Profiling the iron, copper and zinc content in primary neuron and astrocyte cultures by rapid online quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry

Metals often determine the chemical reactivity of the proteins to which they are bound. Each cell in the body tightly maintains a unique metalloproteomic profile, mostly dependent on function. This paper describes an analytical online flow injection quantitative size exclusion chromatography-inductively coupled plasma-mass spectrometry (SEC-ICP-MS) method, which was applied to profiling the metal-binding proteins found in primary cultures of neurons and astrocytes. This method can be conducted using similar amounts of sample to those used for Western blotting (20-150 μg protein), and has a turnaround time of <15 minutes. Metalloprotein standards for Fe (as ferritin), Cu and Zn (as superoxide dismutase-1) were used to construct multi-point calibration curves for online quantification of metalloproteins by SEC-ICP-MS. Homogenates of primary neuron and astrocyte cultures were analysed by SEC-ICP-MS. Online quantification by external calibration with metalloprotein standards determined the mass of metal eluting from the column relative to time (as pg s-1). Total on-column Fe, Cu and Zn detection limits ranged from 0.825 ± 0.005 ng to 13.6 ± 0.7 pg. Neurons and astrocytes exhibited distinct metalloprotein profiles, featuring both ubiquitous and unique metalloprotein species. Separation and detection by SEC-ICP-MS allows appraisal of these metalloproteins in their native state, and online quantification was achieved using this relatively simple external calibration process. © 2013 The Royal Society of Chemistry

Targeting Nrf2 to Suppress Ferroptosis and Mitochondrial Dysfunction in Neurodegeneration

Ferroptosis is a newly described form of regulated cell death, distinct from apoptosis, necroptosis and other forms of cell death. Ferroptosis is induced by disruption of glutathione synthesis or inhibition of glutathione peroxidase 4, exacerbated by iron, and prevented by radical scavengers such as ferrostatin-1, liproxstatin-1, and endogenous vitamin E. Ferroptosis terminates with mitochondrial dysfunction and toxic lipid peroxidation. Although conclusive identification of ferroptosis in vivo is challenging, several salient and very well established features of neurodegenerative diseases are consistent with ferroptosis, including lipid peroxidation, mitochondrial disruption and iron dysregulation. Accordingly, interest in the role of ferroptosis in neurodegeneration is escalating and specific evidence is rapidly emerging. One aspect that has thus far received little attention is the antioxidant transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2). This transcription factor regulates hundreds of genes, of which many are either directly or indirectly involved in modulating ferroptosis, including metabolism of glutathione, iron and lipids, and mitochondrial function. This potentially positions Nrf2 as a key deterministic component modulating the onset and outcomes of ferroptotic stress. The minimal direct evidence currently available is consistent with this and indicates that Nrf2 may be critical for protection against ferroptosis. In contrast, abundant evidence demonstrates that enhancing Nrf2 signaling is potently neuroprotective in models of neurodegeneration, although the exact mechanism by which this is achieved is unclear. Further studies are required to determine to extent to which the neuroprotective effects of Nrf2 activation involve the prevention of ferroptosis

Medical teachers' attitudes towards science and motivational orientation for medical research

BACKGROUND: Research is an important motivating factor for pursuing a career in academic medicine, but the relation between motivation and other factors involved in scientific research are not clear. ----- PURPOSE: To explore the motivational orientation for doing research and its relation with attitudes towards science and publication practice among members of faculty at a medical school. ----- METHODS: We used a Science Attitude Survey and the Work Preference Inventory (intrinsic and extrinsic motivational orientation using 4 Likert-type scales of motivation, possible range 1-5) to survey two groups of teachers at the Zagreb University School of Medicine (n = 327, 66% response rate): professors, elected to tenure-track positions (n = 150), and instructor/research fellows working on or just completing their thesis (n = 177). RESULTS: Overall, teachers scored highest on the Enjoyment subscale of intrinsic motivational orientation (mean score +/- standard deviation 4.3 +/- 0.42 for professors vs 4.1 +/- 0.42 for instructors/research fellows, P = 0.001, t-test). Professors also scored higher than instructors/research fellows on the Challenge subscale of intrinsic motivational orientation (3.8 +/- 0.55 vs. 3.5 +/- 0.64, P < 0.001, t-test), whereas instructors/research fellows scored higher on the Compensation subscale of extrinsic motivational orientation (3.5 +/- 0.74 vs. 3.1 +/- 0.71, P < 0.001, t-test). Multiple linear regression analysis showed that the number of publications was positively associated with scores on the Science Attitude Survey and the Challenge subscale of intrinsic motivation, and negatively associated with scores on the Compensation subscale of extrinsic motivation. ----- CONCLUSIONS: Members of the medical faculty differ in motivational orientation for research depending on their academic status, and their motivation is associated more with requirements for academic advancement than with research. These findings have important implications for developing strategies for enhancing academic research production

Child dental fear and general emotional problems: a pilot study

AIM: This was to investigate the relation between general emotional and behavioural problems of the child and dental anxiety and dental behavioural management problems. BACKGROUND: Dental treatment involves many potentially unpleasant stimuli, which all may lead to the development of dental anxiety and behavioural management problems (BMP). It is still unclear why some children get anxious in the dental situation while others, with a comparable dental history, do not. Besides the latent inhibition theory it is suggested that this can be explained by differences in child rearing and personality traits. METHODS: The sample consisted of 50 children (4-12 years old) and their parents participated in this study. Parents filled out the Child Fear Survey Schedule Dental Subscale (CFSS-DS) and the Child Behaviour Checklist (CBCL) on behalf of their child. Child behaviour during consecutive dental treatments was assessed using the Venham scale. RESULTS: There were 39 children subject to analysis (21 boys) with a mean CFSS score of 40.4. Children aged 4 and 5 years who had sleeping problems, attention problems and aggressive behaviour, as scored by parents on the CBCL, displayed more disruptive behaviour during dental treatment. Children with emotionally/ reactive and attention problems were more anxious. CONCLUSION: In this pilot study a possible relation between general emotional and behavioural problems of young children and dental anxiety was shown. Also a relation between emotional and behavioural problems and dental behavioural management problems was shown. Because of the small number of subjects in our study, further research will be needed to confirm these results

Transformation of detritus by a European native and two invasive alien freshwater decapods

Invasive alien species have the potential to alter biodiversity and ecosystem processes. In freshwaters, detritus decomposition is a major ecosystem service but it remains uncertain whether invasive alien decapods process detritus differently to natives. This study examined leaf litter processing, and cascading effects on biofilms, by the European native white clawed crayfish (Austropotamobius pallipes) compared to two invasive alien decapod species: the American signal crayfish (Pacifastacus leniusculus) and the Chinese mitten crab (Eriocheir sinensis). Invasive alien decapods were responsible for higher leaf litter decomposition than the native. In comparison with native crayfish, invasive alien crab and crayfish showed higher rates of litter consumption, increased production of smaller leaf fragments, fine particulate organic matter (FPOM) and dissolved organic carbon. Nutrients (ammonia and soluble reactive phosphorous) derived from excretion (measured separately in the absence of biofilms) varied among decapod species, being lower for P. leniusculus. However, nutrient concentrations did not vary among species in the detritivory experiments with biofilm, implying nutrients were utilised for biofilm production and respiration as no differences in biomass were evident among decapod treatments. These results show invasive alien decapods have the potential to increase the magnitude of detrital processing to FPOM in rivers, but indirect impacts on primary producers due to nutrient release are uncertain based on this experimental context