Diabetes is a chronic debilitating disease that results from insufficient production of insulin from pancreatic β-cells. Islet cell replacement can effectively treat diabetes but is currently severely limited by the reliance upon cadaveric donor tissue. We have developed a protocol to efficiently differentiate commercially available human embryonic stem cells (hESCs) in vitro into a highly enriched PDX1+ pancreatic progenitor cell population that further develops in vivo to mature pancreatic endocrine cells. Immature pancreatic precursor cells were transplanted into immunodeficient mice with streptozotocin-induced diabetes, and glycemia was initially controlled with exogenous insulin. As graft-derived insulin levels increased over time, diabetic mice were weaned from exogenous insulin and human C-peptide secretion was eventually regulated by meal and glucose challenges. Similar differentiation of pancreatic precursor cells was observed after transplant in immunodeficient rats. Throughout the in vivo maturation period hESC-derived endocrine cells exhibited gene and protein expression profiles that were remarkably similar to the developing human fetal pancreas. Our findings support the feasibility of using differentiated hESCs as an alternative to cadaveric islets for treating patients with diabetes.
This cross-cultural study was designed to examine cultural differences in empathy levels of first-year medical students.
A total of 257 students from the academic year 2010/11, 131 at Jimma University, Ethiopia, and 126 at the Ludwig Maximilian University, Munich, Germany, completed the Balanced Emotional Empathy Scale (BEES), the Reading the Mind in the Eyes (RME-R) test, and a questionnaire on sociodemographic and cultural characteristics. Furthermore, we conducted a qualitative analysis of the students' personal views on the definition of empathy and possible influencing factors. Group comparisons and correlation analyses of empathy scores were performed for the entire cohort and for the Jimma and Munich students separately. We used a regression tree analysis to identify factors influencing the BEES.
The male students in Jimma (39.1 ± 22.3) scored significantly higher in the BEES than those male students from Munich (27.2 ± 22.6; p = 0.0002). There was no significant difference between the female groups. We found a moderate, positive correlation between the BEES and RME-R test, i.e. between emotional and cognitive empathy, within each university. Nevertheless, the RME-R test, which shows only Caucasian eyes, appears not to be suitable for use in other cultures.
The main findings of our study were the influence of culture, religion, specialization choice, and gender on emotional empathy (assessed with the BEES) and cognitive empathy (assessed with the RME-R test) in first-year medical students. Further research is required into the nature of empathy in worldwide medical curricula.
Balanced Emotional Empathy Scale; Cross-cultural; Empathy; Medical Students
We investigated whether a cell-penetrating peptide linked via a disulfide bond to a fluorophore-labeled
cargo peptide can be used to interrogate changes in cellular redox state. A fluorescence resonance energy
transfer (FRET) pair was constructed so that the cargo peptide was labeled with fluorescein amidite (FAM)
and the cell-penetrating peptide was attached to a quencher. Incubation of cells in culture with the FRET
construct was visualized using live-cell, time-lapse imaging, which demonstrated earlier cellular uptake of
the construct when cells were treated with the reducing agent n-acetylcysteine (NAC). The FRET peptide
construct was easily detected in cells cultured in 96-well plates using a plate-reader. Treatment of cells with
various classes of reducing or oxidizing agents resulted in an increase or decrease in FAM fluorescence,
respectively. Changes in FAM fluorescence correlated significantly with redox-sensitive green fluorescent
protein ratios in cells treated with hydrogen peroxide but not NAC. Detection of relative changes in cellular
redox state was enhanced by the fact that uptake of the cell-penetrating peptide occurred more quickly in
relatively reduced compared with oxidized cells. We conclude that cell-penetrating peptides coupled via
disulfide bonds to detectable cargo is a novel and specific approach for assessment of relative changes in
cellular thiol redox state.
Redox; cell-penetrating peptides; model amphipathic peptide; glutathione; FRET.
Antipsychotics, cognitive behavioral therapy (CBT), and omega-3-fatty acids have been found superior to control conditions as regards prevention of psychosis in people at-risk of first-episode psychosis. However, no large-scale trial evaluating the differential efficacy of CBT and antipsychotics has been performed yet. In PREVENT, we evaluate CBT, aripiprazole, and clinical management (CM) as well as placebo and CM for the prevention of psychosis in a randomized, double-blind, placebo-controlled trial with regard to the antipsychotic intervention and a randomized controlled trial with regard to the CBT intervention with blinded ratings. The hypotheses are first that CBT and aripiprazole and CM are superior to placebo and CM and second that CBT is not inferior to aripiprazole and CM combined. The primary outcome is transition to psychosis. By November 2010, 156 patients were recruited into the trial. The subjects were substantially functionally compromised (Social and Occupational Functioning Assessment Scale mean score 52.5) and 78.3% presented with a Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition axis I comorbid diagnosis. Prior to randomization, 51.5% of the participants preferred to be randomized into the CBT arm, whereas only 12.9% preferred pharmacological treatment. First, assessments of audiotaped treatment sessions confirmed the application of CBT-specific skills in the CBT condition and the absence of those in CM. The overall quality rating of the CBT techniques applied in the CBT condition was good. When the final results of the trial are available, PREVENT will substantially expand the current limited evidence base for best clinical practice in people at-risk (prodromal) of first-episode psychosis.
schizophrenia; aripiprazole; cognitive behavior therapy; prodrome; early intervention; prevention; psychosis
First episode schizophrenia (FES) patients tend to be more responsive to treatment. An adequate response has been associated with a favourable long-term course in FES patients. Yet, despite the generally very favourable response profile around one quarter of the patients shows persisting symptoms of psychosis. To improve the outcome and course of psychosis great effort has emerged in identifying biological and clinical variables associated with non-response in order to identify non-responders as early as possible and adopt specific treatment strategies improving illness outcome. Different antipsychotic treatment regimens have been evaluated in terms of their efficacy in reducing symptoms of FES with psychological interventions gaining increasing importance in the treatment concept of patients suffering from their first illness episode. Therefore, aim of this review is to summarize current evidence on the response patterns, the most important predictors of response/non-response as well as on effective treatment interventions in FES patients.
First-episode schizophrenia; Response; Course of illness; Outcome; Predictors; Treatment interventions
Randomized, double-blind, placebo-controlled trials constitute the gold standard in clinical research when testing the efficacy of new psychopharmacological interventions in the treatment of major depression. However, the blinded use of placebo has been found to influence clinical trial outcomes and may bias patient selection.
To improve clinical trial design in major depression so as to reflect clinical practice more closely we propose to present patients with a balanced view of the benefits of study participation irrespective of their assignment to placebo or active treatment. In addition every participant should be given the option to finally receive the active medication. A research agenda is outlined to evaluate the impact of the proposed changes on the efficacy of the drug to be evaluated and on the demographic and clinical characteristics of the enrollment fraction with regard to its representativeness of the eligible population.
We propose a list of measures to be taken to improve the external validity of double-blind, placebo-controlled trials in major depression. The recommended changes to clinical trial design may also be relevant for other psychiatric as well as medical disorders in which expectations regarding treatment outcome may affect the outcome itself.
major depression; clinical trials; randomized controlled trials; psychopharmacology
Impairments in neuropsychological functioning have been described in subjects clinically at high risk for psychosis, but the specific cognitive deficits in different clinical high-risk groups remain to be elucidated. The German Research Network on Schizophrenia employs a heuristic 2-stage model: a putatively late prodromal state (LPS), characterized by the onset of attenuated positive or brief psychotic symptoms, and an early prodromal state (EPS), mainly characterized by the presence of basic symptoms, which are predictive for psychosis within the next 10 years.
A total of 205 subjects met the criteria for either an EPS or an LPS of psychosis and were assessed with a comprehensive neuropsychological test battery. Neurocognitive profiles of high-risk groups were compared with data of 87 healthy controls comparable with regard to gender, age, and premorbid verbal IQ.
Patients in the LPS were impaired in all neurocognitive domains (memory/learning, executive control/processing speed, and working memory) examined, with memory being the worst. Deficits were less pronounced in patients in the EPS, with a specific deficit in the executive control/processing speed domain. Consistent with a progressive neurodevelopmental disorder, some cognitive abilities were already impaired in patients in the EPS, followed by further deterioration in the LPS. Specifically, deficits in executive control functioning were related to the presence of basic symptoms, indicating a vulnerability for psychosis. Memory deficits were associated with the onset of psychotic symptoms indicating further disease progression in the trajectory to psychosis and, thus, may be useful in predicting psychosis and targeting early intervention.
clinical high risk; cognition; neuropsychology; memory; executive function
Infections and immunological processes are likely to be involved in the pathogenesis of Tourette’s syndrome (TS). To determine possible common underlying immunological mechanisms, we focused on innate immunity and studied markers of inflammation, monocytes, and monocyte-derived cytokines.
In a cross-sectional study, we used current methods to determine the number of monocytes and levels of C-reactive protein (CRP) in 46 children, adolescents, and adult patients suffering from TS and in 43 healthy controls matched for age and sex. Tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL-6), soluble CD14 (sCD14), IL1-receptor antagonist (IL1-ra), and serum neopterin were detected by immunoassays.
We found that CRP and neopterin levels and the number of monocytes were significantly higher in TS patients than in healthy controls. Serum concentrations of TNF-alpha, sIL1-ra, and sCD14 were significantly lower in TS patients. All measured values were within normal ranges and often close to detection limits.
The present results point to a monocyte dysregulation in TS. This possible dysbalance in innate immunity could predispose to infections or autoimmune reactions.
Differentiation of human embryonic stem (hES) cells to fully developed cell types holds great therapeutic promise. Despite significant progress, the conversion of hES cells to stable, fully differentiated endocrine cells that exhibit physiologically regulated hormone secretion has not yet been achieved. Here we describe an efficient differentiation protocol for the in vitro conversion of hES cells to functional glucagon-producing α- cells.
RESEARCH DESIGN AND METHODS
Using a combination of small molecule screening and empirical testing, we developed a six-stage differentiation protocol for creating functional α-cells. An extensive in vitro and in vivo characterization of the differentiated cells was performed.
A high rate of synaptophysin expression (>75%) and robust expression of glucagon and the α-cell transcription factor ARX was achieved. After a transient polyhormonal state in which cells coexpress glucagon and insulin, maturation in vitro or in vivo resulted in depletion of insulin and other β-cell markers with concomitant enrichment of α-cell markers. After transplantation, these cells secreted fully processed, biologically active glucagon in response to physiologic stimuli including prolonged fasting and amino acid challenge. Moreover, glucagon release from transplanted cells was sufficient to reduce demand for pancreatic glucagon, resulting in a significant decrease in pancreatic α-cell mass.
These results indicate that fully differentiated pancreatic endocrine cells can be created via stepwise differentiation of hES cells. These cells may serve as a useful screening tool for the identification of compounds that modulate glucagon secretion as well as those that promote the transdifferentiation of α-cells to β-cells.
The purpose of the current study was to determine whether double-stranded adeno-associated virus (dsAAV)-mediated in vivo expression of β-cell growth factors, glucagon-like peptide-1 (GLP-1) and the NK1 fragment of hepatocyte growth factor (HGF/NK1) in β-cells, improves pathology in the db/db mouse model of type 2 diabetes.
RESEARCH DESIGN AND METHODS
The glucoregulatory actions of GLP-1 and full-length HGF are well characterized. Here, we test the ability of HGF/NK1 to induce proliferation of exogenous islets and MIN6 β-cells. In addition, we target both GLP-1 and HGF/NK1 to endogenous β-cells using dsAAV vectors containing the mouse insulin-II promoter. We compare the abilities of these gene products to induce islet proliferation in vitro and in vivo and characterize their abilities to regulate diabetes after AAV-mediated delivery to endogenous islets of db/db mice.
Recombinant HGF/NK1 induces proliferation of isolated islets, and dsAAV-mediated expression of both GLP-1 and HGF/NK1 induces significant β-cell proliferation in vivo. Furthermore, both GLP-1 and HGF/NK1 expressed from dsAAV vectors enhance β-cell mass and insulin secretion in vivo and significantly delay the onset of hyperglycemia in db/db mice.
A single treatment with dsAAV vectors expressing GLP-1 or HGF/NK1 enhances islet growth and significantly improves pathology in a mouse model of type 2 diabetes. This represents the first example of a successful use of HGF/NK1 for diabetes therapy, providing support for direct AAV-mediated in vivo delivery of β-cell growth factors as a novel therapeutic strategy for the treatment of type 2 diabetes.
The PANSS (Positive and Negative Syndrome Scale) is one of the most important rating instruments for patients with schizophrenia. Nevertheless, there is a long and ongoing debate in the psychiatric community regarding its mathematical properties.
All 30 items range from 1 to 7 leading to a minimum total score of 30, implying that the PANSS is an interval scale. For such interval scales straightforward calculation of relative changes is not appropriate. To calculate outcome criteria based on a percent change as, e.g., the widely accepted response criterion, the scale has to be transformed into a ratio scale beforehand. Recent publications have already pointed out the pitfall that ignoring the scale level (interval vs. ratio scale) leads to a set of mathematical problems, potentially resulting in erroneous results concerning the efficacy of the treatment.
A Pubmed search based on the PRISMA statement of the highest-ranked psychiatric journals (search terms "PANSS" and "response") was carried out. All articles containing percent changes were included and methods of percent change calculation were analysed.
This systematic literature research shows that the majority of authors (62%) actually appear to use incorrect calculations. In most instances the method of calculation was not described in the manuscript.
These alarming results underline the need for standardized procedures for PANSS calculations.
PANSS; scale level; literature search
The sodium-calcium exchanger isoform 1 (NCX1) regulates cytoplasmic calcium (Ca2+c) required for insulin secretion in β-cells. NCX1 is alternatively spliced, resulting in the expression of splice variants in different tissues such as NCX1.3 and -1.7 in β-cells. As pharmacological inhibitors of NCX1 splice variants are in development, the pharmacological profile of β-cell NCX1.3 and -1.7 and the cellular effects of NCX1 inhibition were investigated.
RESEARCH DESIGN AND METHODS
The patch-clamp technique was used to examine the pharmacological profile of the NCX1 inhibitor KB-R7943 on recombinant NCX1.3 and -1.7 activity. Ca2+ imaging and membrane capacitance were used to assess the effects of KB-R7943 on Ca2+c and insulin secretion in mouse and human β-cells and islets.
NCX1.3 and -1.7 calcium extrusion (forward-mode) activity was ∼16-fold more sensitive to KB-R7943 inhibition compared with cardiac NCX1.1 (IC50s = 2.9 and 2.4 vs. 43.0 μmol/l, respectively). In single mouse/human β-cells, 1 μmol/l KB-R7943 increased insulin granule exocytosis but was without effect on α-cell glucagon granule exocytosis. KB-R7943 also augmented sulfonylurea and glucose-stimulated Ca2+c levels and insulin secretion in mouse and human islets, although KB-R7943 was without effect under nonstimulated conditions.
Islet NCX1 splice variants display a markedly greater sensitivity to pharmacological inhibition than the cardiac NCX1.1 splice variant. NCX1 inhibition resulted in glucose-dependent increases in Ca2+c and insulin secretion in mouse and human islets. Thus, we identify β-cell NCX1 splice variants as targets for the development of novel glucose-sensitive insulinotropic drugs for type 2 diabetes.
The design of the Positive and Negative Syndrome Scale (PANSS) with item levels ranging from 1 to 7 leads to the trivial result that the 30-item scale’s zero level (no symptoms) is 30. This causes serious problems when ratios are calculated which always implicitly depend on a natural zero point (equals 0). Recent publications concerning efficacy of antipsychotics correctly suggest a subtraction of 30 points to every PANSS before calculating percent change (PC). Nevertheless, the traditional approach using uncorrected scores is still in common practice. This analysis aims to clarify which approach is the most appropriate from a statistical perspective.For analysis, data from a naturalistic study on 400 patients with a schizophrenic spectrum disorder and simulated data sets were used. While calculations concerning absolute score values and their differences are not affected, considerable problems arise in calculations of PC and related response criteria. Even significance levels of estimated treatment effects change, depending on the structure of the data (eg, baseline symptom severity). Using a PANSS version with items ranging from 0 to 6 would avoid such often neglected pitfalls.
scale level; minimum subtraction; percent change; simulation study
Diabetes is a devastating disease that is ultimately caused by the malfunction or loss of insulin-producing pancreatic beta-cells. Drugs capable of inducing the development of new beta-cells or improving the function or survival of existing beta-cells could conceivably cure this disease. We report a novel high-throughput screening platform that exploits multi-parameter high-content analysis to determine the effect of compounds on beta-cell survival, as well as the promoter activity of two key beta-cell genes, insulin and pdx1. Dispersed human pancreatic islets and MIN6 beta-cells were infected with a dual reporter lentivirus containing both eGFP driven by the insulin promoter and mRFP driven by the pdx1 promoter. B-score statistical transformation was used to correct systemic row and column biases. Using this approach and 5 replicate screens, we identified 7 extracts that reproducibly changed insulin and/or pdx1 promoter activity from a library of 1319 marine invertebrate extracts. The ability of compounds purified from these extracts to significantly modulate insulin mRNA levels was confirmed with real-time PCR. Insulin secretion was analyzed by RIA. Follow-up studies focused on two lead compounds, one that stimulates insulin gene expression and one that inhibits insulin gene expression. Thus, we demonstrate that multi-parameter, high-content screening can identify novel regulators of beta-cell gene expression, such as bivittoside D. This work represents an important step towards the development of drugs to increase insulin expression in diabetes and during in vitro differentiation of beta-cell replacements.
The accumulation and aggregation of α-synuclein in nerve cells and glia are characteristic features of a number of neurodegenerative diseases termed synucleinopathies. α-Synuclein is a highly soluble protein which in a nucleation dependent process is capable of self-aggregation. The causes underlying aggregate formation are not yet understood, impairment of the proteolytic degradation systems might be involved.
In the present study the possible aggregate clearing effects of the geldanamycin analogue 17-AAG (17-(Allylamino)-17-demethoxygeldanamycin) was investigated. Towards this, an oligodendroglial cell line (OLN-93 cells), stably expressing human α-synuclein (A53T mutation) was used. In these cells small punctate aggregates, not staining with thioflavine S, representing prefibrillary aggregates, occur characteristically. Our data demonstrate that 17-AAG attenuated the formation of α-synuclein aggregates by stimulating macroautophagy. By blocking the lysosomal compartment with NH4Cl the aggregate clearing effects of 17-AAG were abolished and α-synuclein deposits were enlarged. Analysis of LC3-II immunoreactivity, which is an indicator of autophagosome formation, further revealed that 17-AAG led to the recruitment of LC3-II and to the formation of LC3 positive puncta. This effect was also observed in cultured oligodendrocytes derived from the brains of newborn rats. Inhibition of macroautophagy by 3-methyladenine prevented 17-AAG induced occurrence of LC3 positive puncta as well as the removal of α-synuclein aggregates in OLN-A53T cells.
Our data demonstrate for the first time that 17-AAG not only causes the upregulation of heat shock proteins, but also is an effective inducer of the autophagic pathway by which α-synuclein can be removed. Hence geldanamycin derivatives may provide a means to modulate autophagy in neural cells, thereby ameliorating pathogenic aggregate formation and protecting the cells during disease and aging.
Quetiapine was developed in 1985 by scientists at AstraZeneca (formerly Zeneca) Pharmaceuticals. It received official US Food and Drug Administration approval in September 1997 and approval in Germany in 2000. Since then, quetiapine has been used in the treatment of severe mental illness in approximately 70 countries including Canada, most Western European countries, and Japan. Quetiapine is a dibenzothiazepine derivative with a relatively broad receptor binding profile. It has major affinity to cerebral serotonergic (5HT2A), histaminergic (H1), and dopaminergic D1 and D2 receptors, moderate affinity to α1- und α2-adrenergic receptors, and minor affinity to muscarinergic M1 receptors; it demonstrates a substantial selectivity for the limbic system. This receptor occupancy profile with relatively higher affinity for the 5HT2A receptor compared with the D2 receptor is in part responsible for the antipsychotic characteristics and low incidence of extrapyramidal side-effects of quetiapine. The efficacy of quetiapine in reducing positive and negative symptoms of schizophrenia has been proven in several clinical trials with placebo-controlled comparators. Quetiapine has also demonstrated robust efficacy for treatment of cognitive, anxious-depressive, and aggressive symptoms in schizophrenia. Long-term trials show sustained tolerability for a broad spectrum of symptoms. Quetiapine has also proven efficacy and tolerability in the treatment of moderate to severe manic episodes, and in the treatment of juveniles with oppositional-defiant or conduct disorders, and in the geriatric dementia population. Recent data indicate that quetiapine may also be effective in the treatment of bipolar depressive symptoms without increasing the risk of triggering manic episodes, and in borderline personality disorder. In comparison with other antipsychotics, quetiapine has a favorable side-effect profile. In clinical trials only small insignificant prolongations of the QT interval were observed. Weight-gain liabilities and new-onset metabolic side-effects occupy a middle-ground among newer antipsychotics. As a result of its good efficacy and tolerability profile quetiapine has become well established in the treatment of schizophrenia and manic episodes.
schizophrenia; antipsychotic; quetiapine; efficacy; tolerability
The aim was to investigate the hypothesis that patients with first episode schizophrenic disorders have a more favorable treatment response than those with multiple episodes.
A total of 400 inpatients from an ongoing multi-centre, follow-up program who fulfilled ICD-10 criteria for schizophrenic disorders (F2) were assessed at admission to and discharge from hospital using the Positive and Negative Syndrome Scale (PANSS).
At admission, first episode patients (n = 121) showed higher levels of positive symptoms (PANSS positive subscore) and lower ones of negative symptoms (PANSS negative subscore) than multiple episode patients (n = 279), whereas the global disease severity (PANSS total score) was comparable. Analyses of covariance revealed that treatment response (adjusted symptom levels at discharge) was more favorable in first-episode patients, with respect to both positive and negative symptoms.
The results are compatible with the hypothesis that treatment response becomes less favorable during the course of schizophrenic illness. This finding might be associated with progressive neurobiological alterations.
schizophrenic disorders; first episode; multiple episode; treatment response; PANSS