Glutamate acts on acid-sensing ion channels to worsen ischaemic brain injury.

Glutamate is traditionally viewed as the first messenger to activate NMDAR (N-methyl-D-aspartate receptor)-dependent cell death pathways in stroke1,2, but unsuccessful clinical trials with NMDAR antagonists implicate the engagement of other mechanisms3-7. Here we show that glutamate and its structural analogues, including NMDAR antagonist L-AP5 (also known as APV), robustly potentiate currents mediated by acid-sensing ion channels (ASICs) associated with acidosis-induced neurotoxicity in stroke4. Glutamate increases the affinity of ASICs for protons and their open probability, aggravating ischaemic neurotoxicity in both in vitro and in vivo models. Site-directed mutagenesis, structure-based modelling and functional assays reveal a bona fide glutamate-binding cavity in the extracellular domain of ASIC1a. Computational drug screening identified a small molecule, LK-2, that binds to this cavity and abolishes glutamate-dependent potentiation of ASIC currents but spares NMDARs. LK-2 reduces the infarct volume and improves sensorimotor recovery in a mouse model of ischaemic stroke, reminiscent of that seen in mice with Asic1a knockout or knockout of other cation channels4-7. We conclude that glutamate functions as a positive allosteric modulator for ASICs to exacerbate neurotoxicity, and preferential targeting of the glutamate-binding site on ASICs over that on NMDARs may be strategized for developing stroke therapeutics lacking the psychotic side effects of NMDAR antagonists.

Bioassay-Guided Isolation and Identification of Xanthine Oxidase Inhibitory Constituents from the Leaves of Perilla frutescens.

Activity-directed fractionation and purification processes were employed to identify xanthine oxidase (XO) inhibitory compounds from the leaves of Perilla frutescens. The total extract was evaluated in vitro on XO inhibitory activity and in vivo in an experimental model with potassium oxonate-induced hyperuricemia in mice which was used to evaluate anti-hyperuricemic activity. The crude extract showed expressive urate-lowering activity results. Solvent partitioning of the total extract followed by macroporous resin column chromatography of the n-butanol extract yielded four extracts and eluted parts. Among them, only the 70% ethanol eluted part of the n-butanol extract showed strong activity and therefore was subjected to separation and purification using various chromatographic techniques. Five compounds showing potent activity were identified by comparing their spectral data with literature values to be caffeic acid, vinyl caffeate, rosmarinic acid, methyl rosmarinate, and apigenin. These results indicate that pending further study, these compounds could be used as novel natural product agents for the treatment of hyperuricemia.

Electroencephalography microstate alterations reflect potential double-edged cognitive adaptation in Ménière's disease.

PURPOSE: To explore the microstate characteristics and underlying brain network activity of Ménière's disease (MD) patients based on high-density electroencephalography (EEG), elucidate the association between microstate dynamics and clinical manifestation, and explore the potential of EEG microstate features as future neurobiomarkers for MD. METHODS: Thirty-two patients diagnosed with MD and 29 healthy controls (HC) matched for demographic characteristics were included in the study. Dysfunction and subjective symptom severity were assessed by neuropsychological questionnaires, pure tone audiometry, and vestibular function tests. Resting-state EEG recordings were obtained using a 256-channel EEG system, and the electric field topographies were clustered into four dominant microstate classes (A, B, C, and D). The dynamic parameters of each microstate were analyzed and utilized as input for a support vector machine (SVM) classifier to identify significant microstate signatures associated with MD. The clinical significance was further explored through Spearman correlation analysis. RESULTS: MD patients exhibited an increased presence of microstate class C and a decreased frequency of transitions between microstate class A and B, as well as between class A and D. The transitions from microstate class A to C were also elevated. Further analysis revealed a positive correlation between equilibrium scores and the transitions from microstate class A to C under somatosensory challenging conditions. Conversely, transitions between class A and B were negatively correlated with vertigo symptoms. No significant correlations were detected between these characteristics and auditory test results or emotional scores. Utilizing the microstate features identified via sequential backward selection, the linear SVM classifier achieved a sensitivity of 86.21% and a specificity of 90.61% in distinguishing MD patients from HC. CONCLUSIONS: We identified several EEG microstate characteristics in MD patients that facilitate postural control yet exacerbate subjective symptoms, and effectively discriminate MD from HC. The microstate features may offer a new approach for optimizing cognitive compensation strategies and exploring potential neurobiological markers in MD.

Mechanisms underlying taurine protection against glutamate-induced neurotoxicity.

Taurine appears to exert potent protections against glutamate (Glu)-induced injury to neurons, but the underlying molecular mechanisms are not fully understood. The possibly protected targets consist of the plasma membrane and the mitochondrial as well as endoplasmic reticulum (ER) membranes. Protection may be provided through a variety of effects, including the prevention of membrane depolarization, neuronal excitotoxicity and mitochondrial energy failure, increases in intracellular free calcium ([Ca2+]i), activation of calpain, and reduction of Bcl-2 levels. These activities are likely to be linked spatially and temporally in the neuroprotective functions of taurine. In addition, events that occur downstream of Glu stimulation, including altered enzymatic activities, apoptotic pathways, and necrosis triggered by the increased [Ca2+]i, can be inhibited by taurine. This review discusses the possible molecular mechanisms of taurine against Glu-induced neuronal injury, providing a better understanding of the protective processes, which might be helpful in the development of novel interventional strategies.

Synergistic effects of psyllium husk powder and different levels of methylcellulose on the storage stability of sodium caseinate emulsion.

The study investigated the effects of compound fibers composed of psyllium husk powder (PHP, 0.3%) and methylcellulose (MC, 0, 0.3, 0.6, 0.9, and 1.2%) on the storage stability, rheology, and microstructure of sodium caseinate emulsions. Results showed that the emulsion stability was enhanced with the increased concentrations of MC, especially at the concentration of 1.2%. The oil droplet size in the emulsions was decreased as the concentrations of compound fibers increased, which was further confirmed by the optical microscope analysis. The rheological measurements and cryo-scanning electron microscopy results indicated that compound fibers improved the viscosity of the emulsions, and formed a strong three-dimensional network structure. The results of confocal laser scanning microscope and surface protein concentration measurements showed that compound fibers were evenly distributed into the oil droplet surface. The above results demonstrate that compound fibers are an effective thickener and emulsifier in enhancing the stability properties of oil-in-water (O/W) emulsions stabilized by sodium caseinate.

Measuring endogenous levels of unconjugated bilirubin released from isolated murine brain tissue during oxygen-glucose deprivation.

Quantitative assessment of endogenously synthesized and released bilirubin from brain tissue remains a challenge. Here, we present a sensitive and reproducible experimental paradigm to quantify, in real time, unconjugated bilirubin (UCB) from isolated murine brain tissue during oxygen-glucose deprivation (OGD). We describe steps for perfusion, brain dissection, brain slice preparation and incubation, glucose depletion, and OGD processing. We then detail procedures for standard calibration plotting and sample UCB measurement. For complete details on the use and execution of this protocol, please refer to Liu et al.1.

Surface Functional Modification by Ti3 C2 Tx MXene on PLLA Nanofibers for Optimizing Neural Stem Cell Engineering.

Optimizing cell substrates by surface modification of neural stem cells (NSCs), for efficient and oriented neurogenesis, represents a promising strategy for treating neurological diseases. However, developing substrates with the advanced surface functionality, conductivity, and biocompatibility required for practical application is still challenging. Here, Ti3 C2 Tx MXene is introduced as a coating nanomaterial for aligned poly(l-lactide) (PLLA) nanofibers (M-ANF) to enhance NSC neurogenesis and simultaneously tailor the cell growth direction. Ti3 C2 Tx MXene treatment provides a superior conductivity substrate with a surface rich in functional groups, hydrophilicity, and roughness, which can provide biochemical and physical cues to support NSC adhesion and proliferation. Moreover, Ti3 C2 Tx MXene coating significantly promotes NSC differentiation into both neurons and astrocytes. Interestingly, Ti3 C2 Tx MXene acts synergistically with the alignment of nanofibers to promote the growth of neurites, indicating enhanced maturation of these neurons. RNA sequencing analysis further reveals the molecular mechanism by which Ti3 C2 Tx MXene modulates the fate of NSCs. Notably, surface modification by Ti3 C2 Tx MXene mitigates the in vivo foreign body response to implanted PLLA nanofibers. This study confirms that Ti3 C2 Tx MXene provides multiple advantages for decorating the aligned PLLA nanofibers to cooperatively improve neural regeneration.

Bilirubin gates the TRPM2 channel as a direct agonist to exacerbate ischemic brain damage.

Stroke prognosis is negatively associated with an elevation of serum bilirubin, but how bilirubin worsens outcomes remains mysterious. We report that post-, but not pre-, stroke bilirubin levels among inpatients scale with infarct volume. In mouse models, bilirubin increases neuronal excitability and ischemic infarct, whereas ischemic insults induce the release of endogenous bilirubin, all of which are attenuated by knockout of the TRPM2 channel or its antagonist A23. Independent of canonical TRPM2 intracellular agonists, bilirubin and its metabolic derivatives gate the channel opening, whereas A23 antagonizes it by binding to the same cavity. Knocking in a loss of binding point mutation for bilirubin, TRPM2-D1066A, effectively antagonizes ischemic neurotoxicity in mice. These findings suggest a vicious cycle of stroke injury in which initial ischemic insults trigger the release of endogenous bilirubin from injured cells, which potentially acts as a volume neurotransmitter to activate TRPM2 channels, aggravating Ca2+-dependent brain injury.

Bilirubin induces auditory neuropathy in neonatal guinea pigs via auditory nerve fiber damage.

Bilirubin can cause temporary or permanent sensorineural deafness in newborn babies with hyperbilirubinemia. However, the underlying targets and physiological effects of bilirubin-induced damage in the peripheral auditory system are unclear. Using cochlear functional assays and electron microscopy imaging of the inner ear in neonatal guinea pigs, we show here that bilirubin exposure resulted in threshold elevation in both compound action potential (CAP) and auditory brainstem response (ABR), which was apparent at 1 hr and peaked 8 hr after drug administration. The threshold elevation was associated with delayed wave latencies and elongated interwave intervals in ABR and CAP. At 72 hr postinjection, these measures returned to control levels, except for the CAP amplitude. Cochlear microphonics remained unchanged during the experiment. Morphological abnormalities were consistent with the electrophysiological dysfunction, revealing fewer auditory nerve fibers (ANFs) in the basal turn, myelin sheath lesions of spiral ganglion neurons (SGNs) and ANFs, and loss of type 1 afferent endings beneath inner hair cells (IHCs) without loss of hair cells at 8 hr posttreatment. Similar to the electrophysiological findings, morphological changes were mostly reversed 10 days after treatment, except for the ANF reduction in the basal turn. These results suggest that hyperbilirubinemia in neonatal guinea pigs impaired auditory peripheral neuromechanisms that targeted mainly the IHC synapses and the myelin sheath of SGNs and their fibers. Our observations indicate a potential connection between hyperbilirubinemia and auditory neuropathy.

Electroencephalography Microstate Alterations in Otogenic Vertigo: A Potential Disease Marker.

Objectives: A huge population, especially the elderly, suffers from otogenic vertigo. However, the multi-modal vestibular network changes, secondary to periphery vestibular dysfunction, have not been fully elucidated. We aim to identify potential microstate electroencephalography (EEG) signatures for otogenic vertigo in this study. Materials and Methods: Patients with recurrent otogenic vertigo and age-matched healthy adults were recruited. We performed 256-channel EEG recording of all participants at resting state. Neuropsychological questionnaires and vestibular function tests were taken as a measurement of patients' symptoms and severity. We clustered microstates into four classes (A, B, C, and D) and identified their dynamic and syntax alterations of them. These features were further fed into a support vector machine (SVM) classifier to identify microstate signatures for vertigo. Results: We compared 40 patients to 45 healthy adults, finding an increase in the duration of Microstate A, and both the occurrence and time coverage of Microstate D. The coverage and occurrence of Microstate C decreased significantly, and the probabilities of non-random transitions between Microstate A and D, as well as Microstate B and C, also changed. To distinguish the patients, the SVM classifier, which is built based on these features, got a balanced accuracy of 0.79 with a sensitivity of 0.78 and a specificity of 0.8. Conclusion: There are several temporal dynamic alterations of EEG microstates in patients with otogenic vertigo, especially in Microstate D, reflecting the underlying process of visual-vestibular reorganization and attention redistribution. This neurophysiological signature of microstates could be used to identify patients with vertigo in the future.

Synthesis of novel thiazolyl-pyrimidines and their anticancer activity in vitro.

A series of novel compounds 7-43 were prepared via the condensation of enaminones 4a-h and the guanidines carbonate 6a-f. The structures of these newly synthesized compounds were confirmed by (1) H-NMR, MS, EA and IR. All the compounds were tested for their cytotoxic activity in vitro against human cancer cell lines including Ishikawa, A549, BEL-7404, SPC-A-01 and SGC-7901. Most of them showed moderate cytotoxic against the tested cell lines. Among them, the most potent compounds 9 and 30 exhibited more efficient activity against Ishikawa, A549.

4-{4-Methyl-2-[(meth-yl)(2-methyl-phen-yl)amino]-1,3-thia-zol-5-yl}-N-(3-methyl-phen-yl)pyrimidin-2-amine.

In the title compound, C(23)H(23)N(5)S, the thia-zole ring and pyrimidine ring are almost coplanar, making a dihedral angle of 4.02 (9)°. in the crystal, weak inter-molecular N-H⋯N inter-actions link pairs of molecules into centrosymmetric dimers.

4-{2-[(3,4-Dichloro-phen-yl)(meth-yl)amino]-4-methyl-1,3-thia-zol-5-yl}-N-(3-methyl-phen-yl)pyrimidin-2-amine.

In the title compound, C(22)H(19)Cl(2)N(5)S, the thia-zole and pyrimidine rings are almost co-planar, making a dihedral angle of 6.48 (7)°. In the crystal, intermolecular N-H⋯N hydrogen bonds link pairs of molecules into centrosymmetric dimers..

N-(3-Meth-oxy-phen-yl)-4-{4-methyl-2-[(meth-yl)(4-methyl-phen-yl)amino]-1,3-thia-zol-5-yl}pyrimidin-2-amine.

The asymmetric unit of the title compound, C(23)H(23)N(5)OS, contains two independent mol-ecules. In one mol-ecule, the thia-zole and pyrimidine rings are almost co-planar, making a dihedral angle of 2.48 (8)°. In the other mol-ecule, the corresponding dihedral angle is 12.82 (8)°. The crystal structure is stabilized by weak inter-molecular N-H⋯N and C-H⋯O inter-actions that extend along the b axis.

Stepwise Induction of Inner Ear Hair Cells From Mouse Embryonic Fibroblasts via Mesenchymal- to-Epithelial Transition and Formation of Otic Epithelial Cells.

Although embryonic stem cells or induced pluripotent stem cells are able to differentiate into inner ear hair cells (HCs), they have drawbacks limiting their clinical application, including a potential risk of tumourigenicity. Direct reprogramming of fibroblasts to inner ear HCs could offer an alternative solution to this problem. Here, we present a stepwise guidance protocol to induce mouse embryonic fibroblasts to differentiate into inner ear HC-like cells (HCLs) via mesenchymal-to-epithelial transition and then acquisition of otic sensory epithelial cell traits by overexpression of three key transcription factors. These induced HCLs express multiple HC-specific proteins, display protrusions reminiscent of ciliary bundle structures, respond to voltage stimulation, form functional mechanotransduction channels, and exhibit a transcriptional profile of HC signature. Together, our work provides a new method to produce functional HCLs in vitro, which may have important implications for studies of HC development, drug discovery, and cell replacement therapy for hearing loss.

Multisensory Exercise Improves Balance in People with Balance Disorders: A Systematic Review.

OBJECTIVE: To examine the effect of multisensory exercise on balance disorders. METHODS: PubMed, Scopus and Web of Science were searched to identify eligible studies published before January 1, 2020. Eligible studies included randomized control trials (RCTs), non-randomized studies, case-control studies, and cohort studies. The methodological quality of the included studies was evaluated using JBI Critical Appraisal Checklists for RCTs and for Quasi-Experimental Studies by two researchers independently. A narrative synthesis of intervention characteristics and health-related outcomes was performed. RESULTS: A total of 11 non-randomized studies and 9 RCTs were eligible, including 667 participants. The results supported our assumption that multisensory exercise improved balance in people with balance disorders. All of the 20 studies were believed to be of high or moderate quality. CONCLUSION: Our study confirmed that multisensory exercise was effective in improving balance in people with balance disorders. Multisensory exercises could lower the risk of fall and enhance confidence level to improve the quality of life. Further research is needed to investigate the optimal strategy of multisensory exercises and explore the underlying neural and molecular mechanisms of balance improvement brought by multisensory exercises.

Synthesis and anticancer evaluation of thiazolyl-chalcones.

Thirty-seven (E)-1-(4-methyl-2-arylaminothiazol-5-yl)-3-arylprop-2-en-1-ones were synthesized via Claisen-Schmidt condensation of 1-(4-methyl-2-(arylamino)thiazol-5-yl)ethanone with the corresponding arylaldehydes. All these thiazolyl-chalcones were characterized and evaluated by MTT assay on human cancer cell lines BGC-823, PC-3, NCI-H460, BEL-7402 in vitro. Compounds 5, 8, 26, 37 and 41 are effective against cancer cell lines with IC(50)s below 10 µM. The antitumor activity in ICR mice bearing sarcoma 180 tumors indicates compounds 10 and 41 have moderate in vivo activity with 22-25% tumor-weight inhibition.

Fibula osteal flap with proximal peroneal perforator skin paddle for composite oromandibular reconstruction: A case report.

RATIONALE: Cutaneous perforators of peroneal vessels are divided into proximal and distal perforators on the basis of perforator distributions and musculocutaneous or septocutaneous properties. The traditional fibular osteocutaneous free flap is raised over the distal two-thirds of the fibula with a skin paddle based on distal perforators, which is affixed to the posterior crural septum. However, the skin pedicle may not be available due to anatomic variations or intraoperative injuries. Herein, because of the absence of distal perforators, we reserved and expropriated proximal perforators originating from the musculocutaneous branch of the superior part of the peroneal artery before it divided into nutrient and arcuate arteries and successfully harvested a separate osteal fibula and proximal perforator skin paddle with a single vascular pedicle-peroneal vessel. PATIENT CONCERNS: A 62-year-old man with a 6-month history of mandibular swelling and soft tissue invasion was referred to us. DIAGNOSIS: Panoramic radiography and computed tomography showed an irregular radiolucent lesion of the mandibular body, and histopathological analysis confirmed a follicular-pattern ameloblastoma. INTERVENTIONS: The diseased mandible and soft tissue were resected and reconstructed with a vascularized fibular osteal flap with the proximal perforator skin paddle. OUTCOMES: The mandibular contour was successfully restored; the skin paddle in the mouth was in good condition after 8 months of follow-up. LESSONS: The proximal perforator is reliable and practical for supplying a skin paddle and has significant potential for future applications. We recommend reserving the proximal perforator skin paddle as a backup flap when planning to raise a fibula flap, since unavailability or injury of the traditional fibular skin island based on distal perforators occurs frequently. This approach can avoid the exploration for a second donor site, save surgical time, and reduce surgical complexity. Moreover, we anticipate more frequent use of the proximal perforator flap in the future because of its flexibility and large volume, and since it can be combined with the osteal fibula or fibular osteocutaneous flap. However, an understanding of the traits of the proximal perforator and determination of its peroneal origin by computed tomography angiography is crucial for predesigning fibular osteal flaps with a proximal perforator skin paddle.

Removal of biofilm is essential for long-term ventilation tube retention.

BACKGROUND: Although long-term retention of a ventilation tube is required in many ear diseases, spontaneous removal of conventional ventilation tube is observed in patients within 3 to 12 mo. To address this issue, we aimed to determine a new method for long-term retention of the ventilation tube. AIM: To explore the value of removing the biofilm for long-term retention of tympanostomy ventilation tubes. METHODS: A case-control study design was used to evaluate the safety and effectiveness of long-term tube retention by directly removing the biofilm (via surgical exfoliation) in patients who underwent myringotomy with ventilation tube placement. The patients were randomly divided into two groups: Control group and treatment group. Patients in the treatment group underwent regular biofilm exfoliation surgery in the clinic, whereas those in the control group did not have their biofilm removed. Only conventional ventilation tubes were placed in this study. Outcome measures were tube position and patency. Tube retention time and any complications were documented. RESULTS: Eight patients with biofilm removal and eight patients without biofilm removal as a control group were enrolled in the study. The tympanostomy tube retention time was significantly longer in the treatment group (43.5 ± 26.4 mo) than in the control group (9.5 ± 6.9 mo) (P = 0.003). More tympanostomy tubes were found to be patent and in correct position in the treatment group during the follow-up intervals than in the control group (P = 0.01). CONCLUSION: Despite the use of short-term ventilation tubes, direct biofilm removal can be a well-tolerated and effective treatment for long-term tube retention of tympanostomy ventilation tubes in patients who underwent myringotomy.

LncRNA PEAMIR inhibits apoptosis and inflammatory response in PM2.5 exposure aggravated myocardial ischemia/reperfusion injury as a competing endogenous RNA of miR-29b-3p.

The sensitivity of myocardium is enhanced to ischemia/reperfusion (I/R) injury under PM2.5 exposure. It is still under prelude for lncRNA-miRNA pair in the study of aggravated myocardial I/R injury under PM2.5 exposure. In this study, we first built a rat model of 30 min ischemia and 24 h reperfusion followed PM2.5 (6.0 mg/kg) exposure. We found PM2.5 exposure could obviously aggravate I/R injury in the fields of myocardium damage, apoptosis levels and cardiac function which were evaluated by TTC staining, TUNEL and echocardiography, respectively. Then, based on results of sequencing and RT-qPCR, we selected NONRATT003473.2 in the follow-up experiments and named this lncRNA as PM2.5 exposure aggravated myocardial I/R injury lncRNA (PEAMIR). Consistent with the results rat model, we confirmed PEAMIR to be a protective lncRNA against PM + HR triggered damages in H9c2 cells. Next, according to the bioinformatics analysis from miRanda database and a series of gain- and loss-of-function experiments, we proved PEAMIR to be a ceRNA for miR-29b-3p to inhibit cardiac inflammation and apoptosis. Finally, using Target-Scan database, the conserved binding sites for miR-29b-3p was identified in the 3'UTR of PI3K (p85a), a key protein of apoptosis. Our subsequent experiments validated the regulatory relationship between PEAMIR-miR-29b-3p ceRNA pair and PI3K (p85a)/Akt/GSK3b/p53 cascade pathway. In conclusion, our study demonstrated the role and mechanism of PEAMIR in the augment of I/R injury under PM2.5 exposure, suggesting a promising strategy for the prevention and treatment of I/R injury under PM2.5 exposure.