Membrane-Based In-Gel Loop-Mediated Isothermal Amplification (mgLAMP) System for SARS-CoV-2 Quantification in Environmental Waters.

Since the COVID-19 pandemic is expected to become endemic, quantification of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in ambient waters is critical for environmental surveillance and for early detection of outbreaks. Herein, we report the development of a membrane-based in-gel loop-mediated isothermal amplification (mgLAMP) system that is designed for the rapid point-of-use quantification of SARS-CoV-2 particles in environmental waters. The mgLAMP system integrates the viral concentration, in-assay viral lysis, and on-membrane hydrogel-based RT-LAMP quantification using enhanced fluorescence detection with a target-specific probe. With a sample-to-result time of less than 1 h, mgLAMP successfully detected SARS-CoV-2 below 0.96 copies/mL in Milli-Q water. In surface water, the lowest detected SARS-CoV-2 concentration was 93 copies/mL for mgLAMP, while the reverse transcription quantitative polymerase chain reaction (RT-qPCR) with optimal pretreatment was inhibited at 930 copies/mL. A 3D-printed portable device is designed to integrate heated incubation and fluorescence illumination for the simultaneous analysis of nine mgLAMP assays. Smartphone-based imaging and machine learning-based image processing are used for the interpretation of results. In this report, we demonstrate that mgLAMP is a promising method for large-scale environmental surveillance of SARS-CoV-2 without the need for specialized equipment, highly trained personnel, and labor-intensive procedures.

Effectiveness and safety of perampanel as adjunctive therapy among Chinese patients with focal-onset epilepsy: A real-world prospective observational study.

OBJECTIVE: Perampanel (PER) has previously been shown to be effective and tolerable when used as an adjunctive therapy for patients with focal-onset seizures (FOS). This study aimed to evaluate the effect of PER as adjunctive therapy for patients with FOS in the Chinese population under real-world conditions for 1 year. METHODS: A prospective, single-center, 1-year observational study was conducted at Huashan Hospital, enrolling both under age (≥4 years old) and adult patients with FOS. Response to PER was assessed at 3-, 6-, and 12-month checkpoints by analyzing the 50 % responder rate, the seizure-free rate, and reduction in seizure frequency. RESULTS: One hundred and eight patients (mean age: 26.6 years, 56.5 % males) with FOS were included, with seventy-six patients finishing the 1-year follow-up (retention rate: 70.4 %, mean PER dose: 4.3 mg/day). The seizure frequency was reduced significantly at 3, 6, and 12 months relative to baseline (p < 0.001 for each seizure type). At 12 months, the responder rate was 65.8 %, and the seizure-free rate was 39.5 %. A significantly higher responder rate was found in patients with focal to bilateral tonic-clonic seizures (p = 0.024), among which the percentage of patients with sleep-related epilepsy was significantly high (p = 0.045). Responders had a lower number of concomitant anti-seizure medications (ASMs) than the non-responders (p = 0.009). Drug-related adverse events (AEs) were reported in 37 % of patients, mostly mild or moderate, and the patients who experienced AEs had a higher daily dose of PER than those who did not (p = 0.026). CONCLUSION: Perampanel, an add-on therapy for focal-onset seizures, was found to be effective and tolerable in Chinese patients at 12 months.

Rapid Detection Methods for Bacterial Pathogens in Ambient Waters at the Point of Sample Collection: A Brief Review.

The world is currently facing a serious health burden of waterborne diseases, including diarrhea, gastrointestinal diseases, and systemic illnesses. The control of these infectious diseases ultimately depends on the access to safe drinking water, properly managed sanitation, and hygiene practices. Therefore, ultrasensitive, rapid, and specific monitoring platforms for bacterial pathogens in ambient waters at the point of sample collection are urgently needed. We conducted a literature review on state-of-the-art research of rapid in-field aquatic bacteria detection methods, including cell-based methods, nucleic acid amplification detection methods, and biosensors. The detection performance, the advantages, and the disadvantages of the technologies are critically discussed. We envision that promising monitoring approaches should be automated, real-time, and target-multiplexed, thus allowing comprehensive evaluation of exposure risks attributable to waterborne pathogens and even emerging microbial contaminants such as antibiotic resistance genes, which leads to better protection of public health.

Synthesis and application of superabsorbent polymer microspheres for rapid concentration and quantification of microbial pathogens in ambient water.

Even though numerous methods have been developed for the detection and quantification of waterborne pathogens, the application of these methods is often hindered by the very low pathogen concentrations in natural waters. Therefore, rapid and efficient sample concentration methods are urgently needed. Here we present a novel method to pre-concentrate microbial pathogens in water using a portable 3D-printed system with super-absorbent polymer (SAP) microspheres, which can effectively reduce the actual volume of water in a collected sample. The SAP microspheres absorb water while excluding bacteria and viruses by size exclusion and charge repulsion. To improve the water absorption capacity of SAP in varying ionic strength waters (0-100 mM), we optimized the formulation of SAP to 180 g⋅L-1 Acrylamide, 75 g⋅L-1 Itaconic Acid and 4.0 g⋅L-1 Bis-Acrylamide for the highest ionic strength water as a function of the extent of cross-linking and the concentration of counter ions. Fluorescence microscopy and double-layer agar plating respectively showed that the 3D-printed system with optimally-designed SAP microspheres could rapidly achieve a 10-fold increase in the concentration of Escherichia coli (E. coli) and bacteriophage MS2 within 20 min with concentration efficiencies of 87% and 96%, respectively. Fold changes between concentrated and original samples from qPCR and RT-qPCR results were found to be respectively 11.34-22.27 for E. coli with original concentrations from 104 to 106 cell·mL-1, and 8.20-13.81 for MS2 with original concentrations from 104 to 106 PFU·mL-1. Furthermore, SAP microspheres can be reused for 20 times without performance loss, significantly decreasing the cost of our concentration system.

Effects of UGT1A9 genetic polymorphisms on monohydroxylated derivative of oxcarbazepine concentrations and oxcarbazepine monotherapeutic efficacy in Chinese patients with epilepsy.

AIM: The human UDP-glucuronosyltransferase which is genetically polymorphic catalyzes glucuronidations of various drugs. The interactions among UGT1A4, UGT1A6, UGT1A9, and UGT2B15 genetic polymorphisms, monohydroxylated derivative (MHD) of oxcarbazepine (OXC) plasma concentrations, and OXC monotherapeutic efficacy were explored in 124 Chinese patients with epilepsy receiving OXC monotherapy. METHOD: MHD is the major active metabolite of OXC, and its plasma concentration was measured using high-performance liquid chromatography when patients reached their maintenance dose of OXC. Genomic DNA was extracted from whole blood and SNP genotyping performed using PCR followed by dideoxy chain termination sequencing. We followed the patients for at least 1 year to evaluate the OXC monotherapy efficacy. Patients were divided into two groups according to their therapeutic outcome: group 1, seizure free; group 2, not seizure free. The data were analyzed using T test, one-way analysis of variance (ANOVA), Kruskal-Wallis test, chi-square test, Fisher's exact test, correlation analysis, and multivariate regression analysis. RESULT: T test analysis showed that MHD plasma concentrations were significantly different between the two groups (p = 0.002). One-way ANOVA followed by Bonferroni post hoc testing of four candidate SNPs revealed that carriers of the UGT1A9 variant allele I399 C > T (TT 13.28 ± 7.44 mg/L, TC 16.41 ± 6.53 mg/L) had significantly lower MHD plasma concentrations and poorer seizure control than noncarriers (CC 22.24 ± 8.49 mg/L, p < 0.05). CONCLUSION: In our study, we have demonstrated the effects of UGT1A9 genetic polymorphisms on MHD plasma concentrations and OXC therapeutic efficacy. Through MHD monitoring, we can predict OXC therapeutic efficacy, which may be useful for the personalization of OXC therapy in epileptic patients.

MicroRNA-181c negatively regulates the inflammatory response in oxygen-glucose-deprived microglia by targeting Toll-like receptor 4.

Cerebral hypoxia/ischemia rapidly induces inflammation in the brain, which is characterized by microglial activation and the release of inflammatory cytokines. We have previously demonstrated that miR-181c can directly regulate tumor necrosis factor (TNF)-α production post-transcriptionally. Here, we determined that hypoxia up-regulated TLR4 expression but down-regulated miR-181c expression in primary microglia. We also demonstrated that miR-181c suppresses TLR4 by directly binding its 3'-untranslated region. In addition, miR-181c inhibited NF-κB activation and the downstream production of proinflammatory mediators, such as TNF-α, IL-1ß, and iNOS. Knocking down TLR4 in microglia significantly decreased TLR4 expression and inhibited NF-κB activation and the downstream production of proinflammatory mediators, whereas ectopic TLR4 expression significantly abrogated the suppressed inflammatory response induced by miR-181c. Therefore, our study identified an important role for the miR-181c-TLR4 pathway in hypoxic microglial activation and neuroinflammation. This pathway could represent a potential therapeutic target for cerebral hypoxic diseases associated with microglial activation and the inflammatory response. Cerebral hypoxia/ischemia induces microglial activation and the release of inflammatory cytokines. We found that hypoxia down-regulated miR-181c in primary microglia. In addition, miR-181c inhibited TLR4 expression through binding to its 3'UTR, thus inhibiting NF-kB activation and the production of downstream proinflammatory mediators. Therefore, the miR-181c-TLR4 pathway may be a potential therapeutic target for the treatment of cerebral hypoxic diseases.

Long-term treatment with Perampanel of Chinese patients with focal-onset seizures, especially in sleep-related epilepsy: a prospective real-world observational study.

Background: There is currently a lack of studies examining the long-term therapeutic effectiveness of the third-generation anti-sezure medication, perampanel (PER), for focal-onset seizures (FOS), particularly in Chinese patients with sleep-related epilepsy (SRE). Additionally, the appropriate dosage, plasma concentration, and the relationship between dose and plasma concentration of PER in Chinese patients are still uncertain. Methods: A prospective, single-center, 24-month observational study was conducted in patients diagnosed with FOS, with a focus on patients with SRE. Changes in seizure frequency from baseline, adverse events, and retention rates were analyzed at 12 and 24 months following the start of the treatment. Tolerability was evaluated based on adverse events and discontinuation profiles. PER plasma concentrations were used to assess dose-concentration-response relationships. Results: A total of 175 patients were included (median age: 25 years; range: 4-72 years; 53. 1% males and 46.9% females), with the SRE population accounting for 49. 1% (n = 86). The patients diagnosed with SRE showed considerably higher response rates than those who did not have this diagnosis (p = 0.025, odds ratio = 3.8). Additionally, the SRE group adhered better to PER treatment (r = 0.0009). Patients with a shorter duration of epilepsy (median: 3 years; range:2-7 years) demonstrated a more favorable therapeutic response to PER (p = 0.032). Throughout the administration of maintenance doses, among the entire FOS population, the concentration of PER (C0) ranged between 101.5 and 917.4 ng/mL (median, 232.0 ng/mL), and the mean plasma concentration of PER in the responders was 292.8 ng/mL. We revealed a linear relationship between PER dose and plasma concentration, regardless of whether PER was used as monotherapy or add-on therapy. The retention rates were 77.7% and 65. 1% at 12 and 24 months, respectively. Drug-related adverse events occurred in 45.0% of the patients and were mostly manageable. Conclusion: PER effectively reduced seizure frequency in Chinese patients with FOS, particularly in those with SRE, over a 24-month period. The treatment was well-tolerated and had a clear linear dose-plasma concentration relationship.

The clinical features of familial focal epilepsy with variable foci and NPRL3 gene variant.

OBJECTIVE: Familial focal epilepsy with variable foci (FFEVF) is a rare type of focal epilepsy syndrome; it is associated with NPRL3 variant. However, relevant reports are rare in China. We aimed to analyze the clinical features of Chinese patients with FFEVF to understand further the differences between various NPRL3 variants and explored the effect of NPRL3 variant on mRNA. METHODS: We ran a full workup on a family with FFEVF (four patients, one healthy member): an inquiry of medical history, cranial magnetic resonance imaging (MRI), electroencephalogram (EEG), and whole exon sequencing. Their clinical features were compared with those of other FFEVF patients in published reports. The mRNA splicing changes were analyzed quantitatively and qualitatively using real-time quantitative-polymerase chain reaction (q-PCR) and reverse transcription (RT)-PCR and compared between our patients and healthy individuals. RESULTS: Patients with NPRL3: c.1137dupT variant had a wide range of onset age (4 months to 31 years), diverse seizure types, variable foci (frontal lobe/temporal lobe), different seizure times (day/night) and frequencies (monthly/seldom/every day), different therapeutic effects (refractory epilepsy/almost seizure free), normal MRI, and abnormal EEG (epileptiform discharge, slow wave). The phenotypic spectrum with different NPRL3 variants was either similar or different. Significantly different relative quantities of mRNA were found between patients and healthy individuals in real-time qPCR. Abnormal splicing was observed in patients compared with healthy individual in RT-PCR. Despite having the same gene variant, different family members had different mRNA splicing, possibly causing different phenotypes. CONCLUSION: The clinical features of FFEVF varied, and auxiliary inspection was atypical. NPRL3: c.1137dupT could change the relative quantity of mRNA and cause abnormal splicing, which might produce different phenotypes in different family members.

A high-performance liquid chromatography-tandem mass spectrometry method for quantification of perampanel in human plasma: Effect of concomitant anti-seizure medications on perampanel concentration in patients with epilepsy.

Perampanel is a first-in-class α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor antagonist and a novel anti-seizure medication. It is currently used as adjunctive treatment for partial seizures in patients over 12 years of age. With the increasing clinical application of perampanel, monitoring its concentration under certain clinical conditions is important. This study developed a rapid and sensitive high-performance liquid chromatography-tandem mass spectrometry method to quantify perampanel in human plasma. Protein precipitation with acetonitrile was performed for sample preparation. Perampanel and perampanel-d5 (internal standard) were analyzed under gradient conditions using a C18 column. The mobile phase was composed of 0.1% (v/v) formic acid in water (solvent A) and 0.1% (v/v) formic acid in acetonitrile (solvent B) at a flow rate of 0.4 mL/min. Mass detection was performed using multiple reaction monitoring in the positive ionization mode. The proposed method was validated over a range of 0.5-500 ng/mL for perampanel. The linearity (r2 value) was higher than 0.999, and the linear equation was y = 0.00116x + 0.0116. The accuracy of the low-, middle-, and high-quality control samples was between 103% and 113%, and the intra- and inter-day precisions were below 6.81%. The quality of the proposed method was evaluated in accordance with the US Food and Drug Administration Bioanalytical Method Validation Guidance for Industry. The plasma concentrations of perampanel in 25 patients were successfully determined to be 38.7-577.7 ng/mL using the validated method.

Pregnancy characteristics and adverse outcomes in offspring of women with epilepsy: a prospective registry study from Mainland China.

Objective: This study aimed to explore the influencing factors of adverse outcomes in the offspring of women with epilepsy (WWE) and to analyze the changes brought about by the epilepsy knowledge popularization campaign in China (EKPCIC). Methods: This nested case-control study focused on WWE and their offspring from a female epilepsy cohort in mainland China. From January 2009 to August 2022, WWE was prospectively enrolled in 32 study centers. This study aimed to observe the health outcomes of their offspring within 1 year of age. The main outcome measure assessed the health status of the offspring within their first year of age. We aimed to analyze the effects of seizures, anti-seizure medicines (ASMs), and a lack of folic acid supplementation on adverse outcomes in the offspring of WWE and to explore the changes in perinatal management and adverse outcomes of the offspring after dissemination of the EKPCIC in 2015. Additionally, subgroup analyses were conducted to compare seizure control during pregnancy between the valproate and non-valproate groups. Results: In total, 781 pregnancies in 695 WWE were included, of which 186 (23.69%) had adverse outcomes. The National Hospital Epilepsy Severity Scale score, number of seizures, status epilepticus, ASM type, and valproate and folic acid doses were associated with a high risk of adverse outcomes. After the EKPCIC, the use of ASMs (P = 0.013) and folic acid (P < 0.001), the seizure-free rate during pregnancy (P = 0.013), and the breastfeeding rate (P < 0.001) increased, whereas the incidence of complications during pregnancy decreased (P = 0.013). However, there was no significant difference in the incidence of adverse outcomes between the analyzed offspring pre-/post-EKPCIC. Additionally, there was no association between the frequency of seizures at different time points during pregnancy and the use of valproate (F = 1.514, P = 0.221). Conclusion: Possible factors influencing adverse outcomes in the offspring of WWE include seizures, type and number of ASM usage, and a lack of folic acid supplementation. Although the management of WWE during pregnancy is now more standardized, further efforts are needed to reduce adverse outcomes in offspring.

Comparison of Lamotrigine and Oxcarbazepine Monotherapy Among Chinese Adult Patients With Newly-Diagnosed Focal-Onset Epilepsy: A Prospective Observational Study.

Objective: We performed a prospective cohort study to compare the efficacy, safety, effect on mood, and quality of life between lamotrigine (LTG) and oxcarbazepine (OXC) monotherapy among Chinese adult patients with newly-diagnosed focal-onset epilepsy (FOE) with or without secondarily generalized tonic-clonic seizures. Methods: We enrolled 106 adult patients with new-onset FOE, of whom 56 were in the OXC group and 50 in the LTG group. Their clinical characteristics were detailly recorded especially basic seizure frequency, seizure types, and drug-related adverse events. Efficacy was evaluated as seizure-free (no seizure for 6 months), effective (seizure reduction by more than 50%), and ineffective (seizure reduction by less than 50%). Both intention-to-treat and per-protocol analyses were performed. We also assessed their mood state with the Zung Self-rating Scale for anxiety (Z-SAS) and Zung Self-rating Scale for Depression (Z-SDS) and quality of life (QOL) with Quality of Life in Epilepsy (QOLIE-31) at their baseline visit, 3-month visits, and 6-month visit. Intra-group comparisons in each group and inter-group comparisons between the two groups were made. Correlation analysis and multiple regression analysis were also conducted. Results: Except for gender, the two groups were well matched in any other characteristics such as primary seizure frequency and seizure types. In terms of efficacy, 33 patients in the OXC group were evaluated as seizure-free and 15 as effective, while in the LTG group, 31 were seizure-free, and nine were effective. No significant differences could be observed in efficacy between the two groups (P = 0.429). Through multiple logistic regression analysis, we found that OXC monotherapy was more likely to predict a seizure-free state (OR = 1.76) than LTG, but the difference didn't reach statistical significance (P = 0.322) after correcting for other clinical variables. Both groups had adverse events such as fatigue, drowsiness, dizziness, rash, and gastrointestinal discomfort, most of which were mild and transient. In the OXC group, the scores of SAS (P = 0.067) and SDS (P = 0.004) reduced at the 6-month visit, while the score of QOLIE-31 significantly increased (P = 0.001). In the LTG group, a significant decrease in SAS and SDS scores and an increase in QOLIE-31 scores could be witnessed (All P < 0.001). The inter-group comparison showed that improvement of SAS and SDS in the LTG group was more evident than that in the OXC group, which was of statistical significance. Correlational analysis indicated that the improvement of mood and life quality scales in both groups was independent of baseline seizure frequency and treatment efficacy. Multiple linear regression analysis indicated that LTG monotherapy was the only independent factor that could predict a better SAS (P = 0.01) and SDS (P = 0.019) outcome. Conclusions: OXC and LTG are effective as monotherapy and can be considered first-line selection among adult patients with new-onset FOE. Most adverse events are mild, transient, and tolerable. The two drugs improve the mood state of patients, though LTG is superior to OXC in this respect. OXC and LTG have great power in enhancing patients' quality of life. The positive effect on the psychosocial well-being of epilepsy patients may be one of the intrinsic pharmacological properties of LTG and OXC.

Development of a proof-of-concept microfluidic portable pathogen analysis system for water quality monitoring.

Waterborne diseases cause millions of deaths worldwide, especially in developing communities. The monitoring and rapid detection of microbial pathogens in water is critical for public health protection. This study reports the development of a proof-of-concept portable pathogen analysis system (PPAS) that can detect bacteria in water with the potential application in a point-of-sample collection setting. A centrifugal microfluidic platform is adopted to integrate bacterial cell lysis in water samples, nucleic acid extraction, and reagent mixing with a droplet digital loop mediated isothermal amplification assay for bacteria quantification onto a single centrifugal disc (CD). Coupled with a portable "CD Driver" capable of automating the assay steps, the CD functions as a single step bacterial detection "lab" without the need to transfer samples from vial-to-vial as in a traditional laboratory. The prototype system can detect Enterococcus faecalis, a common fecal indicator bacterium, in water samples with a single touch of a start button within 1 h and having total hands-on-time being less than 5 min. An add-on bacterial concentration cup prefilled with absorbent polymer beads was designed to integrate with the pathogen CD to improve the downstream quantification sensitivity. All reagents and amplified products are contained within the single-use disc, reducing the opportunity of cross contamination of other samples by the amplification products. This proof-of-concept PPAS lays the foundation for field testing devices in areas needing more accessible water quality monitoring tools and are at higher risk for being exposed to contaminated waters.

Determinants of quality of life in people with epilepsy and their gender differences.

Improving the patient's quality of life (QOL) is the most important goal of epilepsy management. We performed this study to determine the factors associated with QOL in people with epilepsy and to assess whether there are gender differences in these determinants. Patients were interviewed using the Quality of Life in Epilepsy Inventory-31(QOLIE-31), the Adverse Event Profile (AEP), the Self-Rating Anxiety Scale (SAS), and the Hamilton Depression Rating Scale (HAMD). Two hundred forty-seven patients (152 men, 95 women) were included in the analysis. Among all patients, regressive analyses showed that AEP score was the strongest predictor of the QOLIE-31 overall score, accounting for 10.4% of the variance. The next strongest predictors were the number of currently used antiepileptic drugs (AEDs) (3.6%), the HAMD score (2.5%), and the SAS score (1.2%). Importantly, there were gender differences in these predictors of QOL. The strongest predictors of the QOLIE-31 overall score in women were the AEP score and the number of AEDs. In contrast, the strongest predictors in men were the SAS score, the AEP score and the frequency of seizures. These results indicate that perceived adverse effects of treatments and number of AEDs exerted greater effects on QOL in women, whereas anxiety and seizure-related variables had a stronger impact on QOL among men. Accordingly, it may be necessary to individualize interventions to improve the QOL of people with epilepsy.

[Factors associated with quality of life in epileptics and the variations between men and women, younger and older people].

OBJECTIVE: To determine factors associated with quality of life (QOL) in epileptics and the variations between men and women, younger and older people. METHODS: A total of 204 patients (160 younger, 49 older; 125 men, 79 women) were interviewed by the quality of life in epilepsy-31 (QOLIE-31), side effect profile (SEP), self-rating anxiety scale (SAS) and Hamilton depression scale (HAMD). Medical and socio-demographic data were acquired from patient records. Multivariate linear regressive analyses were used to determine the set of best predictors of composite QOLIE-31 score. RESULTS: No statistical difference was revealed in QOLIE-31 overall score either between younger and older patients or between men and women. Among all patients, regressive analyses revealed that SEP (beta = - 0.395, P = 0.000) and SAS (beta = -0.152, P = 0.016) were two strong predictors of QOLIE-31 overall score. Grouped by gender, among men, epilepsy duration (beta = -0.165, P = 0.028) and seizure frequency (beta = -0.284, P = 0.001) respectively associated with QOLIE-31 overall score and "social function" score while the number of AEDs (antiepileptic drugs) substantially correlated with QOLIE-31 overall score among women (beta= -0.238, P = 0.006). Grouped by ages, seizure frequency (beta = -0.284, P = 0.000) and education level (beta = 0.203, P = 0.005) predicted QOLIE-31 "social function" score only among younger patients; among older patients, a significant association were found between the number of AEDs and QOLIE-31 overall score (beta = - 0.363, P = 0.004). CONCLUSION: Side effects of AEDs and number of AEDs exert greater effect on QOL in women and older patients. And such seizure-related variables as epileptic duration and seizure frequency influence QOL only among men and younger patients.

Anti-N-methyl-D-aspartate receptor encephalitis with accompanying ovarian teratoma in female patients from East China: Clinical features, treatment, and prognostic outcomes.

PURPOSE: The association between anti-N-methyl-D-aspartate receptor encephalitis (anti-NMDAR encephalitis) and teratoma is formally recognized. This study compared the clinical features, treatments and outcomes between female patients with or without accompanying teratomas and determined the potential influences of coexisting teratomas. METHOD: Fifty-six female patients diagnosed with anti-NMDAR encephalitis were enrolled in two major tertiary hospitals in East China from January 2013 through March 2018 and were grouped as patients with or without teratoma. The clinical features were reviewed, and follow-up studies were performed. Comparisons were made between the two groups. RESULTS: Patients with teratoma reported fewer viral prodromes (p = 0.0085) and stronger positive intensity of anti-NMDAR antibodies in cerebrospinal fluid (CSF) (p = 0.0368), while nontumor patients tended to demonstrate lymphocytic pleocytosis in CSF (p = 0.0306). Seizure types varied between individuals, with complex partial seizures more common in teratoma patients (p = 0.0105). Nontumor patients frequently required combinations of first-line and second-line immunotherapy (p = 0.0014), which may be attributed to higher mRS scores at admission (p = 0.0300). Also, they had higher mean mRS scores since the 12-month follow-up and greater probability of relapse than did patients with teratomas (p = 0.0286). CONCLUSIONS: Symptomatology and auxiliary findings indicate that ovarian teratoma may be the immunologic trigger for anti-NMDAR encephalitis patients, while viral infection is likely to play a major part in pathogenesis for those without any detectable tumor. Overall, anti-NMDAR encephalitis patients with teratomas present with milder neurological symptoms and have better long-term outcomes after tumor removal.

Multicenter double-blind, randomized, placebo-controlled trial of levetiracetam as add-on therapy in Chinese patients with refractory partial-onset seizures.

PURPOSE: To evaluate efficacy and tolerability of levetiracetam (LEV; Keppra) as add-on therapy in Chinese patients with refractory partial-onset seizures. METHODS: In this multicenter, double-blind, randomized, placebo-controlled trial, 206 patients aged 16-70 years with uncontrolled partial-onset seizures were randomized to receive LEV (n =103) or placebo (n =103); 202 patients (LEV, n =102; placebo, n = 100) comprised the intent-to-treat population. An 8-week historical baseline period confirmed eligibility according to seizure count. The 16-week treatment period consisted of a 4-week up-titration period (LEV, 1,000-3,000 mg/day in two equal divided doses) followed by a 12-week maintenance period. Efficacy assessments were based on weekly frequency of partial-onset seizures during the 16-week treatment period. RESULTS: LEV significantly decreased weekly partial-onset seizure frequency over placebo by 26.8% (p < 0.001). Median percentage reductions in weekly partial-onset seizure frequency from historical baseline were 55.9% for LEV and 13.7% for placebo (p < 0.001). The >or=50% responder rates were 55.9% for LEV, compared with 26.0% for placebo (p < 0.001). Freedom from partial-onset seizures during treatment period was achieved by 11 LEV patients (10.8%) and 2 placebo patients (2.0%) (p = 0.012). Adverse events were reported by 65 LEV-treated patients (63.1%) and 62 placebo-treated patients (60.2%); most were of mild-to-moderate intensity. The most common adverse events were somnolence (LEV, 17.5%; placebo, 17.5%), decreased platelet count (LEV, 9.7%; placebo, 9.7%), and dizziness (LEV, 7.8%; placebo, 13.6%). DISCUSSION: Add-on LEV was effective and well-tolerated in Chinese patients with refractory partial-onset seizures.

The anticonvulsant and neuroprotective effects of kir2.3 activation in PTZ-induced seizures and the kainic acid model of TLE.

PURPOSE: To elucidate the role of activating the inwardly rectifying K+ channel 2.3 (Kir2.3) in acute seizure and chronic epilepsy, we investigated the effect of a Kir2.3 agonist (tenidap) on epileptic and electrophysiological activities in mice. Neuronal excitability and damage were also evaluated. METHODS: A Pentylenetetrazole (PTZ)-induced acute seizure model and a kainic acid (KA)-induced temporal epilepsy model were used in adult mice. The mice were given tenidap 30 min before PTZ injection or were given tenidap for 7 days after entering the chronic stage of the KA model. Video monitoring and EEG recordings were performed for comparisons. Immunofluorescence of c-fos was detected in the PTZ model, and Nissl staining was performed in the KA model. RESULTS: Tenidap intervention significantly reduced the duration and severity of PTZ-induced acute seizures, which conformed with the power-spectrum analyses of the EEG and the quantification of spikes on EEG. C-fos expression representing neuronal excitability was also reduced with tenidap pretreatment. However, the latency time to seizure onset was unaltered. Seven days of tenidap treatment in the chronic KA model significantly attenuated seizure and spike frequencies compared to the same animal before administration. Nissl staining showed reduced hilar neuron loss in the tenidap-intervention group but showed no difference in the width of the granule cell layer. CONCLUSION: To our knowledge, few studies have reported the relevance of Kir2.3 to epilepsy. The present data suggested that activation of Kir2.3 exerts an anticonvulsant effect in acute seizures and the chronic stage of TLE, which makes this channel a potent therapeutic target.

Activation of Kir2.3 Channels by Tenidap Suppresses Epileptiform Burst Discharges in Cultured Hippocampal Neurons.

BACKGROUND & OBJECTIVE: Tenidap, a selective human inwardly rectifying potassium (Kir) 2.3 channel opener, has been reported to have antiepileptic effect in the pilocarpine temporal lobe epilepsy rat model in our previous study. However, the effect of tenidap on neurons and its relationship with the epileptiform bursting charges in neuron is still required to be explored. METHODS: In this study, cyclothiazide (CTZ) induced cultured hippocampal neuron epileptic model was used to study the antiepileptic effect of tenidap and the relationship between Kir2.3 channel and the neuronal epileptiform burst. RESULTS: Patch clamp recording showed that both acute (2h) and chronic (48h) CTZ pre-treatment all significantly induced robust epileptiform burst activities in cultured hippocampal neurons, and tenidap acutely application inhibited this highly synchronized abnormal activities. The effect of tenidap is likely due to increased activity of Kir2.3 channels, since tenidap significantly enhanced kir current recorded from those neurons. In addition, neurons overexpressing Kir2.3 channels, by transfection with Kir2.3 plasmid, showed a significant large increase of the Kir current, prevented CTZ treatment to induce epileptiform burst discharge. CONCLUSION: Our current study demonstrated that over activation of Kir2.3 channel in hippocampal neurons could positively interference with epileptiform burst activities, and tenidap, as a selective Kir2.3 channel opener, could be a potential candidate for seizure therapy.

Propidium monoazide pretreatment on a 3D-printed microfluidic device for efficient PCR determination of live versus dead'microbial cells.

Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simu-lations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing her-ringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.

[Effect of dizocilpine on P-glycoprotein expression in hippocampus in limbic seizure: experiment with rats].

OBJECTIVE: To observe the effect of dizocilpine (MK801), a noncompetitive antagonist of N-methyl-D-aspartic acid (NMDA) receptor, on P-glycoprotein (P-gp) expression after limbic seizure, and to explore whether NMDA receptor play a role in the regulation of P-gp expression during limbic seizure. METHODS: 120 Wistar rats were randomly divided into 2 equal sets. 50 rats in Set 1 underwent intraperitoneal injection of lithium chloride, scopolamine, and pilocarpine so as to cause status epilepticus (SE) for 90 min. Then diazepam was given to terminate the SE. The rats were killed 0, 3, 6, 14, and 72 h after the SE respectively. The hippocampus was isolated. Realtime fluorescent quantitative RT-PCR (qRT-PCR) was used to detect the expression of multidrug resistance gene 1a (mdr1a) and mdr1b. Immunohistochemistry was used to detect the P-gp. The rats were used as controls. Another 60 rats (Set 2) were randomly divided into 3 equal groups: control group, given with normal saline (NS) only, SE group, given with NS 20 min before administration of pilocarpine, and MK801 group, given with MK801 20 min before administration of pilocarpine. The 3 groups in Set 2 were further divided into 2 equal subgroups of 10 rats to be killed 6 or 24 h after SE. RESULTS: The mdr1a expression in hippocampus within 72 h after seizure was much higher at each time point: the level of mdr1a expression instantly after the seizure was terminated was [5.6 (2.9) x 10(5) mRNA copies/40 ng total RNA], significantly higher than that of the controls [2.4 (1.1) x 10(5) mRNA copies/40 ng total RNA, P < 0.05], increased to the level of [7.6 (6.3) x 10(5), P < 0.01] 3 h after, and kept at such level till 72 h after. The msr1b expression transiently increased 2.2 and 2.4 times that of the controls respectively 3 h and 6 h after the seizure was terminated [(3.3 +/- 0.4) x 10(4), and (3.6 +/- 1.0) x 10(4), both P < 0.01)]. The expression level of mdr1a 6 h after the seizure was terminated of the MK801 group was (4.3 +/- 0.8) x 10(5) and the expression level of mdr1b 6 h after the seizure was terminated of the MK801 group was (2.0 +/- 0.7) x 10(4), both significantly lower than those of the SE group (both P < 0.01). The P-gp expression level 24 h after the seizure was terminated of the MK801 group was 26.6 +/- 5.0 pieces of microvessels/400 times field, significantly lower than that of the SE group (39.0 +/- 4.1, P < 0.01). CONCLUSION: MK801 down-regulates the overexpression of P-gp after seizure, which indicates that NMDA receptor may be involved in the regulation of P-gp expression during seizure. Therefore, it is possible to prevent the overexpression of P-gp after seizure by inhibiting NMDA receptor's overactivation effectively.