Gastric Enteric Glial Cells: A New Contributor to the Synucleinopathies in the MPTP-Induced Parkinsonism Mouse.

Accumulating evidence has shown that Parkinson's disease (PD) is a systemic disease other than a mere central nervous system (CNS) disorder. One of the most important peripheral symptoms is gastrointestinal dysfunction. The enteric nervous system (ENS) is regarded as an essential gateway to the environment. The discovery of the prion-like behavior of α-synuclein makes it possible for the neurodegenerative process to start in the ENS and spread via the gut-brain axis to the CNS. We first confirmed that synucleinopathies existed in the stomachs of chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)/probenecid (MPTP/p)-induced PD mice, as indicated by the significant increase in abnormal aggregated and nitrated α-synuclein in the TH-positive neurons and enteric glial cells (EGCs) of the gastric myenteric plexus. Next, we attempted to clarify the mechanisms in single MPTP-injected mice. The stomach naturally possesses high monoamine oxidase-B (MAO-B) activity and low superoxide dismutase (SOD) activity, making the stomach susceptible to MPTP-induced oxidative stress, as indicated by the significant increase in reactive oxygen species (ROS) in the stomach and elevated 4-hydroxynonenal (4-HNE) in the EGCs after MPTP exposure for 3 h. Additionally, stomach synucleinopathies appear before those of the nigrostriatal system, as determined by Western blotting 12 h after MPTP injection. Notably, nitrated α-synuclein was considerably increased in the EGCs after 3 h and 12 h of MPTP exposure. Taken together, our work demonstrated that the EGCs could be new contributors to synucleinopathies in the stomach. The early-initiated synucleinopathies might further influence neighboring neurons in the myenteric plexus and the CNS. Our results offer a new experimental clue for interpreting the etiology of PD.

E46K Mutant α-Synuclein Is Degraded by Both Proteasome and Macroautophagy Pathway.

Genetic studies have revealed that rare mutations and multiplications of the gene locus in α-synuclein (α-syn) are implicated in the pathogenesis of Parkinson's disease (PD). However, the pathological effects of α-syn are still obscure. The neurotoxicity of α-syn is mainly determined by its protein levels, which depend on a balance between synthesis and degradation. Therefore, verifying the possible routes contributing to the clearance of α-syn is important for PD therapy. In this study, we established stable lines overexpressing human wild-type (WT) and E46K mutant α-syn in rat PC12 cells and investigated the degradation pathways of α-syn by using a panel of inhibitors and inducers of lysosome and proteasome function. We also monitored the degradation kinetics of α-syn by using cycloheximide to block protein synthesis. Our data showed that both proteasome and chaperon-mediated autophagy (CMA) are responsible for the degradation of the WT α-syn. Meanwhile, E46K mutant α-syn is mainly degraded by the proteasome and macroautophagy pathway. Compared with the WT protein, E46K mutant α-syn turned over more slowly in PC12 cells. In addition, overexpression of E46K mutant α-syn increased vulnerability of PC12 cells to apoptosis insults when compared with WT α-syn. Our findings may verify the possible routes contributing to the degradation of the E46K mutant α-syn.

A Review of the Extraction and Determination Methods of Thirteen Essential Vitamins to the Human Body: An Update from 2010.

Vitamins are a class of essential nutrients in the body; thus, they play important roles in human health. The chemicals are involved in many physiological functions and both their lack and excess can put health at risk. Therefore, the establishment of methods for monitoring vitamin concentrations in different matrices is necessary. In this review, an updated overview of the main pretreatments and determination methods that have been used since 2010 is given. Ultrasonic assisted extraction, liquid⁻liquid extraction, solid phase extraction and dispersive liquid⁻liquid microextraction are the most common pretreatment methods, while the determination methods involve chromatography methods, electrophoretic methods, microbiological assays, immunoassays, biosensors and several other methods. Different pretreatments and determination methods are discussed.

20C, a bibenzyl compound isolated from Gastrodia elata, protects PC12 cells against rotenone-induced apoptosis via activation of the Nrf2/ARE/HO-1 signaling pathway.

AIM: Our preliminary study shows that a bibenzyl compound isolated from Gastrodia elata, 2-[4-hydroxy-3-(4-hydroxybenzyl)benzyl]-4-(4-hydroxybenzyl)phenol (designated 20C), protects PC12 cells against H2O2-induced injury. In this study we investigated whether 20C exerted neuroprotective action in a cell model of Parkinson's disease. METHODS: A cell model of Parkinson's disease was established in PC12 cells by exposure to rotenone (4 µmol/L) for 48 h. Cell viability and apoptosis were assessed, and intracellular ROS level and the mitochondrial membrane potential (MMP) were detected. The expression of apoptosis-related proteins Bax, Bcl-2, cytochrome c, cleaved caspase-3, and oxidative stress-related proteins Nrf2, HO-1 and NQO1 were examined using Western blotting. The mRNA levels of HO-1 and NQO1 were determined with RT-PCR. The nuclear translocation of Nrf2 was observed with immunofluorescence staining. RESULTS: Treatment with rotenone significantly increased the number of apoptotic cells, accompanied by marked increases in the Bax/Bcl-2 ratio, cytochrome c release and caspase-3 activation. Rotenone also increased ROS accumulation, reduced MMP, and increased the nuclear translocation of Nrf2 as well as the mRNA and protein levels of the Nrf2 downstream target genes HO-1 and NQO1 in PC12 cells. Co-treatment with 20C (0.01-1 µmol/L) dose-dependently attenuated rotenone-induced apoptosis and oxidative stress in PC12 cells. Nrf2 knockdown by siRNA partially reversed the protective effects of 20C in rotenone-treated PC12 cells. CONCLUSION: The bibenzyl compound 20C protects PC12 cells from rotenone-induced apoptosis, at least in part, via activation of the Nrf2/ARE/HO-1 signaling pathway.

Protopanaxtriol protects against 3-nitropropionic acid-induced oxidative stress in a rat model of Huntington's disease.

AIM: Protopanaxtriol (Ppt) is extracted from Panax ginseng Mayer. In the present study, we investigated whether Ppt could protect against 3-nitropropionic acid (3-NP)-induced oxidative stress in a rat model of Huntington's disease (HD) and explored the mechanisms of action. METHODS: Male SD rats were treated with 3-NP (20 mg/kg on d 1, and 15 mg/kg on d 2-5, ip). The rats received Ppt (5, 10, and 20 mg/kg, po) daily prior to 3-NP administration. Nimodipine (12 mg/kg, po) or N-acetyl cysteine (NAC, 100 mg/kg, po) was used as positive control drugs. The body weight and behavior were monitored within 5 d. Then the animals were sacrificed, neuronal damage in striatum was estimated using Nissl staining. Hsp70 expression was detected with immunohistochemistry. Reactive oxygen species (ROS) generation was measured using dihydroethidium (DHE) staining. The levels of components in the Nrf2 pathway were measured with immunohistochemistry and Western blotting. RESULTS: 3-NP resulted in a marked reduction in the body weight and locomotion activity accompanied by progressive striatal dysfunction. In striatum, 3-NP caused ROS generation mainly in neurons rather than in astrocytes and induced Hsp70 expression. Administration of Ppt significantly alleviated 3-NP-induced changes of body weight and behavior, decreased ROS production and restored antioxidant enzymes activities in striatum. Moreover, Ppt directly scavenged free radicals, increased Nrf2 entering nucleus, and the expression of its downstream products heme oxygenase-1 (HO-1) and NAD(P)H quinone oxidase 1 (NQO1) in striatum. Similar effects were obtained with the positive control drugs nimodipine or NAC. CONCLUSION: Ppt exerts a protective action against 3-NP-induced oxidative stress in the rat model of HD, which is associated with its anti-oxidant activity.

The applied principles of EEG analysis methods in neuroscience and clinical neurology.

Electroencephalography (EEG) is a non-invasive measurement method for brain activity. Due to its safety, high resolution, and hypersensitivity to dynamic changes in brain neural signals, EEG has aroused much interest in scientific research and medical fields. This article reviews the types of EEG signals, multiple EEG signal analysis methods, and the application of relevant methods in the neuroscience field and for diagnosing neurological diseases. First, three types of EEG signals, including time-invariant EEG, accurate event-related EEG, and random event-related EEG, are introduced. Second, five main directions for the methods of EEG analysis, including power spectrum analysis, time-frequency analysis, connectivity analysis, source localization methods, and machine learning methods, are described in the main section, along with different sub-methods and effect evaluations for solving the same problem. Finally, the application scenarios of different EEG analysis methods are emphasized, and the advantages and disadvantages of similar methods are distinguished. This article is expected to assist researchers in selecting suitable EEG analysis methods based on their research objectives, provide references for subsequent research, and summarize current issues and prospects for the future.

External Ba2+ block of the two-pore domain potassium channel TREK-1 defines conformational transition in its selectivity filter.

TREK-1 is a member of the two-pore domain potassium channel family that is known as a leak channel and plays a key role in many physiological and pathological processes. The conformational transition of the selectivity filter is considered as an effective strategy for potassium channels to control the course of potassium efflux. It is well known that TREK-1 is regulated by a large volume of extracellular and intracellular signals. However, until now, little was known about the selectivity filter gating mechanism of the channel. In this research, it was found that Ba(2+) blocked the TREK-1 channel in a concentration- and time-dependent manner. A mutagenesis analysis showed that overlapped binding of Ba(2+) at the assumed K(+) binding site 4 (S4) within the selectivity filter was responsible for the inhibitory effects on TREK-1. Then, Ba(2+) was used as a probe to explore the conformational transition in the selectivity filter of the channel. It was confirmed that collapsed conformations were induced by extracellular K(+)-free and acidification at the selectivity filters, leading to nonconductive to permeable ions. Further detailed characterization demonstrated that the two conformations presented different properties. Additionally, the N-terminal truncated isoform (ΔN41), a product derived from alternative translation initiation, was identified as a constitutively nonconductive variant. Together, these results illustrate the important role of selectivity filter gating in the regulation of TREK-1 by the extracellular K(+) and proton.

Inferences of carboplatin response-related signature by integrating multiomics data in ovarian serous cystadenocarcinoma.

Platinum-based chemotherapy, especially carboplatin, is the primary measure to treat patients with ovarian cancer (OC). However, OC patients still have an adverse prognosis due to emergency of chemotherapy resistance. Ovarian serous cystadenocarcinoma (OSC) is the most common histological subtype of OC. Therefore, identifying the key factors that affect chemotherapy resistance and searching novel treatments had become a top priority. In this study, we analyzed carboplatin response-related mRNA, miRNA, DNA methylation, and alternative splicing (AS) and established a drug-resistant signature for carboplatin in OSC. This drug-resistant signature was obviously higher in resistant group than in non-resistant group and had accuracy predictive performance, which demonstrated that this signature could be considered as a superior indicator for OSC patients with carboplatin resistance. Furthermore, we selected three potential small molecule drugs including liranaftate, siguazodan, and tramiprostate to inhibit carboplatin resistance of OSC. In addition, we also identified ZINC00000205417, ZINC00000140928, and ZINC00021908260 were potential small molecule compounds for SLC17A7 based on Molecular Operating Environment (MOE) virtual screening. Finally, we confirmed the drug-like properties of these small molecule drugs via evaluating absorption, distribution, metabolism, elimination, and toxicity (ADMET) property. In summary, the signature could be used as biomarker for carboplatin resistance and small molecule drugs targeting these genes could improve clinical treatment for OSC in the future.

Progress in Pretreatment and Analysis of Cephalosporins: An Update Since 2005.

Cephalosporins, one of the classic ß-lactam antibiotics, has been widely concerned by global population as the most commonly used broad-spectrum antibiotic. Serious health hazards are possible to be posed by quality control problems caused by its unstable structure, as well as food and environmental pollution introduced by improper use. Given the reasons above, the sensitive and valid methods for monitor and determination of cephalosporins in different matrices are urgently required. This review aims to provide a comprehensive overview on the current status of the pretreatment and analysis methods of cephalosporins in bulk drug, pharmaceutics, animal-derived foodstuffs including eggs, milk, meat, environment samples, and biological samples. Pretreatment methods including simple steps (protein precipitation, centrifugation, filtration), liquid-liquid extraction, solid phase extraction, QuECHERS, and detection methods covering LC, LC-MS/MS, voltammetric sensor, capillary electrophoresis spectroscopy, biological methods from January 2005 to October 2018 are updated, elaborated and compared here. Moreover, advanced materials and prospects for development are discussed. HighlightsDetermination of cephalosporins in different newly found matrix are represented.Comparisons between different mass analyzers and progress in HRMS methods are in detailed.Optimization of experimental conditions are discussed.Newly emerged eco-friendly methods are introduced.

Development of alternative splicing signature in lung squamous cell carcinoma.

Increasing evidence demonstrated that alternative splicing (AS) plays a vital role in tumorigenesis and clinical outcome of patient. However, systematical analysis of AS in lung squamous cell carcinoma (LUSC) is lacking and greatly necessary. Thus, this study was to systematically estimate the function of AS events served as prognostic indicators in LUSC. Among 31,345 mRNA AS events in 9633 genes, we detected 1996 AS in 1409 genes which have significant connection with overall survival (OS) of LUSC patients. Then, prognostic model based on seven types of AS events was established and we further constructed a combined prognostic model. The Kaplan-Meier curve results suggested that seven types of AS signatures and the combined prognostic model could exhibit robust performance in predicting prognosis. Patients in the high-risk group had significantly shorter OS than those in the low-risk group. The ROC showed all prognostic models had high accuracy and powerful predictive performance with different AUC ranging from 0.837 to 0.978. Moreover, the combined prognostic model had highest performance in risk stratification and predictive accuracy than single prognostic models and had higher accuracy than other mRNA model. Finally, a significant correlation network between survival-related AS genes and prognostic splicing factors (SFs) was established. In conclusion, our study provided several potential prognostic AS models and constructed splicing network between AS and SFs in LUSC, which could be used as potential indicators and treatment targets for LUSC patients.

Prevalence of Oral Mucosal Diseases in Older Adults in Mainland China: A Meta-Analysis of Observational Studies.

Oral mucosal disease (OMD) is a public health challenge globally, but the epidemiological findings in older adults have been inconsistent in China. Thus, this meta-analysis was carried out to explore the prevalence of OMD and its moderating factors in this population. An electronic literature search was conducted of both international (PubMed, PsycINFO, and EMBASE) and Chinese (China National Knowledge Infrastructure and WanFang) databases from inception to November 1, 2019. The Der-Simonian and Laird random effects model was used to synthesize the prevalence of OMD and its 95% confidence intervals (95% CI). Twenty-four studies covering 23,653 older adults were included. The pooled prevalence of OMD was 23% (95% confidence interval: 17.9%-29.0%) Subgroup analyses and meta-analysis revealed that the prevalence of OMD was significantly associated with the reporting sampling, year of publication, and survey (all p values <0.05). This meta-analysis found that the prevalence of OMD among older adults in mainland China was significantly high. Early detection and effective intervention of OMD in older adults have public health and clinical importance.

Automated Detector of High Frequency Oscillations in Epilepsy Based on Maximum Distributed Peak Points.

High frequency oscillations (HFOs) are considered as biomarker for epileptogenicity. Reliable automation of HFOs detection is necessary for rapid and objective analysis, and is determined by accurate computation of the baseline. Although most existing automated detectors measure baseline accurately in channels with rare HFOs, they lose accuracy in channels with frequent HFOs. Here, we proposed a novel algorithm using the maximum distributed peak points method to improve baseline determination accuracy in channels with wide HFOs activity ranges and calculate a dynamic baseline. Interictal ripples (80-200[Formula: see text]Hz), fast ripples (FRs, 200-500[Formula: see text]Hz) and baselines in intracerebral EEGs from seven patients with intractable epilepsy were identified by experienced reviewers and by our computer-automated program, and the results were compared. We also compared the performance of our detector to four well-known detectors integrated in RIPPLELAB. The sensitivity and specificity of our detector were, respectively, 71% and 75% for ripples and 66% and 84% for FRs. Spearman's rank correlation coefficient comparing automated and manual detection was [Formula: see text] for ripples and [Formula: see text] for FRs ([Formula: see text]). In comparison to other detectors, our detector had a relatively higher sensitivity and specificity. In conclusion, our automated detector is able to accurately calculate a dynamic iEEG baseline in different HFO activity channels using the maximum distributed peak points method, resulting in higher sensitivity and specificity than other available HFO detectors.

Overexpression of human E46K mutant α-synuclein impairs macroautophagy via inactivation of JNK1-Bcl-2 pathway.

Parkinson's disease (PD) is pathologically characterized by selective loss of dopaminergic neurons in the midbrain and the existence of intracellular protein inclusions termed Lewy bodies, largely composed of α-synuclein. Genetic studies have revealed that rare point mutations in the gene encoding α-synuclein including A30P, A53T, and E46K are associated with familial forms of PD, indicating a pathological role for mutant α-synuclein in PD etiology. However, the mechanisms underlying the neuronal toxicity of mutant α-synuclein are still to be elucidated. Growing evidence has suggested a deleterious effect of mutant α-synuclein on the autophagy-lysosome pathway. In this study, we discovered that overexpression of human E46K mutant α-synuclein impaired macroautophagy in mammalian cells. Our data showed that overexpression of E46K mutant α-synuclein impaired autophagy at an early stage of autophagosome formation via the c-Jun N-terminal kinase 1 (JNK1)-Bcl-2 but not the mammalian target of rapamycin (mTOR) pathway. Overexpressed E46K mutant α-synuclein inhibited JNK1 activation, leading to a reduced Bcl-2 phosphorylation and increased association between Bcl-2 and Beclin1, further disrupting the formation of Beclin1/hVps34 complex, which is essential for autophagy initiation. Furthermore, overexpression of E46K mutant α-synuclein increased the vulnerability of differentiated PC12 cells to rotenone treatment, which would be partly due to its inhibitory effects on autophagy. Our findings may shed light on the potential roles of mutant α-synuclein in the pathogenesis of PD.

[Heterotopic complex membrane induces bone formation].

OBJECTIVE: The composite membrane was made by recombinant human bone morphogenetic protein-2 (rhBMP-2), collagen (Co), polylactic-co-glycolic acid (PLGA). To observe the ectopic bone formation and evaluate the capability of rhBMP-2/Co/PLGA complex membrane. METHODS: 48 male Kunming adult mice were divided into two groups randomly: Co/PLGA complex membrane group and rhBMP-2/Co/PLGA complex membrane group. After etherized, an incision was made on the outer muscle and two groups of complex membranes were implanted into the right side hind limb muscle pouch. 8 mice from each group were sacrificed at 7, 14, 28 d. The hind limbs were removed, examined by soft X-ray, HE staining and light microscope examination. RESULTS: Wounds healed well, no infections and rejections were observed. In the Co/PLGA group, partial implanted membrane degradated and absorbed at 7 d. The membrane collagen fiber appeared loose at 14 d. The membrane lost its intact to disrupted at 28 d. No ectopic bone formation was found. In the rhBMP-2/Co/PLGA group, ectopic bone formation was found by eye, soft X-ray and histological examinations. The rhBMP-2/Co/PLGA complex membrane degraded slower than Co/PLGA complex membrane. CONCLUSION: Co/PLGA can provide a carrier for rhBMP-2 to have stronger effect of biological activity.

Involvement of tissue transglutaminase in endothelin 1-induced hypertrophy in cultured neonatal rat cardiomyocytes.

A potential link between tissue-type transglutaminase (tTG) and cardiac hypertrophy was suggested recently. However, whether tTG is implicated in hypertrophic agonist-induced cardiac hypertrophy is not yet known. The purpose of this study was to investigate the effects of tTG on cardiomyocyte hypertrophy induced by endothelin (ET) 1. Real-time quantitative RT-PCR and Western blot analysis demonstrated that ET-1 increased the expression of tTG mRNA and protein in cardiomyocytes by activating ET(A) receptors. ET-1 failed to cause increases in cell size and [(3)H]leucine uptake, sarcomere reorganization, and gene induction of the atrial natriuretic factor when cardiomyocytes were treated with monodansylcadaverine, a competitive inhibitor of tTG. Furthermore, the effects of ET-1 on multifunctional activities of tTG were determined by evaluating the incorporation of [(3)H]putrescine into N,N'-dimethylated casein and charcoal absorption, respectively. The results showed that ET-1 did not influence the basal transglutaminase activity of cardiomyocytes but significantly inhibited the 0.1-mmol/L Ca(2+)-stimulated transglutaminase activity. Otherwise, ET-1 elevated the activity of GTPase in a concentration- and time-dependent manner. In vivo, right ventricular hypertrophy induced by 2 weeks of chronic hypoxia was depressed by the tTG inhibitor cystamine (10 to 30 mg/kg, 2 times per day, IP) in a dose-dependent manner. Taken together, our data strongly supported the notion that tTG may act as a positive regulator of the hypertrophic program in response to ET-1. This is probably attributable to the signaling activity of tTG rather than transglutaminase activity.