with CSDHs. We retrospectively assessed 605 consecutive patients 7 years of age and older who were diagnosed with a CSDH between 2002 and 2014. Twelve patients (2%) had ACs, and 10 of the 12 patients were 7-40 years of age. Patients with ACs as a complication of CSDHs were significantly younger than those without ACs (p < 0.05). Three different relationships between the location of CSDHs and ACs were found: a CSDH contacting an AC, an ipsilateral CSDH apart from an AC, and a CSDH contralateral to an AC. In 21 patients with CSDHs who were 7-40 years of age, 10 (47.6%) had ACs (AC group) and 7 (33.3%) had no associated illnesses (non-AC group). All 10 young patients with ACs showed ipsilateral CSDHs including a CSDH apart from an AC. All 17 patients in both the AC and non-AC groups showed headache but no paresis at admission. The pathogenesis of CSDHs associated with ACs may be different among the three types of locations. The clinical characteristics of patients with a combination of a CSDH and an AC including headache as a major symptom may be attributed to young age in the majority of patients with ACs.

Biochemical Computation for Spine Structural Plasticity.

The structural plasticity of dendritic spines is considered to be essential for various forms of synaptic plasticity, learning, and memory. The process is mediated by a complex signaling network consisting of numerous species of molecules. Furthermore, the spatiotemporal dynamics of the biochemical signaling are regulated in a complicated manner because of geometrical restrictions from the unique morphology of the dendritic branches and spines. Recent advances in optical techniques have enabled the exploration of the spatiotemporal aspects of the signal regulations in spines and dendrites and have provided many insights into the principle of the biochemical computation that underlies spine structural plasticity.

Density of the cerebral cortex in computed tomography angiography source images and clinical outcomes in Grade V subarachnoid hemorrhage.

OBJECTIVE: Among patients diagnosed with Grade V subarachnoid hemorrhage (SAH) according to the World Federation of Neurosurgical Societies (WFNS), the identification of those with the possibility of recovery is not feasible, although approximately one-fifth of these patients have favorable outcomes according to a recently published series of surgical papers. We hypothesized that computed tomography angiography (CTA) source images (SIs), which have applied to the detection of acute ischemia, might be useful for selecting Grade V patients with possibilities for favorable outcomes. METHODS: We retrospectively assessed 170 SAH patients who underwent surgery between January 2009 and February 2012 and quantitatively measured their mean cerebral cortical densities from the initial CTA-SIs. RESULTS: The cortical densities of 123 patients of Grades I-IV were strongly affected by the following two CTA-SI findings that were revealed by multivariate analysis: the density of the internal carotid artery (ICA) just proximal to the petrous portion and the ratio of the subarachnoid (SA) space to intracranial volume (P < 0.05). Favorable outcomes were obtained in 9 of the 47 Grade V patients (19.1%), and the predictors of the favorable outcomes according to multivariate analysis were increased cortical densities on the CTA-SIs and female patients (P < 0.05). CONCLUSION: The outcomes of Grade V patients could be partly predicted based on the estimates of cortical density from the CTA-SIs, and this measure might be useful for the selection of Grade V patients for surgery, provided that both ICA density and SA space ratio are confirmed before the evaluation of cortical density.

Wide-area scanner for high-speed atomic force microscopy.

High-speed atomic force microscopy (HS-AFM) has recently been established. The dynamic processes and structural dynamics of protein molecules in action have been successfully visualized using HS-AFM. However, its maximum scan ranges in the X- and Y-directions have been limited to ~1 µm and ~4 µm, respectively, making it infeasible to observe the dynamics of much larger samples, including live cells. Here, we develop a wide-area scanner with a maximum XY scan range of ~46 × 46 µm(2) by magnifying the displacements of stack piezoelectric actuators using a leverage mechanism. Mechanical vibrations produced by fast displacement of the X-scanner are suppressed by a combination of feed-forward inverse compensation and the use of triangular scan signals with rounded vertices. As a result, the scan speed in the X-direction reaches 6.3 mm/s even for a scan size as large as ~40 µm. The nonlinearity of the X- and Y-piezoelectric actuators' displacements that arises from their hysteresis is eliminated by polynomial-approximation-based open-loop control. The interference between the X- and Y-scanners is also eliminated by the same technique. The usefulness of this wide-area scanner is demonstrated by video imaging of dynamic processes in live bacterial and eukaryotic cells.

Two-dimensional differential calibration method for a neutron dosemeter using a thermal neutron beam.

A new thermal neutron calibration method to experimentally determine the energy response function of a neutron detector using a pulse parallel beam and the time-of-flight (TOF) technique is developed. The calibration method was experimentally demonstrated for a (3)He proportional counter and an electric personal dosemeter using a pulsed thermal neutron beam from the research reactor JRR-3M. The responses of the detectors were successfully obtained as a function of neutron energy. However, detailed information on the detector structure is required to obtain the spatial response distribution for the detector. The authors further propose an improved calibration method obtaining the spatial response distribution using a pulsed narrow beam, the TOF technique and a beam scanning technique.

Selective and regulated gene expression in murine Purkinje cells by in utero electroporation.

Cerebellar Purkinje cells, which convey the only output from the cerebellar cortex, play an essential role in cerebellar functions, such as motor coordination and motor learning. To understand how Purkinje cells develop and function in the mature cerebellum, an efficient method for molecularly perturbing them is needed. Here we demonstrate that Purkinje cell progenitors at embryonic day (E)11.5 could be efficiently and preferentially transfected by spatially directed in utero electroporation (IUE) with an optimized arrangement of electrodes. Electrophysiological analyses indicated that the electroporated Purkinje cells maintained normal membrane properties, synaptic responses and synaptic plasticity at postnatal days 25-28. By combining the L7 promoter and inducible Cre/loxP system with IUE, transgenes were expressed even more specifically in Purkinje cells and in a temporally controlled manner. We also show that three different fluorescent proteins could be simultaneously expressed, and that Bassoon, a large synaptic protein, could be expressed in the electroporated Purkinje cells. Moreover, phenotypes of staggerer mutant mice, which have a deletion in the gene encoding retinoid-related orphan receptor α (RORα1), were recapitulated by electroporating a dominant-negative form of RORα1 into Purkinje cells at E11.5. Together, these results indicate that this new IUE protocol, which allows the selective, effective and temporally regulated expression of multiple foreign genes transfected into Purkinje cell progenitors in vivo, without changing the cells' physiological characteristics, is a powerful tool for elucidating the molecular mechanisms underlying early Purkinje cell developmental events, such as dendritogenesis and migration, and synaptic plasticity in mature Purkinje cells.

Glioblastoma associated with intratumoral abscess formation. Case report.

A 45-year-old man presented with a rare case of glioblastoma associated with intratumoral abscess formation manifesting as headache and vomiting after an appendectomy. Computed tomography and magnetic resonance imaging demonstrated a ring-enhanced lesion mimicking malignant glioma. Craniotomy and tumor removal were performed. Abscess formation within the intra-axial tumor was found intraoperatively. Histological examination revealed glioblastoma with abscess, and the etiological agent was anaerobic Gram-negative bacilli. The suspected route of microbial migration and colonization in this tumor was bacteremia from appendicitis.