Development and construction of the Multidimensional Self-Efficacy Scale for Epilepsy (MSESE) and its psychometric properties.

PURPOSE: It has become evident that patients with epilepsy require strong self-efficacy support in various domains, including work, social interaction, and academic performance, to ensure their complete social functioning. Nevertheless, previous studies have predominantly assessed the self-efficacy of individuals with epilepsy from a singular perspective of disease management. This study aimed to develop the Multidimensional Self-Efficacy Scale for Epilepsy (MSESE) to assess multiple dimensions and establish its psychometric properties. METHODS: We compiled a total of 25 questions for the initial version of the questionnaire based on a review of the literature and insights from experts, patients, and family members. The study included 180 adult patients with epilepsy who met the research criteria, with 126 of them serving as pre-test samples. All participants completed the MSESE, Brief Symptom Rating Scale-50 (BSRS-50), Rosenberg Self-Esteem Scale-Chinese version (RSES-C), and General Self-Efficacy Scale (GSES). RESULTS: The final scale consisted of 12 items across four dimensions, with item factor loadings ranging from .51 to .90. Most of the fit indices indicated a good fit. Construct validity was established through significant correlations with the BSRS-50, RSES-C, and GSES (r = -0.51 to 0.69, p < 0.01). Internal consistency coefficients for the MSESE were strong at .90, with individual dimensions ranging from 0.71 to 0.89. The MSESE also demonstrated a satisfactory test-retest reliability of 0.72. CONCLUSIONS: The MSESE is a convenient, multidimensional, and easy-to-use scale with good psychometric properties, making it suitable for both clinical assessments and research purposes.

Ni-Catalyzed Asymmetric C-P Cross-Coupling Reaction for the Synthesis of Chiral Heterocyclic Phosphine Oxides.

Nickel performs excellently in C-C and C-X cross-coupling reactions. Here, we disclose a Ni(II)-catalyzed asymmetric C-P cross-coupling reaction to afford valuable chiral heterocyclic tertiary phosphine oxides. The method is mild and efficient, which invokes a self-sustained nickel catalytic cycle without an external reductant, light irradiation, or electricity.

Betel Quid Dependence Effects on Working Memory and Remote Memory in Chewers with Concurrent Use of Cigarette and Alcohol.

OBJECTIVE: The current study asked whether BQ dependence level could affect working memory (WM) and remote memory for the chewers with concurrent use of cigarettes and alcohol, a common phenomenon in Taiwan. METHODS: The standardized neuropsychological tests (Wechsler Memory Scale III (WMS-III) and Remote Memory Test) were adopted to address the BQ chewers' verbal WM, spatial WM and remote memory. The Spatial Span Test and the Digit Span Test from WMS-III and the Remote Memory Test were adopted. The Betel Nut Dependency Scale (BNDS), the Fagerstrom Test for Nicotine Dependence (FTND), and the Alcohol Use Disorders Identification Test (AUDIT) were adopted to measure the dependence levels. RESULTS: The BQ dependence level and Last BQ did not affect spatial WM, verbal WM, and remote memory. Last Cigarette is critical in affecting WM; namely, longer interval led to worse performance. Finally, higher alcohol dependence level could lead to better remote memory. CONCLUSIONS: To our knowledge, there are no BQ studies addressing the effects of concurrent use of cigarettes and alcohol on memory. The current results suggest that cigarette smoking and alcohol drinking, rather than BQ chewing, are critical for memory performance.

The effect of facial expression intensity on emotion recognition and psychosocial performance in patients with frontal or temporal lobe epilepsy.

PURPOSE: No studies have examined the relationship between the intensity of facial emotion expression and theory of mind (ToM) ability in people with epilepsy. This study aimed to explore facial emotion recognition in a group of patients with frontal (FLE) or temporal lobe epilepsy (TLE) and its relationship with the intensities of perceived facial emotion expressions, ToM, and social functioning. METHODS: Twenty-six patients with FLE or TLE and 30 matched controls were included in the study. All participants completed the facial emotion recognition test, Faux Pas Recognition (FPR) test measuring advanced ToM, Symptom Checklist-90-Revised, Social and Occupational Functioning Scale for Epilepsy (SOFSE), and background neuropsychological tests. RESULTS: The patient group was significantly worse than the control group in recognizing facial expressions of negative emotions, particularly for medium-intensity facial expression of fear. There was no significant difference between the groups in recognizing high-intensity fear facial expressions. The scores of FPR (overall and affective ToMs) in the patient group were significantly lower than those in the control group. Additionally, the facial emotion recognition was significantly associated with the total score of FPR, and the FPR total score remarkably correlated with the Communication subscale score of the SOFSE. CONCLUSIONS: Patients with FLE or TLE had impaired ability to recognize medium-intensity facial expressions of fear. Moreover, patients' ToM deficit significantly correlated not only with their emotion recognition problem but also with their social-communicative competence. Nevertheless, we also found that increasing the intensity of expression can improve the accuracy of emotion recognition in patients with epilepsy. These findings may provide considerations for further longitudinal studies and interventions on the social difficulties of people with epilepsy.

Treatment with the combination of clavulanic acid and valproic acid led to recovery of neuronal and behavioral deficits in an epilepsy rat model.

Epilepsy, which is caused by abnormal neuronal firing in the brain, is a common neurological disease and affects motor and cognitive functions. Excessive levels of glutamate and insufficient levels of inhibitory GABA are involved in its pathophysiology. Valproic acid (Val), a GABAergic agonist, is one of the first-line antiepileptic drugs, but it shows many adverse side effects at the clinical dose. Clavulanic acid (CA), a ß-lactamase inhibitor, has been demonstrated to increase glutamate transporter-1 expression. This study evaluated the effects of CA and Val in an epilepsy rat model. Male Wistar rats received intraperitoneal injections of pentylenetetrazol (PTZ, 35 mg/kg, every other day, IP, for 13 days) to induce kindling epilepsy. After four times of PTZ injection, rats received daily treatment with CA (1 or 10 mg/kg, IP), Val (50 or 100 mg/kg, IP), or the combination of CA (1 mg/kg) and Val (50 mg/kg) for 7 consecutive days. Motor, learning, and memory functions were measured. Rats with PTZ-induced kindling exhibited seizures, motor dysfunction, cognitive impairment, and cell loss and reduction of neurogenesis in the hippocampus. Neither 1 mg/kg CA nor 50 mg/kg Val treatment was effective in alleviating behavioral and neuronal deficits. However, treatment with 10 mg/kg CA, 100 mg/kg Val, and the combination of 1 mg/kg CA and 50 mg/kg Val improved these behavioral and neuronal deficits. Particularly, the combination of CA and Val showed synergistic effects on seizure suppression, suggesting the potential for treating epilepsy and related neuronal damage and motor and cognitive deficits.

Treatment effects of the combination of ceftriaxone and valproic acid on neuronal and behavioural functions in a rat model of epilepsy.

NEW FINDINGS: What is the central question of this study? Imbalance of activities between GABAergic and glutamatergic systems is involved in epilepsy. It is not known whether simultaneously increasing GABAergic and decreasing glutamatergic activity using valproic acid and ceftriaxone, respectively, leads to better seizure control. What is the central question of this study? Ceftriaxone suppressed seizure and cognitive deficits and restored neuronal density and the number of newborn cells in the hippocampus in a rat model of epilepsy. Combined treatment with ceftriaxone and valproic acid showed additive effects in seizure suppression. ABSTRACT: The pathophysiology of epilepsy is typically considered as an imbalance between inhibitory GABA and excitatory glutamate neurotransmission. Valproic acid (Val), a GABA agonist, is one of the first-line antiepileptic drugs in the treatment of epilepsy, but it exhibits adverse effects. Ceftriaxone (CEF) elevates expression of glutamate transporter-1, enhances the reuptake of synaptic glutamate, increases the number of newborn cells and exhibits neuroprotective effects in animal studies. In this study, we evaluated effects of the combination of CEF and Val on behavioural and neuronal measures in a rat epilepsy model. Male Wistar rats were injected i.p. with pentylenetetrazol (35 mg/kg, every other day for 13 days) to induce the epilepsy model. Ceftriaxone (10 or 50 mg/kg), Val (50 or 100 mg/kg) or the combination of CEF and Val were injected daily after the fourth pentylenetetrazol injection for seven consecutive days. Epileptic rats exhibited seizure and impairments in motor and cognitive functions. Treatment with CEF and Val reduced the seizure and enhanced motor and cognitive functions in a dose-dependent manner. The combination of CEF (10 mg/kg) and Val (50 mg/kg) improved behaviours considerably. Histologically, compared with control animals, epileptic rats exhibited lower neuronal density and a reduction in hippocampal newborn cells but higher apoptosis in the basolateral amygdala, all of which were restored by the treatment with CEF, Val or the combination of CEF and Val. The study findings demonstrated that the combination of low doses of CEF and Val has beneficial effects on seizure suppression, neuroprotection and improvement in motor and cognitive functions in epilepsy.

Cerebroprotection by dioscin after experimental subarachnoid haemorrhage via inhibiting NLRP3 inflammasome through SIRT1-dependent pathway.

BACKGROUND AND PURPOSE: Dioscin has multiple biological activities and is beneficial for cardiovascular and cerebral vascular diseases. Here, we investigated the protective effects of dioscin against subarachnoid haemorrhage and the molecular mechanisms involved. EXPERIMENTAL APPROACH: Dioscin was administered after subarachnoid haemorrhage induced in rats. MCC950, a potent selective nod-like receptor pyrin domain-containing 3 (NLRP3) inhibitor, was used to suppress NLRP3 and EX527 (selisistat) was used to inhibit sirtuin 1 (SIRT1). KEY RESULTS: In vivo, dioscin inhibited acute inflammatory response, oxidative damage, neurological impairment and neural cell degeneration after subarachnoid haemorrhage along with dramatically suppressing NLRP3 inflammasome activation. While pretreatment with MCC950 reduced the inflammatory response and improved neurological outcomes it did not lessen ROS production. However, giving dioscin after MCC950 reduced acute brain damage and ROS production. Dioscin increased SIRT1 expression after subarachnoid haemorrhage, whereas EX527 abolished the up-regulation of SIRT1 induced by dioscin and offset the inhibitory effects of dioscin on NLRP3 inflammasome activation. EX527 pretreatment also reversed the neuroprotective effects of dioscin against subarachnoid haemorrhage. Similarly, in vitro, dioscin dose-dependently suppressed inflammatory response, oxidative damage and neuronal degeneration and improved cell viability in neurons and microglia co-culture system. These effects were associated with inhibition of the NLRP3 inflammasome and stimulation of SIRT1 signalling, which could be inhibited by EX527 pretreatment. CONCLUSION AND IMPLICATIONS: Dioscin provides protection against subarachnoid haemorrhage via the suppression of NLRP3 inflammasome activation through SIRT1-dependent pathway. Dioscin may be a new candidate to ameliorate early brain injury after subarachnoid haemorrhage.

Astaxanthin ameliorates oxidative stress and neuronal apoptosis via SIRT1/NRF2/Prx2/ASK1/p38 after traumatic brain injury in mice.

BACKGROUND AND PURPOSE: Oxidative stress and neuronal apoptosis play key roles in traumatic brain injury. We investigated the protective effects of astaxanthin against traumatic brain injury and its underlying mechanisms of action. EXPERIMENTAL APPROACH: A weight-drop model of traumatic brain injury in vivo and hydrogen peroxide exposure in vitro model were established. Brain oedema, behaviour tests, western blot, biochemical analysis, lesion volume, histopathological study and cell viability were performed. KEY RESULTS: Astaxanthin significantly reduced oxidative insults on Days 1, 3 and 7 after traumatic brain injury. Neuronal apoptosis was also ameliorated on Day 3. Additionally, astaxanthin improved neurological functions up to 3 weeks after traumatic brain injury. Astaxanthin treatment dramatically enhanced the expression of peroxiredoxin 2 (Prx2), nuclear factor-erythroid 2-related factor 2 (NRF2/Nrf2) and sirtuin 1 (SIRT1), while it down-regulated the phosphorylation of apoptosis signal-regulating kinase 1 (ASK1) and p38. Inhibition of Prx2 by siRNA injection reversed the beneficial effects of astaxanthin against traumatic brain injury. Additionally, Nrf2 knockout prevented the neuroprotective effects of astaxanthin in traumatic brain injury. In contrast, overexpression of Prx2 in Nrf2 knockout mice attenuated the secondary brain injury after traumatic brain injury. Moreover, inhibiting SIRT1 by EX527 dramatically inhibited the neuroprotective effects of astaxanthin and suppressed SIRT1/Nrf2/Prx2/ASK1/p38 pathway both in vivo and in vitro. CONCLUSION AND IMPLICATIONS: Astaxanthin improved the neurological functions and protected the brain from injury after traumatic brain injury, primarily by reducing oxidative stress and neuronal death via SIRT1/Nrf2/Prx2/ASK1/p38 signalling pathway and might be a new candidate to ameliorate traumatic brain injury.

Establishing Indicators and an Analytic Method for Moisture Susceptibility and Rutting Resistance Evaluation Using a Hamburg Wheel Tracking Test.

The Hamburg wheel tracking test (HWTT) is widely used to evaluate the performance of asphalt mixtures. According to HWTT specifications, the stripping inflection point (SIP) and the rut depth at a certain number of load cycles are two common indicators for evaluating the moisture susceptibility and rutting resistance of asphalt mixtures, respectively. Although these indicators have been used extensively by several transportation institutions, the reliability and stability in evaluating asphalt mixture behaviors of these indicators have been questioned. To more effectively evaluate the performance of asphalt mixture in the HWTT, this study introduces a novel method of analysis for the HWTT and novel indicators of rutting resistance and moisture susceptibility. The proposed method and indicators were employed to analyze the HWTT results of 14 field core specimens, and the proposed indicators were compared with conventional HWTT indicators to assess their capability of distinction between asphalt mixtures with different performance behaviors in the HWTT. The results indicate that the conventional HWTT indicators cannot effectively evaluate the asphalt mixtures with different performance in the HWTT. By contrast, the proposed analytic method and indicators have significant advantages to effectively evaluate and distinguish the rutting resistance and moisture susceptibility of asphalt mixtures.

Social functioning and health-related quality of life trajectories in people with epilepsy after epilepsy surgery.

OBJECTIVE: An improvement in quality of life (QoL) over time after epilepsy surgery has been demonstrated in people with epilepsy; however, social functioning has been less investigated. We conducted this study to examine whether postsurgical improvement is parallel between QoL and social functioning in patients with epilepsy. METHODS: We retrospectively reviewed patients who underwent epilepsy surgery. All participants completed a comprehensive neuropsychological assessment, the Quality of Life in Epilepsy Inventory (QOLIE-89) questionnaire, and the Social and Occupational Functioning Scale for Epilepsy (SOFSE) before surgery and at 3 months, 6 months, and 1 year after surgery. Demographic and epilepsy-related information was also collected. Generalized estimating equations with identity links were used to model the QOLIE-89 and SOFSE over time and possible associated factors. A p < 0.05 was considered statistically significant. RESULTS: A total of 76 patients, including 36 males and 43 females aged 18 to 62 years were collected. Both total QOLIE-89 and overall SOFSE improved over time after epilepsy surgery (adjusted p value < 0.001 and 0.002, respectively, with Bonferroni's correction). Total QOLIE-89 improved 3 months after surgery, while overall SOFSE showed no significant improvement until 6 months after surgery. The presurgical Full-Scale Intelligence Quotient (FSIQ) of the Wechsler Adult Intelligence Scale-III and years of education were significantly associated with time-dependent improvement for both total QOLIE-89 and overall SOFSE (p value < 0.001). At one year after surgery, overall SOFSE and total QOLIE-89 scores were significantly higher in the seizure-free group than in the nonseizure-free group (p value = 0.040 and 0.032, respectively). CONCLUSION: Social functioning significantly improved in people with epilepsy after surgery as QoL, but it took more time to exhibit improvement. People with better FSIQ and more years of education had better improvement in social functioning over time. The early intervention of rehabilitation programs after epilepsy surgery might be necessary to facilitate the improvement in social functioning.

Meta-Analysis of Pterional Versus Supraorbital Keyhole Approach for Clipping Intracranial Aneurysms: Direct Comparison of Approach-Related Complications.

BACKGROUND: The supraorbital keyhole approach (SKA) and pterional approach (PA) have been recommended for clipping intracranial aneurysms (IAs). We conducted a meta-analysis to systematically and comprehensively compare the clinical outcomes between the 2 approaches. METHODS: We retrieved potential academic studies that had compared the clinical outcomes of SKA and PA for clipping IAs from the Cochrane Library, Medline, PubMed, and EMBASE databases. The references of the identified studies were carefully reviewed to ensure that all available documents had been included in the present study. The meta-analysis was conducted in accordance with the acknowledged PRISMA (prioritized reported items for systematic review and meta-analysis) guidelines. RESULTS: A total of 8 studies with 1016 participants were included in our study. Of these 1016 patients with IAs, 518 were assigned to the SKA group and 498 to the PA group. Our results showed that the SKA is preferable for clipping IAs compared with the PA because of its shorter operative time (weighted mean difference, -3.163; 95% confidence interval [CI], -5.577 to -0.749; P = 0.01) and length of stay, especially for the patients with unruptured IAs (weighted mean difference, -52.107; 95% CI, -81.597 to -22.618). However, a lower risk of postoperative infection was found in the SKA group (4.6% vs. 8.5%; odds ratio, 0.398; 95% CI, 0.186-0.850; P = 0.017), which seemed to derive from the ruptured IAs (OR, 0.330; 95% CI, 0.136-0.800; P = 0.014). However, no significant difference was found in the incidence of frontal sinus opening, completed occlusion, procedural complications, intraoperative rupture, postoperative hematoma, or cerebral vessel spasm. CONCLUSIONS: The SKA was associated with a shorter length of stay and operation time and a lower risk of postoperative infection. Hence, the SKA seems to be equally effective and safe for clipping IAs compared with PA and should be recommended for surgeons who have acquired sufficient experience with this technique owing to its advantages compared with the PA.

Unambiguous Sequence Characterization of a Monoclonal Antibody in a Single Analysis Using a Nonspecific Immobilized Enzyme Reactor.

Accurate sequence characterization is essential for the development of therapeutic antibodies by the pharmaceutical industry. Presented here is a methodology to obtain comprehensive sequence analysis of a monoclonal antibody. An enzyme reactor of immobilized Aspergillopepsin I, a highly stable nonspecific protease, was used to cleave reduced antibody subunits into a peptide profile ranging from 1 to 20 kDa. Utilizing the Thermo Orbitrap Fusion's unique instrument architecture combined with state-of-the-art instrument control software allowed for dynamic instrument methods that optimally characterize eluting peptides based on their size and charge density. Using a data-dependent instrument method, both collisional dissociation and electron transfer dissociation were used to fragment the appropriate charge state of analyte peptides. The instrument layout also allowed for scans to be taken in parallel using both the ion trap and Orbitrap concurrently, thus allowing larger peptides to be analyzed in high resolution using the Orbitrap while simultaneously analyzing tryptic-like peptides using the ion trap. We harnessed these capabilities to develop a custom method to optimally fragment the eluting peptides based on their mass and charge density. Using this approach, we obtained 100% sequence coverage of the total antibody in a single chromatographic analysis, enabling unambiguous sequence assignment of all residues.

Specific and high-resolution identification of monoclonal antibody fragments detected by capillary electrophoresis-sodium dodecyl sulfate using reversed-phase HPLC with top-down mass spectrometry analysis.

In recent years, capillary electrophoresis-sodium dodecyl sulfate (cSDS) has been widely used for high resolution separation and quantification of the fragments and aggregates of monoclonal antibodies (mAbs) to ensure the quality of mAb therapeutics. However, identification of the low-molecular-weight (LMW) and high-molecular-weight (HMW) species detected in cSDS electropherograms has been based primarily on the approximate MWs calculated from standard curves using known MW standards and correlations with fragments and aggregates identified by other methods. It is not easy to collect sufficient amounts of H/LMW species from cSDS for analysis by orthogonal methods and the direct coupling of cSDS with mass spectrometry (MS) is very difficult due to interference from SDS. In this study, we describe the precise identification of H/LMW species detected by cSDS using reversed-phase high performance liquid chromatography (RP-HPLC) coupled with top-down tandem MS analysis. The H/LMW species were first identified by on-line RP-HPLC MS analysis and the RP-HPLC fractions were then analyzed by cSDS to connect the identified H/LMW species with the peaks in the cSDS electropherogram. With this method, 58 unique H/LMW species were identified from an immunoglobulin G1 (IgG1) mAb. The identified fragments ranged from 10 kDa single chain fragments to 130 kDa triple chain fragments, including some with post-translational modifications. This is the first study to clearly identify the antibody fragments, including the exact clipping sites, observed in cSDS electropherograms. The methodology and results presented here should be applicable to most other IgG1 mAbs.

Preliminary comparison of neuropsychological performance in patients with non-small-cell lung cancer treated with chemotherapy or targeted therapy.

PURPOSE: This cross-sectional pilot study aimed to compare the effects of chemotherapy and targeted therapy on neuropsychological performance and psychiatric symptoms in patients with non-small-cell lung cancer (NSCLC). PATIENTS AND METHODS: A total of 113 patients with NSCLC were recruited. According to their type of cancer treatment, the patients were classified into chemotherapy (n=40), targeted therapy (n=33), and untreated control (n=40) groups. All participants completed five objective tests measuring various domains of cognitive function, a subjective cognitive functioning scale (Functional Assessment of Cancer Therapy-Cognitive Function; FACT-cog), and the Hospital Anxiety and Depression Scale (HADS) either within 6 months after diagnosis (for the untreated group) or about 18 months after treatment. RESULTS: Overall, there were no significant intergroup differences in the proportions of patients with abnormal cognitive performance and psychiatric disturbances. Among the untreated NSCLC patients, 35% had impaired performance in at least one cognitive domain, and a comparable finding (30%-35%) was made for the other two treatment groups. The proportion of patients with impaired psychomotor speed was the highest (10%-15%) across various cognitive domains. Moreover, a significant proportion of NSCLC patients (15%-20%) exhibited HADS-defined anxiety and depression disorder. Finally, significant correlations were found between FACT-cog total scores and the HADS Depression subscale across all three groups. CONCLUSION: This study demonstrated that 1) a substantial proportion of NSCLC patients exhibited cognitive impairments (especially regarding psychomotor speed) and psychiatric disturbances; 2) no significant differences were observed among the three patient groups for any subjective or objective measure of cognitive deficit; and 3) perceived cognitive impairment was significantly associated with depression or anxiety. Prompt treatment of psychiatric disorders to minimize their impact is therefore recommended.

Systematic and Comprehensive Comparison of Incidence of Restenosis Between Carotid Endarterectomy and Carotid Artery Stenting in Patients with Atherosclerotic Carotid Stenosis.

OBJECTIVE: The purpose of the present study was to conduct a meta-analysis to systematically compare the incidence rates of in-stent restenosis after carotid artery stenting (CAS) and restenosis after carotid endarterectomy (CEA) for patients with atherosclerotic carotid stenosis. METHODS: We retrieved potential academic reports comparing restenosis between CEA and CAS from the MEDLINE, PubMed, and EMBASE databases and the Cochrane Library from the date of the first CEA (January 1951) to July 20, 2018. The references of the identified studies were carefully reviewed to ensure that all available reports were included in the present study. RESULTS: Our meta-analysis included 27 studies (15 randomized controlled trials, 12 nonrandomized controlled trials) and 20,479 participants with atherosclerotic carotid stenosis. A statistically significant difference was found in the cumulative incidence of restenosis >70% between CEA and CAS (risk difference, -0.033, 95% confidence interval [CI] -0.054 to -0.013; P = 0.002). For the restenosis >70% outcomes, although CEA was relevant with a lower rate of restenosis than CAS within 6 months (odds ratio [OR], 0.495; 95% CI, 0.285-0.861; P = 0.013) and 1 year (OR, 0.626; 95% CI, 0.483-0.811; P < 0.001), no statistically significant differences were found at 1.5 years (P = 0.210), 2 years (P = 0.123), 4 years (P = 0.124), 5 years (P = 0.327), or 10 years (P = 0.839). For the restenosis >50% outcomes, a significant difference was found in the rate of restenosis between the CEA and CAS groups within 1 year (OR, 0.317; 95% CI, 0.228-0.441; P < 0.001) but not at 1.5 years (P = 0.301), 2 years (P = 0.686), or 5 years (P = 0.920). No nominally significant effects were demonstrated with respect to the cumulative incidence of occlusion (P = 0.195) or the cumulative incidence of restenosis for symptomatic patients (P = 0.170) between CEA and CAS. CONCLUSIONS: Although CAS was preferred over CEA, regardless of restenosis >50% or >70% after revascularization within 1 year, no significant difference was observed with extension of the follow-up period to >1 year. CAS was not associated with a greater cumulative incidence of occlusion or the cumulative incidence of restenosis for symptomatic patients.

O-GlcNAc Site Mapping by Using a Combination of Chemoenzymatic Labeling, Copper-Free Click Chemistry, Reductive Cleavage, and Electron-Transfer Dissociation Mass Spectrometry.

As a dynamic post-translational modification, O-linked ß- N-acetylglucosamine ( O-GlcNAc) modification (i.e., O-GlcNAcylation) of proteins regulates many biological processes involving cellular metabolism and signaling. However, O-GlcNAc site mapping, a prerequisite for site-specific functional characterization, has been a challenge since its discovery. Herein we present a novel method for O-GlcNAc enrichment and site mapping. In this method, the O-GlcNAc moiety on peptides was labeled with UDP-GalNAz followed by copper-free azide-alkyne cycloaddition with a multifunctional reagent bearing a terminal cyclooctyne, a disulfide bridge, and a biotin handle. The tagged peptides were then released from NeutrAvidin beads upon reductant treatment, alkylated with (3-acrylamidopropyl)trimethylammonium chloride, and subjected to electron-transfer dissociation mass spectrometry analysis. After validation by using standard synthetic peptide gCTD and model protein α-crystallin, such an approach was applied to the site mapping of overexpressed TGF-ß-activated kinase 1/MAP3K7 binding protein 2 (TAB2), with four O-GlcNAc sites unambiguously identified. Our method provides a promising tool for the site-specific characterization of O-GlcNAcylation of important proteins.

A disposable breath sensing tube with on-tube single-nanowire sensor array for on-site detection of exhaled breath biomarkers.

This paper presents a novel disposable breath-sensing tube with an on-tube single-nanowire (NW) sensor array for noninvasive, simple, and on-site detection of exhaled breath biomarkers. Although various noninvasive detection methods for lung cancer biomarkers in breath samples exist, they are unsuitable for self-diagnostics and immediate detection because they entail complicated handling procedures and are time intensive. In this study, we simulated, fabricated, and characterized disposable nanosensors by using single TiO2 and Ag NWs in flexible plastic tubes. The proposed sensors simultaneously detect 2-propyl-1-pentanol (2-PP) lung cancer biomarkers and changes in the humidity, flow rate, and temperature of the flowing gas. The optimal dimension of the tubes was determined and verified through dynamic simulations and experiments. The current tube design decreases sensing variation and moisture interference by 43.28% and 78.77%, respectively, compared with previous designs. In the future, the proposed breath sensor, which has a response time of less than 10 s, can be used in a tube for simple and quick screening of lung cancer patients with the 2-PP concentration exceeding 100 ppb. The developed breath-sensing platform can be applied in noninvasive on-site disease screening.

Analysis of Monoclonal Antibody Sequence and Post-translational Modifications by Time-controlled Proteolysis and Tandem Mass Spectrometry.

Methodology for sequence analysis of ∼150 kDa monoclonal antibodies (mAb), including location of post-translational modifications and disulfide bonds, is described. Limited digestion of fully denatured (reduced and alkylated) antibody was accomplished in seconds by flowing a sample in 8murea at a controlled flow rate through a micro column reactor containing immobilized aspergillopepsin I. The resulting product mixture containing 3-9 kDa peptides was then fractionated by capillary column liquid chromatography and analyzed on-line by both electron-transfer dissociation and collisionally activated dissociation mass spectrometry (MS). This approach enabled identification of peptides that cover the complete sequence of a murine mAb. With customized tandem MS and ProSightPC Biomarker search, we verified 95% amino acid residues of this mAb and identified numerous post-translational modifications (oxidized methionine, pyroglutamylation, deamidation of Asn, and several forms ofN-linked glycosylation). For disulfide bond location, native mAb is subjected to the same procedure but with longer digestion times controlled by sample flow rate through the micro column reactor. Release of disulfide containing peptides from accessible regions of the folded antibody occurs with short digestion times. Release of those in the interior of the molecule requires longer digestion times. The identity of two peptides connected by a disulfide bond is determined using a combination of electron-transfer dissociation and ion-ion proton transfer chemistry to read the two N-terminal and two C-terminal sequences of the connected peptides.