Utility of labial salivary gland biopsy in the histological diagnosis of neuronal intranuclear inclusion disease.

BACKGROUND AND PURPOSE: Neuronal intranuclear inclusion disease (NIID) poses a diagnostic challenge because of its diverse clinical manifestations. Detection of intranuclear inclusions remains the primary diagnostic criterion for NIID. Skin biopsies have traditionally been used, but concerns exist regarding postoperative complications and scarring. We sought to investigate the diagnostic utility of labial salivary gland biopsy, a less invasive alternative. METHODS: This study included a total of 19 patients and 11 asymptomatic carriers who underwent labial gland biopsies, while 10 patients opted for skin biopsies. All these individuals were confirmed to have pathogenic GGC repeat expansions in the NOTCH2NLC gene. The control group comprised 20 individuals matched for age and sex, all with nonpathogenic GGC repeat expansions, and their labial gland tissue was sourced from oral surgery specimens. RESULTS: Labial gland biopsies proved to be a highly effective diagnostic method in detecting eosinophilic intranuclear inclusions in NIID patients. The inclusions showed positive staining for p62 and ubiquitin, confirming their pathological significance. The presence of uN2CpolyG protein in the labial gland tissue further supported the diagnosis. Importantly, all patients who underwent lip gland biopsy experienced fast wound healing without any noticeable scarring. In contrast, skin biopsies led to varying degrees of scarring and one instance of a localized infection. CONCLUSION: Labial salivary gland biopsy emerged as a minimally invasive, efficient diagnostic method for NIID, with rapid healing and excellent sensitivity.

Palladium-Catalyzed Iodine Assisted Carbonylation of Indoles with ClCF2CO2Na and Alcohols.

A palladium-catalyzed iodine-assisted carbonylation reaction of indoles with readily available ClCF2CO2Na and alcohols has been developed. This protocol provides a practical and efficient approach to highly regioselective indole-3-carboxylates via a preiodination strategy of indoles. Different from classic carbonylation using toxic and difficult-to-handle carbon monoxide, this operationally simple and scalable reaction employed difluorocarbene as the carbonyl surrogate.

Current advances in neuronal intranuclear inclusion disease.

Neuronal intranuclear inclusion disease (NIID) is a rare but probably underdiagnosed neurodegenerative disorder due to pathogenic GGC expansions in the NOTCH2NLC gene. In this review, we summarize recent developments in the inheritance features, pathogenesis, and histopathologic and radiologic features of NIID that subvert the previous perceptions of NIID. GGC repeat sizes determine the age of onset and clinical phenotypes of NIID patients. Anticipation may be absent in NIID but paternal bias is observed in NIID pedigrees. Eosinophilic intranuclear inclusions in skin tissues once considered pathological hallmarks of NIID can also present in other GGC repeat diseases. Diffusion-weighted imaging (DWI) hyperintensity along the corticomedullary junction once considered the imaging hallmark of NIID can frequently be absent in muscle weakness and parkinsonism phenotype of NIID. Besides, DWI abnormalities can appear years after the onset of predominant symptoms and may even disappear completely with disease progression. Moreover, continuous reports of NOTCH2NLC GGC expansions in patients with other neurodegenerative diseases lead to the proposal of a new concept of NOTCH2NLC-related GGC repeat expansion disorders (NRED). However, by reviewing the previous literature, we point out the limitations of these studies and provide evidence that these patients are actually suffering from neurodegenerative phenotypes of NIID.

Palladium-Catalyzed Carbonylative [5+1] Cycloaddition of N-Tosyl Vinylaziridines: Solvent-Controlled Divergent Synthesis of α,ß- and ß,γ-Unsaturated δ-Lactams.

A palladium-catalyzed carbonylative [5+1] cycloaddition of N-tosyl vinylaziridines with CO has been developed. This protocol affords an efficient and practical approach for solvent-controlled divergent synthesis of α,ß-unsaturated δ-lactams in dimethylformamide and ß,γ-unsaturated δ-lactams in tetrahydrofuran in good to excellent yields. Significantly, the step- and atom-economical reactions are more regioselective toward [5+1] cycloaddition than toward [3+1] cycloaddition.

Rap2B promotes cell adhesion, proliferation, migration and invasion of human glioma.

PURPOSE: Rap2B, a member of the GTP-binding proteins, is generally up-regulated in numerous types of tumors. Nevertheless, the influence and regulatory mechanisms of Rap2B in gliomas are still not corroborated. Therefore, we analyzed the expression of Rap2B in glioma tissues and cells, and researched its significance in adhesion, proliferation, migration and invasion of the glioma cell line. METHODS: We analyzed the expression of Rap2B in different pathologic grades of glioma tissues by tissue microarray and immunohistochemistry. We assessed the expression of Rap2B in glioma tissue and non-tumor tissue by Western blot. And the expression of Rap2b protein in glioma cells and normal human astrocytes (NHA) was detected by Western blot. In addition, we disclosed the effect of Rap2B knockdown on cell adhesion, proliferation, migration and invasion by using cell attachment assay, CCK-8 assay, cell migration assay and Wound Healing assay, cell invasion assay, respectively. Western blot was used to detect the changes of expression level of NF-kB, MMP-2 and MMP-9 protein when downregulated the expression of Rap2B. RESULTS: The tissue microarray immunohistochemical results of glioma showed that the expression of Rap2B had no significant correlations between Rap2B expression and the clinicopathologic variables, including patient age (P = 0.352), gender (P = 0.858), WHO Grade (P = 0.693) and histology type (P = 0.877). Western blot analysis showed that the glioma tissue had a dramatically increase of Rap2B expression compared with the non-tumor tissues (P < 0.01). And the expression of Rap2B was markedly up-regulated in all 5 glioma cell lines compared with that in normal human astrocytes (NHA) (P < 0.01). We found that the ability of adhesion, proliferation, migration and invasion of glioma cells were significantly decreased after downregulated Rap2B expression compared with the control group (P < 0.05). In addition, Western blot results showed that the expression levels of NF-kB, MMP-2 and MMP-9 in the interference group were significantly lower than those in the negative control group (P < 0.05). CONCLUSIONS: Rap2B expression is up-regulated in glioma tissues and glioma cell lines. Knockdown of Rap2B inhibits glioma cells' adhesion and proliferation in vitro. Knockdown of Rap2B inhibits glioma cells' migration in vitro. Knockdown of Rap2B inhibits glioma cells' invasion and MMPs activity through NF-kB pathway.

Winter plant phenology in the alpine meadow on the eastern Qinghai-Tibetan Plateau.

Background and Aims: There is no knowledge of winter plant phenology and its controlling factors on the Qinghai-Tibetan Plateau (QTP). Thus, we conducted a 4 year winter phenology and growth dynamics study in the alpine meadow on the eastern QTP. Methods: From November 2013 to March 2017, the phenology of the 'winter-growth' and 'winter-green' species was recorded every 5 d. In November-February from 2014 to 2015, the above-ground biomass (AGB) in random plots was calculated to distinguish different growth patterns among winter growing species. The percentage of winter abundance relative to the summer population for forbs and the percentage of absolute coverage for grasses (W/S) were calculated to describe the importance of the winter population to the summer population. The soil moisture (SM) and soil temperature (ST) were used to explore the controlling factors on the AGB. Pearson's correlation analysis between winter phenology data and environmental variables, including air temperature (Tair), snow cover fraction (SCF), SM and ST, was used to investigate the factors affecting winter phenology during November-February from 2014 to 2017. Key Results: There were 107 species in total in the sites, including ten 'winter-growth' species and four 'winter-green' species. Among the 'winter-green' species, Festuca ovina and Deschampsia cespitosa were the dominant species in the sites. The 'winter-growth' species grew new leaves or ramets or transitioned to reproductive growth. Gentiana spathulifolia even flowered in winter. 'Winter-growth' made important contributions to the annual AGB, e.g. winter growth of G. spathulifolia accounted for 23.26 % of its annual AGB, while 14.74 % of the annual AGB of G. crassuloides was from winter growth. In addition, winter warming and snowfall reduction under global climate change on the eastern QTP may decrease the AGB increment of the 'winter-growth' and delay the green-up onset date of 'winter-green' species. Also, winter warming and snowfall reduction may advance the first flowering date of 'winter-growth' species. Conclusions: In contrast to previous views that plants on the QTP were generally considered to remain dormant in winter, our study revealed that alpine meadow plants had strong winter growth which suggested the importance of re-evaluating the dynamics of ecosystem function of alpine meadow, including its contribution to the global carbon balance. It was also shown that soil moisture availability is more important than warmer temperature in controlling the green-up onset of 'winter-green' species on the eastern QTP, which contrasts with the traditional view that warmer winters could advance green-up. As snowmelt is the only source of soil water in winter, the prediction of the green-up trend may be further complicated by snowfall variation in winter.

ROS/TXNIP pathway contributes to thrombin induced NLRP3 inflammasome activation and cell apoptosis in microglia.

There is no effective therapy for intracerebral hemorrhage (ICH) because of poor understanding of the mechanisms of brain injury after hemorrhage. The NLRP3 inflammasome, as a vital component of innate immune system, which is associated with a wide range of human CNS disorders, including ICH. But its detailed mechanisms in ICH remain mainly unclear. In this study, BV2 cells with thrombin exposure were used to investigate the role of NLRP3 inflammasome in thrombin-induced brain injury. We used western blot to detect NLRP3 inflammasome activation and the expression of thioredoxin binding protein (TXNIP), DCFH-DA to investigate intracellular reactive oxygen species (ROS), flow cytometry to analyze apoptosis. Our results showed that ROS inhibitor N-acetyl-l-cysteine (NAC) suppressed the upregulation of intracellular ROS and TXNIP expression. Furthermore, the cell apoptosis and expression of apoptotic protein were significantly attenuated after treatment of thrombin with NAC or NLRP3 antagonist (MCC950). Thrombin activates ROS/TXNIP/NLRP3 signaling in BV2 cells, which may indicate a mechanism that pro-inflammatory and pro-apoptotic contributes to the development of ICH.

Remote limb ischemic postconditioning inhibits microglia pyroptosis by modulating HGF after acute ischemia stroke.

The repetitive inflation-deflation of a blood pressure cuff on a limb is known as remote limb ischemic postconditioning (RIPostC). It prevents brain damage induced by acute ischemia stroke (AIS). Pyroptosis, executed by the pore-forming protein gasdermin D (GSDMD), is a type of regulated cell death triggered by proinflammatory signals. It contributes to the pathogenesis of ischemic brain injury. However, the effects of RIPostC on pyroptosis following AIS remain largely unknown. In our study, linear correlation analysis confirmed that serum GSDMD levels in AIS patients upon admission were positively correlated with NIHSS scores. RIPostC treatment significantly reduced GSDMD level compared with patients without RIPostC at 3 days post-treatment. Besides, middle cerebral artery occlusion (MCAO) surgery was performed on C57BL/6 male mice and RIPostC was induced immediately after MCAO. We found that RIPostC suppressed the activation of NLRP3 inflammasome to reduce the maturation of GSDMD, leading to decreased pyroptosis in microglia after AIS. Hepatocyte growth factor (HGF) was identified using the high throughput screening. Importantly, HGF siRNA, exogenous HGF, and ISG15 siRNA were used to reveal that HGF/ISG15 is a possible mechanism of RIPostC regulation in vivo and in vitro.

Effect of Programmed Comprehensive Nursing for Postoperative Delirium in Intensive Care Unit Patients.

Background: This study is to evaluate the effect of programmed comprehensive nursing for postoperative delirium in intensive care unit (ICU) patients. Methods: A total of 90 cases of ICU surgery patients admitted to our hospital from July 2019 to July 2020 were recruited and assigned via the random number table method (1 : 1) to receive either conventional nursing (control group) or programmed comprehensive nursing (experimental group). The delirium assessment method was used to record the incidence of delirium events at different time points after the intervention. Outcome measures included delirium events, the Visual Analogue Scale (VAS) scores, the Pittsburgh sleep quality index (PSQI) scores, the activities of daily living (ADL) scale scores, nursing satisfaction, and total nursing efficiency. Results: Programmed comprehensive nursing was associated with significantly better nursing satisfaction, ADL scores, VAS scores, PSQI scores, and nursing efficiency than conventional nursing (all P < 0.05). Programmed comprehensive nursing resulted in a significantly lower incidence of postoperative delirium than conventional nursing (P < 0.05). Conclusion: The application of programmed comprehensive nursing for postoperative delirium in ICU patients shows a promising efficiency, effectively alleviates the clinical symptoms of patients, and optimizes various clinical indicators of patients; hence, it is worthy of further application and promotion.

Palladium-Catalyzed Regioselective [5 + 1] Annulation of Vinyl Aziridines/Epoxides with ClCF2COONa.

Palladium-catalyzed regioselective [5 + 1] annulation reactions of vinyl aziridines/epoxides with ClCF2COONa have been developed. Significantly, vinyl aziridines/epoxides act as heteroatom-containing five-atom synthons, and commercially available and cheap ClCF2COONa acts as the source of carbonyl serving as a difluorocarbene precursor. This protocol provides an efficient and practical method for the synthesis of δ-lactams and δ-lactones in good yields.

Loss of function of CMPK2 causes mitochondria deficiency and brain calcification.

Brain calcification is a critical aging-associated pathology and can cause multifaceted neurological symptoms. Cerebral phosphate homeostasis dysregulation, blood-brain barrier defects, and immune dysregulation have been implicated as major pathological processes in familial brain calcification (FBC). Here, we analyzed two brain calcification families and identified calcification co-segregated biallelic variants in the CMPK2 gene that disrupt mitochondrial functions. Transcriptome analysis of peripheral blood mononuclear cells (PBMCs) isolated from these patients showed impaired mitochondria-associated metabolism pathways. In situ hybridization and single-cell RNA sequencing revealed robust Cmpk2 expression in neurons and vascular endothelial cells (vECs), two cell types with high energy expenditure in the brain. The neurons in Cmpk2-knockout (KO) mice have fewer mitochondrial DNA copies, down-regulated mitochondrial proteins, reduced ATP production, and elevated intracellular inorganic phosphate (Pi) level, recapitulating the mitochondrial dysfunction observed in the PBMCs isolated from the FBC patients. Morphologically, the cristae architecture of the Cmpk2-KO murine neurons was also impaired. Notably, calcification developed in a progressive manner in the homozygous Cmpk2-KO mice thalamus region as well as in the Cmpk2-knock-in mice bearing the patient mutation, thus phenocopying the calcification pathology observed in the patients. Together, our study identifies biallelic variants of CMPK2 as novel genetic factors for FBC; and demonstrates how CMPK2 deficiency alters mitochondrial structures and functions, thereby highlighting the mitochondria dysregulation as a critical pathogenic mechanism underlying brain calcification.

Melatonin Attenuates Thrombin-induced Inflammation in BV2 Cells and Then Protects HT22 Cells from Apoptosis.

Increasing evidence has revealed that the uncontrolled thrombin-induced inflammation following intracerebral hemorrhage (ICH) plays a key role in ICH. Oxidative stress and neuroinflammatory responses are interdependent and bidirectional events. Melatonin is now recognized as an antioxidant and a free radical scavenger due to its roles in various physiological and pathological processes. The aim of this study was to explore the molecular mechanisms underlying the effects of melatonin on thrombin-induced microglial inflammation and its indirect protection of HT22 cells from p53-associated apoptosis. Melatonin treatment attenuated the expression of IL-1ß, IL-18, cleaved caspase-1, and NLRP3 and decreased the production of reactive oxygen species (ROS), revealing its inhibitory effects against ROS-NLRP3 inflammasome activation. In further experiments investigating the protection conferred by melatonin, incubating HT22 cells with conditioned medium (CM) from thrombin-stimulated microglia induced HT22 cell apoptosis, and this effect was reversed after treating CM with either melatonin or N-acetyl-L-cysteine (NAC). Additionally, the Bax/Bcl-2 ratio and the levels of cleaved caspase-3 and p53 were markedly lower in the cells cultured in thrombin + melatonin-CM than in the cells cultured in thrombin-CM. Furthermore, the levels of MMP, ROS, SOD, MDA, and GSH-PX in bystander HT22 cells suggested that melatonin decreased HT22 cell apoptosis instigated via the p53-associated apoptotic pathway. Therefore, these findings strongly indicate the anti-inflammatory properties of melatonin that may suppress ROS-NLRP3 inflammasome activation and protect HT22 cells against apoptosis by inhibiting the ROS-mediated p53-dependent mitochondrial apoptotic pathway.

Activated mGluR5 protects BV2 cells against OGD/R induced cytotoxicity by modulating BDNF-TrkB pathway.

Activated Metabotropic glutamate receptors 5(mGluR5) exhibits protective effects against ischemic brain damage, but the underlying mechanisms are not clearly known. Brain-derived neurotrophic factor (BDNF), as a valuable member of neurotrophic factor family, exerts its protection by combining with its high-affinity receptor tyrosine protein kinase B (TrkB). To investigate the role of activated mGluR5 against oxygen-glucose deprivation (OGD)/reoxygenation (R)-mediated cytotoxicity, the cell viability, apoptosis, the release of inflammatory cytokines and accumulation of reactive oxygen species (ROS) were evaluated in BV2 cells (Microglia cell line) with or without OGD/R exposure. Our data show that CHPG (the selective mGluR5 agonist) pretreatment, as an mGluR5 agonist, protected BV2 cells against OGD/R-induced cytotoxicity, apoptosis, the release of inflammatory cytokines, and the accumulation of ROS. However, these effects were significantly reversed by the mGluR5 antagonist MPEP pretreatment. Our data also show that the expressions of BDNF and TrkB were significantly decreased in BV2 cells with OGD/R exposure. CHPG pretreatment significantly enhanced the expressions of BDNF and TrkB in BV2 cells with OGD/R exposure. However, the increased expressions were significantly abrogated by MPEP pretreatment. In addition, inhibition of BDNF/TrKB pathway by K252a also attenuated the protective effects of activated mGluR5 against OGD/R-induced cytotoxicity, apoptosis and the release of inflammatory cytokines. Morever, pretreatment with exogenous BDNF protected BV2 cells against OGD/R induced apoptosis and release of inflammatory cytokines. These data suggested that BDNF/TrKB pathway may be involved in regulating activated mGluR5' protective effects against OGD/R induced cytotoxicity in BV2 cells.