Roles of p62 in BDNF-dependent autophagy suppression and neuroprotection against mitochondrial dysfunction in rat cortical neurons.

Previously, we have reported that pre-conditioning of primary rat cortical neurons with brain-derived neurotrophic factor (BDNF) may exert neuroprotective effects against 3-nitropropionic acid (3-NP), a mitochondrial complex II inhibitor. However, the underlying mechanisms, especially potential involvements of autophagy, remain elusive. In this work, we tested the hypothesis that BDNF may suppress 3-NP-induced autophagy to exert its neuroprotective effects by inducing the expression of p62/sequestosome-1 in primary cortical neurons. We found that 3-NP increased total level of microtubule-associated protein 1A/1B-light chain (LC)-3 as well as the LC3-II/LC3-I ratio, an index of autophagy, in primary cortical neurons. BDNF decreased LC3-II/LC3-I ratio and time-dependently induced expression of p62. Knockdown of p62 by siRNA restored LC3-II/LC3-I ratio and increased total LC3 levels associated with BDNF exposure; p62 knockdown also abolished BDNF-dependent neuroprotection against 3-NP. Upstream of p62, we found that BDNF triggered phosphorylation of mammalian target of rapamycin (mTOR) and its downstream mediator p70S6K; importantly, the mTOR inhibitor rapamycin reduced both BDNF-dependent p62 induction as well as 3-NP resistance. BDNF is known to induce c-Jun in cortical neurons. We found that c-Jun knockdown in part attenuated BDNF-mediated p62 induction, whereas p62 knockdown had no significant effects on c-Jun expression. In addition to suppressing p62 induction, rapamycin also partially suppressed BDNF-induced c-Jun expression, but c-Jun knockdown failed to affect mTOR activation. Together, our results suggested that BDNF inhibits 3-NP-induced autophagy via, at least in part, mTOR/c-Jun-dependent induction of p62 expression, together contributing to neuroprotection against mitochondrial inhibition.

Oncostatin M-dependent Mcl-1 induction mediated by JAK1/2-STAT1/3 and CREB contributes to bioenergetic improvements and protective effects against mitochondrial dysfunction in cortical neurons.

Oncostatin M (OSM), a cytokine in the interleukin-6 (IL-6) family, has been proposed to play a protective role in the central nervous system, such as attenuation of excitotoxicity induced by N-methyl-D-aspartate (NMDA) and glutamate. However, the potential neuroprotective effects of OSM against mitochondrial dysfunction have never been reported. In the present study, we tested the hypothesis that OSM may confer neuronal resistance against 3-nitropropionic acid (3-NP), a plant toxin that irreversibly inhibits the complex II of the mitochondrial electron transport chain, and characterized the underlying molecular mechanisms. We found that OSM preconditioning dose- and time-dependently protected cortical neurons against 3-NP toxicity. OSM stimulated expression of myeloid cell leukemia-1 (Mcl-1), an anti-apoptotic Bcl-2 family member expressed in differentiating myeloid cells, that required prior phosphorylation of Janus kinase-1 (JAK1), JAK2, extracellular signal-regulated kinase-1/2 (ERK1/2), signal transducer and activator of transcription-3 (STAT3), STAT1, and cAMP-response element-binding protein (CREB). Pharmacological inhibitors of JAK1, JAK2, ERK1/2, STAT3, STAT1, and CREB as well as the siRNA targeting at STAT3 and Mcl-1 all abolished OSM-dependent 3-NP resistance. Finally, OSM-dependent Mcl-1 induction contributed to the enhancements of mitochondrial bioenergetics including increases in spare respiratory capacity and ATP production. In conclusion, our findings indicated that OSM induces Mcl-1 expression via activation of ERK1/2, JAK1/2, STAT1/3, and CREB; furthermore, OSM-mediated Mcl-1 induction contributes to bioenergetic improvements and neuroprotective effects against 3-NP toxicity in cortical neurons. OSM may thus serve as a novel neuroprotective agent against mitochondrial dysfunction commonly associated with pathogenic mechanisms underlying neurodegeneration.

Incidence, prevalence, and medical expenditures of classical amyotrophic lateral sclerosis in Taiwan, 1999-2008.

BACKGROUND/PURPOSE: Amyotrophic lateral sclerosis (ALS) is a rare disease, which makes the estimation of incidence and prevalence difficult in Taiwan. This study was conducted to investigate the incidence, prevalence, and medical expenditure of ALS in Taiwan. METHODS: Patients who had at least one service claim either as an outpatient or inpatient between the years 2004 and 2007 and were over 15 years of age with a primary diagnosis of ALS were identified from the National Health Insurance Research Database. Additionally, ALS patients with serious disability database certificates over 15 years of age were included for the calculation of incidence and prevalence between the years 1999 and 2008. Lastly, the total medical expenditure, including ventilator use and riluzole, were reported. RESULTS: In 2006 and 2008, the average annual incidence and prevalence of ALS was 0.51 and 1.97 (per 10(5)), respectively, in Taiwan. The male-to-female ratio of incidence for ALS was 1.67. The average medical expenditure for ALS patients stayed steady at 16-fold greater than the general population of Taiwan in 2008. The percentage of ventilator and riluzole expenditure as a proportion of total medical expense decreased from 55% in 2000 to 33% in 2008. CONCLUSION: The incidence and average medical expenditure of ALS patients remained stable over the years in Taiwan, however, as a proportion of total medical expenses, expenditure on ventilator and riluzole decreased over the study period.

Elevated serum autoantibody against high mobility group box 1 as a potent surrogate biomarker for amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a complicate and progressive onset devastating neurodegenerative disease. Its pathogenic mechanisms remain unclear and there is no specific test for diagnosis. For years, researchers have been vigorously searching for biomarkers associated with ALS to assist clinical diagnosis and monitor disease progression. Some specific inflammatory processes in the central nervous system have been reported to participate in the pathogenesis of ALS. As high mobility group box 1 (HMGB1) is elevated in spinal cord tissues of patients with ALS, we hypothesized, therefore, that serum autoantibody against HMGB1 (HMGB1 autoAb) might represent an effective biomarker for ALS. Patients with ALS, Alzheimer's disease, Parkinson's disease, and healthy age-matched control subjects were recruited for this study. ALS group consisted of 61 subjects, the other groups each consisted of forty subjects. We generated a polyclonal antibody against HMGB1 and developed an ELISA-based methodology for screening serum samples of these subjects. All samples were coded for masked comparison. For statistic analyses, two-tailed Student's t-test, ANOVA, Bonferroni multiple comparison test, Spearman correlation, and receiver operating characteristic curve were applied. We discovered that the level of HMGB1 autoAb significantly increased in patients with ALS as compared with that of patients with Alzheimer's disease, Parkinson's disease, and healthy control subjects. The differences between all groups were robust even at the early stages of ALS progression. More importantly, higher HMGB1 autoAb level was found in more severe disease status with significant correlation. Our study demonstrates that serum HMGB1 autoAb may serve as a biomarker for the diagnosis of ALS and can be used to monitor disease progression.

Riluzole and prognostic factors in amyotrophic lateral sclerosis long-term and short-term survival: a population-based study of 1149 cases in Taiwan.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a rare disease in Taiwan; thus, estimation of ALS mortality is difficult. We evaluated factors associated with ALS survival in Taiwan. METHODS: The study enrolled 1149 Taiwanese with a primary diagnosis of ALS during 1999-2008. Follow-up information was available for all patients; mean (SD) duration of follow-up was 2.91 (2.62) years. Medical interventions, including noninvasive positive pressure ventilation (NIPPV), tracheotomy, gastrostomy, and riluzole, were included in time-dependent survival analysis. RESULTS: Of the 1149 ALS patients, 438 (38.12%) died during follow-up. Mortality in the first year was 16%, which was 13 times (95% CI 11.1-15.2) the age- and sex-standardized rate of the general population in Taiwan. The average annual crude mortality rate was 13.1% (person-years). Factors significantly associated with increased mortality were male sex, advanced age, rural residence, lower economic status, no tracheotomy, and no riluzole treatment. Significant predictors of long-term versus average survival were younger age at diagnosis, being a dependent or receiving social welfare, and NIPPV support. Significant predictors of short-term versus average survival were older age, being employed, no tracheotomy, and no riluzole use. CONCLUSIONS: The results support the use of riluzole to improve ALS survival. Patients who received riluzole and underwent tracheotomy had the best survival.

Eosinophil-derived neurotoxin is elevated in patients with amyotrophic lateral sclerosis.

BACKGROUND AND OBJECTIVES: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by loss of motor neurons in the brainstem, motor cortex, and spinal cord. Oxidative stress and neuroinflammation have been implicated in the pathophysiology of ALS. Members of the family of damage-associated molecular patterns, including reactive oxygen species, high-mobility group box 1, and eosinophil-derived neurotoxin (EDN), may participate in pathological conditions. In this study, we aim to discover new biomarker for detecting ALS. MATERIALS AND METHODS: We examined 44 patients with ALS, 41 patients with Alzheimer's disease, 41 patients with Parkinson's disease, and 44 healthy controls. The concentration of serum EDN was measured using an enzyme-linked immunosorbent assay. RESULTS: EDN levels were significantly increased 2.17-fold in the serum of patients with ALS as compared with healthy controls (P < 0.05). No correlation between the levels of serum EDN and various clinical parameters of ALS was found. Moreover, the levels of serum EDN in patients with Parkinson's disease and Alzheimer's disease and healthy controls were similar. CONCLUSION: A higher level of serum EDN was found specifically in patients with ALS, indicating that EDN may participate in the pathophysiology of ALS.

Evaluation of the Photoplethysmogram-Based Deep Learning Model for Continuous Respiratory Rate Estimation in Surgical Intensive Care Unit.

The respiratory rate (RR) is a significant indicator to evaluate a patient's prognosis and status; however, it requires specific instrumentation or estimates from other monitored signals. A photoplethysmogram (PPG) is extensively used in clinical environments as well as in intensive care units (ICUs) to primarily monitor peripheral circulation while capturing indirect information about intrathoracic pressure changes. This study aims to apply and evaluate several deep learning models using a PPG for the continuous and accurate estimation of the RRs of patients. The dataset was collected twice for 2 min each in 100 patients aged 18 years and older from the surgical intensive care unit of a tertiary referral hospital. The BIDMC and CapnoBase public datasets were also analyzed. The collected dataset was preprocessed and split according to the 5-fold cross-validation. We used seven deep learning models, including our own Dilated Residual Neural Network, to check how accurately the RR estimates match the ground truth using the mean absolute error (MAE). As a result, when validated using the collected dataset, our model showed the best results with a 1.2628 ± 0.2697 MAE on BIDMC and RespNet and with a 3.1268 ± 0.6363 MAE on our dataset, respectively. In conclusion, RR estimation using PPG-derived models is still challenging and has many limitations. However, if there is an equal amount of data from various breathing groups to train, we expect that various models, including our Dilated ResNet model, which showed good results, can achieve better results than the current ones.

c-Jun-dependent sulfiredoxin induction mediates BDNF protection against mitochondrial inhibition in rat cortical neurons.

In current study, we tested the hypothesis that c-Jun-dependent sulfiredoxin expression mediates protective effects of brain-derived neurotrophic factor (BDNF) against neurotoxicity induced by 3-nitropropionic acid (3-NP), a mitochondrial complex II inhibitor, in primary rat cortical cultures. We found that BDNF-dependent c-Jun expression and nuclear translocation required prior phosphorylation of extracellular signal-regulated kinase (ERK)1/2, but not Akt. BDNF also transiently activated the expression of sulfiredoxin, an ATP-dependent antioxidant enzyme, at both mRNA and protein levels. Furthermore, both c-Jun siRNA and ERK1/2 inhibitor PD98059 suppressed BDNF-induced sulfiredoxin expression. Finally, PD98059, c-Jun siRNA, and sulfiredoxin siRNA all abrogated BDNF-mediated 3-NP resistance. Together, these results established a signaling cascade of "BDNF → ERK1/2-Pi → c-Jun → sulfiredoxin → 3-NP resistance". We therefore conclude that c-Jun-induced sulfiredoxin mediates the BDNF-dependent neuroprotective effects against 3-NP toxicity in primary rat cortical neurons, at least in part.

Erythropoietin and sonic hedgehog mediate the neuroprotective effects of brain-derived neurotrophic factor against mitochondrial inhibition.

Brain-derived neurotrophic factor (BDNF) deficiency and mitochondrial dysfunction have been implicated in the pathogenesis of Huntington's disease (HD). 3-Nitropropionic acid (3-NP) is a mitochondrial inhibitor commonly used as a pharmacological model mimicking HD. We have recently reported that preconditioning of primary rat cortical cultures with BDNF induces sonic hedgehog (SHH), which contributes to the protective effects of BDNF against 3-NP neurotoxicity. Because carbamylated erythropoietin (EPO) may induce SHH, we investigated whether BDNF-dependent SHH expression and 3-NP resistance require prior induction of EPO. We found that BDNF induced EPO expression at both mRNA and protein levels. BDNF-mediated SHH induction and 3-NP resistance were abolished by the soluble EPO receptor (sEPO-R), an EPO inhibitor. Recombinant rat EPO (rEPO) induced SHH and attenuated 3-NP neurotoxicity. The rEPO-dependent neuroprotection was suppressed by the SHH inhibitor cyclopamine (CPM); however, sEPO-R failed to affect SHH neuroprotection. Furthermore, the rEPO-dependent neuroprotection was not suppressed by the BDNF neutralizing antibody, which completely abolished BDNF-mediated 3-NP resistance at the same dosage. Overall, our results demonstrate that BDNF-dependent SHH expression and 3-NP resistance require prior induction of EPO, thus establishing a signaling cascade of "BDNF-->EPO-->SHH-->3-NP resistance" in rat cortical neurons.

Sonic hedgehog mediates BDNF-induced neuroprotection against mitochondrial inhibitor 3-nitropropionic acid.

Sonic hedgehog (SHH), a morphogen critical for embryogenesis, has also been shown to be neuroprotective. We have recently reported that pretreatment of rat cortical neurons for 8 h with brain-derived neurotrophic factor (BDNF; 100 ng/ml) affords protection against neurotoxicity of 3-nitropropionic acid (3-NP; 2.5 mM for 24 h), a mitochondrial complex II inhibitor. However, whether SHH is involved in BDNF-mediated neuroprotection remains unknown. Herein we tested whether BDNF induces SHH expression and if so, whether BDNF induction of SHH contributes to the observed neuroprotective effects. We found BDNF (100 ng/ml) increased SHH expression at both mRNA and protein levels. BDNF protection against 3-NP was abolished by cyclopamine (CPM; 5 microM), the SHH pathway inhibitor. Preconditioning of cortical neurons with N-terminal fragment of SHH (SHH-N; 0.1-1 ng/ml) was sufficient to confer resistance. These results indicate that BDNF induces SHH expression, which contributes to neuroprotection against 3-NP toxicity in rat cortical neurons.

Olfactory neuropathy in severe acute respiratory syndrome: report of A case.

This case was a 27 years old female with severe acute respiratory syndrome (SARS). She suffered from typical symptoms of SARS. Although she got almost complete recovery from most symptoms after treatment, she noted acute onset complete anosmia 3 weeks after the onset of her first symptom. Her brain MRI examination did not show definite lesion except an incidental finding of left temporal epidermoid cyst. Her anosmia persisted for more than 2 years during following up. Peripheral neuropathy and myopathy have been reported as a concomitant problem during the convalescent stage of SARS, while the sequel of permanent ansomia in SARS was not reported before. Olfactory neuropathy, which rarely occurred in typical peripheral neuropathy, could be a special type of neuropathy induced by corona virus infection in SARS. Olfactory function test should be taken into routine check-up for patients with SARS. The pathophysiology and therapeutic strategy of this special type of permanent olfactory dysfunction deserve further investigation.

Decreased level of serum autoantibody against LG72 is a biosignature of amyotrophic lateral sclerosis.

AIM: LG72 can increase mitochondrial ROSs and oxidative stress has been implicated in the pathophysiology of amyotrophic lateral sclerosis (ALS). The serum level of LG72 or LG72-related molecules might therefore be associated with ALS. Here, we aim to determine the serum autoantibody against LG72 has potential as a biomarker for the diagnosis of ALS. MATERIALS: Seventy-eighty patients with ALS, 45 patients with AD, 43 patients with PD and 88 healthy adults were enrolled. RESULTS: The concentration of serum autoantibody against LG72 was more than fourfold lower in ALS than other control groups (p < 0.001). The AUC was 0.9627 when the cut-off value for autoantibody concentration was 0.167 µg/ml. CONCLUSION: This finding suggests that the autoantibody against LG72 might serve as a surrogate biomarker for ALS.

A genome-wide association study on amyotrophic lateral sclerosis in the Taiwanese Han population.

Identification of mutations in patients with amyotrophic lateral sclerosis (ALS) in a genome-wide association study can reveal possible biomarkers of such a rapidly progressive and fatal neurodegenerative disease. It was observed that significant single nucleotide polymorphisms vary when the tested population changes from one ethnic group to another. To identify new loci associated with ALS susceptibility in the Taiwanese Han population, we performed a genome-wide association study on 94 patients with sporadic ALS and 376 matched controls. We uncovered two new susceptibility loci at 13q14.3 (rs2785946) and 11q25 (rs11224052). In addition, we analyzed the functions of all the associated genes among 54 significant single nucleotide polymorphisms using Gene Ontology annotations, and the results showed several statistically significant neural- and muscle-related Gene Ontology terms and the associated diseases.

Nuclear Factor-kappaB-Dependent Sestrin2 Induction Mediates the Antioxidant Effects of BDNF Against Mitochondrial Inhibition in Rat Cortical Neurons.

Brain-derived neurotrophic factor (BDNF), in addition to its neurotrophic action, also possesses antioxidant activities. However, the underlying mechanisms remain to be fully defined. Sestrin2 is a stress-responsive gene implicated in the cellular defense against oxidative stress. Currently, the potential functions of sestrin2 in nervous system, in particular its correlation with neurotrophic factors, have not been well established. In this study, we hypothesized that BDNF may enhance sestrin2 expression to confer neuronal resistance against oxidative stress induced by 3-nitropropionic acid (3-NP), an irreversible mitochondrial complex II inhibitor, and characterized the molecular mechanisms underlying BDNF induction of sestrin2 in primary rat cortical cultures. We found that BDNF-mediated sestrin2 expression in cortical neurons required formation of nitric oxide (NO) with subsequent production of 3',5'-cyclic guanosine monophosphate (cGMP) and activation of cGMP-dependent protein kinase (PKG). BDNF induced localization of nuclear factor-kappaB (NF-κB) subunits p65 and p50 into neuronal nuclei that required PKG activities. Interestingly, BDNF exposure led to formation of a protein complex containing at least PKG-1 and p65/p50, which bound to sestrin2 promoter with resultant upregulation of its protein products. Finally, BDNF preconditioning mitigated production of reactive oxygen species (ROS) as a result of 3-NP exposure; this antioxidative effect of BDNF was dependent upon PKG activity, NF-κB, and sestrin2. Taken together, our results indicated that BDNF enhances sestrin2 expression to confer neuronal resistance against oxidative stress induced by 3-NP through attenuation of ROS formation; furthermore, BDNF induction of sestrin2 requires activation of a pathway involving NO/PKG/NF-κB.

Inhibitor of Differentiation-1 and Hypoxia-Inducible Factor-1 Mediate Sonic Hedgehog Induction by Amyloid Beta-Peptide in Rat Cortical Neurons.

One major pathological hallmark of Alzheimer's disease (AD) is the accumulation of senile plaques mainly composed of neurotoxic amyloid beta-peptide (Aß) in the patients' brains. Sonic hedgehog (SHH) is a morphogen critically involved in the embryonic development of the central nervous system (CNS). In the present study, we tested whether Aß may induce SHH expression and explored its underlying mechanisms. We found that both Aß25-35 and Aß1-42 enhanced SHH expression in the primary cortical neurons derived from fetal rat brains. Immunohistochemistry revealed heightened expression of SHH in the cortex and hippocampus of aged (9 and 12 months old) AD transgenic mouse brains as compared to age-matched littermate controls. Chromatin immunoprecipitation (ChIP) assay demonstrated that Aß25-35 enhanced binding of hypoxia-inducible factor-1 (HIF-1) to the promoter of the Shh gene in primary cortical cultures; consistently, Aß25-35 induction of SHH was abolished by HIF-1α small interfering RNA (siRNA). Aß25-35 also time-dependently induced inhibitor of differentiation-1 (Id1) that has been shown to stabilize HIF-1α; further, Aß25-35-mediated induction of HIF-1α and SHH was both suppressed by Id1 siRNA. Pharmacological induction of HIF-1α by cobalt chloride and application of the cell-permeable recombinant Id1 proteins were both sufficient to induce SHH expression. Finally, both the SHH pathway inhibitor cyclopamine and its neutralizing antibody attenuated Aß cytotoxicity, albeit to a minor extent. These results thus established a signaling cascade of "Aß â†’ Id1 → HIF-1 → SHH" in primary rat cortical cultures; furthermore, SHH may in part contribute to Aß neurotoxicity.

Doppler velocity criteria based on receiver operating characteristic analysis for the detection of threshold carotid stenoses.

BACKGROUND AND PURPOSE: Duplex scanning is an accepted method for noninvasive evaluation of carotid stenosis. However, the ultrasound criteria used for the detection of threshold stenoses vary widely between laboratories, and quality assurance measures to allow adjustment of criteria are often lacking. This study was completed using receiver operating characteristic (ROC) analysis to determine Doppler velocity criteria for threshold carotid stenoses, compared to an accepted standard, and to demonstrate methods to allow adjustment of criteria. METHODS: The study cohort included 134 patients who had carotid endarterectomy. Ultrasound and arteriographic data were collected for both the operated and nonoperated sides. Each carotid artery was treated as an independent case in the final analysis. Angiograms were used as the gold standard in ROC analysis to determine the Doppler velocity criteria for the detection of different threshold stenoses. RESULTS: The ROC analysis results showed that for the detection of 70% stenosis, the best Doppler systolic criterion was 200 cm/s (sensitivity 93.6%, specificity 71.7%, area under the curve [AUC] 87.6%), the best diastolic criterion was 65 cm/s (sensitivity 85.1%, specificity 74.6%, AUC 84.3%), and the best criterion of carotid ratio (CR) (internal carotid artery systolic velocity/common carotid artery systolic velocity) was 3.0 (sensitivity 78.7%, specificity 75.4%, AUC 81.3%). For 50% stenosis, the best systolic criterion was 140 cm/s (sensitivity 90.3%, specificity 95.2%, AUC 97.0%), the best diastolic criterion was 60 cm/s (sensitivity 98.6%, specificity 77.8%, AUC 92.1%), and the best criterion of CR was 2.5 (sensitivity 93.1%, specificity 72.0%, AUC 89.0%). CONCLUSIONS: This study showed that duplex scanning is able to detect threshold carotid stenoses. For the best performance, each laboratory should have its own criteria; however, the criteria provided here could be a helpful reference to those laboratories that have not yet established their own criteria. Most important, this study provides an example of how to evaluate the performance criteria, how to modify them, how such changes can affect performance, and how performance can be modified depending on the goals of the laboratory.

A multiple regression model of combined duplex criteria for detecting threshold carotid stenosis and predicting the exact degree of carotid stenosis.

BACKGROUND AND PURPOSE: Combined duplex criteria are commonly used in vascular laboratories for evaluating carotid stenosis. However, most of these combinations are empirical, and systemic validation is lacking. This study was completed using a multiple regression method to evaluate the accuracy of different combined duplex criteria for detecting threshold carotid stenosis and predicting the exact degree of carotid stenosis on angiography. METHODS: Two hundred sixty-six sets of unilateral carotid duplex and angiographic data were randomly divided into 2 sets: a derivation set and a validation set. The derivation set was used to develop a multiple logistic regression model for detecting 70% threshold carotid stenosis. Age, sex, systolic blood pressure, diastolic blood pressure, Doppler peak systolic velocity (PSV), Doppler and diastolic velocity (EDV), the systolic carotid ratio (SCR), and ophthalmic artery flow direction were tested as independent variables. A multiple linear regression model was also developed for predicting the exact degree of carotid stenosis on angiogram. The validation set was then used to evaluate the accuracy of these models. RESULTS: According to the logistic regression strategy, the best multiple logistic regression model was as follows: probability of threshold carotid stenosis = exp(2.6 PSV - 6.2)/[1 + exp(2.6 PSV - 6.2)]. The best linear regression model was as follows: degree of carotid stenosis = 20.2 PSV - 7.4 EDV + 0.4 SCR + 8.5. Both models proved to be valid following an evaluation of the validation set. CONCLUSIONS: This study illustrates that Doppler parameters may be of use in predicting the exact degree of carotid stenosis and the probability of threshold carotid stenosis. This is important if duplex criteria are going to replace angiography as the only tool for selecting endarterectomy candidates.

Measurement of carotid stenosis: comparisons between duplex and different angiographic grading methods.

BACKGROUND AND PURPOSE: Endarterectomy has been proved to be an effective stroke prevention procedure. However, there are still inconsistencies between the results of different preoperative evaluation methods, which may sometimes complicate treatment plans. This study measured the discrepancies between different angiographic grading methods and attempted to further assess the accuracy of the carotid duplex examination according to these different angiographic grading methods. METHODS: One hundred seventy-one preendarterectomy carotid duplex examinations and angiograms were reviewed. All angiograms were measured blindly by one of the authors using the North American Symptomatic Carotid Endarterectomy Trial (N), the European Carotid Surgery Trial (E), and the common carotid (C) methods. The measurement results were further converted into the area of stenosis indices (N2, E2, and C2, respectively). By using regression testing, all results could be compared. The duplex examination data were then compared with the results of different angiographic measurement methods to evaluate their accuracy. RESULTS: The measurement results of all angiographic grading methods were well correlated. Using different angiographic grading systems as the gold standard, duplex examination for screening endarterectomy candidates produced the following results: According to the N method, accuracy was 74%; according to the E method, accuracy was 90%; and according to the C method, the accuracy rate was 92%. According to N2, accuracy was 88%; according to E2, accuracy was 94%; and according to C2, the accuracy rate was 93%. CONCLUSIONS: The measurement results of all 3 commonly used angiographic grading methods were linearly correlated with one another. (It is important to note that none of those standards should be treated as the only gold standard.) In this study, the duplex criteria have a greater accuracy rate according to E or C rather than N. However, this study also demonstrated that the cut point of the Doppler criteria is the determinant factor for accuracy rather than which gold standard was compared. Through careful adjustment of the cutoff criteria, carotid duplex can be highly accurate, despite the use of different reference standards.

An eye-tracking assistive device improves the quality of life for ALS patients and reduces the caregivers' burden.

Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease. In some cases, patients with ALS retain a normal level of consciousness but disease progression eventually results in generalized paralysis, which first impedes and then prevents oral communication. This communication obstacle can generate a great deal of stress for the patient, family, and caregiver. Here the authors ask whether the use of an eye-tracking assistive device can improve quality of life for ALS patients and relieves burden of their primary caregivers. Subjects were divided into two groups depending on whether they used (n = 10) or did not use (n = 10) an eye-tracking assistive device. The authors assessed patients' quality of life and severity of depression using the ALS Specific Quality of Life Instrument-Revised and the Taiwanese Depression Questionnaire, respectively. The Caregiver Burden Scale was used to assess the burden on caregivers. Our study shows that the eye-tracking assistive device significantly improved patients' quality of life, as compared with patients in the nonuser group (p <.01). The assistive device also reduced the burden on caregivers (p <.05). This is likely a result of the improvement of patient's autonomy and more effective communication between patient and caregiver.

A novel cell-penetrating peptide derived from human eosinophil cationic protein.

Cell-penetrating peptides (CPPs) are short peptides which can carry various types of molecules into cells; however, although most CPPs rapidly penetrate cells in vitro, their in vivo tissue-targeting specificities are low. Herein, we describe cell-binding, internalization, and targeting characteristics of a newly identified 10-residue CPP, denoted ECP(32-41), derived from the core heparin-binding motif of human eosinophil cationic protein (ECP). Besides traditional emphasis on positively charged residues, the presence of cysteine and tryptophan residues was demonstrated to be essential for internalization. ECP(32-41) entered Beas-2B and wild-type CHO-K1 cells, but not CHO cells lacking of cell-surface glycosaminoglycans (GAGs), indicating that binding of ECP(32-41) to cell-surface GAGs was required for internalization. When cells were cultured with GAGs or pre-treated with GAG-digesting enzymes, significant decreases in ECP(32-41) internalization were observed, suggesting that cell-surface GAGs, especially heparan sulfate proteoglycans were necessary for ECP(32-41) attachment and penetration. Furthermore, treatment with pharmacological agents identified two forms of energy-dependent endocytosis, lipid-raft endocytosis and macropinocytosis, as the major ECP(32-41) internalization routes. ECP(32-41) was demonstrated to transport various cargoes including fluorescent chemical, fluorescent protein, and peptidomimetic drug into cultured Beas-2B cells in vitro, and targeted broncho-epithelial and intestinal villi tissues in vivo. Hence this CPP has the potential to serve as a novel vehicle for intracellular delivery of biomolecules or medicines, especially for the treatment of pulmonary or gastrointestinal diseases.