Glycans are not necessary to maintain the pathobiological features of bovine spongiform encephalopathy.

The role of the glycosylation status of PrPC in the conversion to its pathological counterpart and on cross-species transmission of prion strains has been widely discussed. Here, we assessed the effect on strain characteristics of bovine spongiform encephalopathy (BSE) isolates with different transmission histories upon propagation on a model expressing a non-glycosylated human PrPC. Bovine, ovine and porcine-passaged BSE, and variant Creutzfeldt-Jakob disease (vCJD) isolates were used as seeds/inocula in both in vitro and in vivo propagation assays using the non-glycosylated human PrPC-expressing mouse model (TgNN6h). After protein misfolding cyclic amplification (PMCA), all isolates maintained the biochemical characteristics of BSE. On bioassay, all PMCA-propagated BSE prions were readily transmitted to TgNN6h mice, in agreement with our previous in vitro results. TgNN6h mice reproduced the characteristic neuropathological and biochemical hallmarks of BSE, suggesting that the absence of glycans did not alter the pathobiological features of BSE prions. Moreover, back-passage of TgNN6h-adapted BSE prions to BoTg110 mice recovered the full BSE phenotype, confirming that the glycosylation of human PrPC is not essential for the preservation of the human transmission barrier for BSE prions or for the maintenance of BSE strain properties.

Physical activity, weight and functional limitations in elderly Spanish people: the National Health Survey (2009-2014).

Background: The purpose of this study was to analyze physical activity (PA), functional limitations, weight status, self-perceived health status and disease or chronic health problems in older people aged 65 and over using the European Health Survey in Spain (EHSS) conducted one in 2009 and one in 2014. Methods: This study included 12,546 older people, 6026 [2330 men and 3696 women; age (Mean, SD) =75.61 ± 7.11 years old] in 2009 and 6520 [2624 men and 3896 women; age (Mean, SD) =75.90 ± 7.59 years old] in 2014. The sample was divided into three age groups: 65-74 years old, 75-84 years old and ≥85 years old. Results: In 2014, participants exhibited lower values for moderate PA, and self-perceived health status compared to 2009. Moreover, in 2014 more people with disease or chronic health problems, and severe difficulty walking 500 m without assistance were found and severe difficulty going up and down 12 stairs than people in 2009. In relation to weight status there were no significant differences between older people in 2009 and 2014. Conclusions: From 2009 to 2014, the PA levels of Spanish older people have decreased, while the BMI has not increased. That fact is in consonance with a worst perception of health status in 2014 and with an increase of their disease levels. The current data highlight the importance of incorporating exercise programmes at an early stage of ageing in order to preserve physical performance, and to prevent the negative consequences of ageing.

Prevalence and determinants in cytology testing for cervical cancer screening in Spain (2006-14).

Background: Cervical cancer has decreased in developed countries thanks to cytology screening programmes. The aims of this study were To analyse the frequency and evolution of performing cytology tests and to determine the variables that influence their use. Methods: Cross-sectional study of non-institutionalized women who participated in the national health survey (2006, 2011/12) and the European Health Survey in Spain (2009, 2014). Study variables: cytology-testing, time since last cytology-test, reason for performing the test, age, nationality, marital status, social status, education level and place of residence. Results: The study evaluated 53 628 women in Spain over 15 years old, with a mean age of 52.68 (SD ± 19.12). About 94.1% were Spanish, 49.2% were married and 77.2% lived with a partner. In 2014, 72% had a cytology test, a number that increased significantly. Women aged 25-65 were 5.13 times more likely to undergo a cytology test than those aged 15-24 years old (odds ratio (OR): 5.13; P < 0.001); women with university educations were 9.23 times more likely to undergo a cytology test than those without university educations (P > 0.001); those of social classes I and II (high) were 1.2 more likely to undergo a cytology test than those of low social class (P = 0.026); and Spanish women were 1.74 times more likely to undergo a cytology test than foreigners living in Spain (P < 0.001). Conclusion: Frequency of cytology testing has increased in the last few years. Screening for cervical cancer is associated with higher social status, education level, age, and not being foreign.

PrP(C) regulates epidermal growth factor receptor function and cell shape dynamics in Neuro2a cells.

The prion protein (PrP) plays a key role in prion disease pathogenesis. Although the misfolded and pathologic variant of this protein (PrP(SC)) has been studied in depth, the physiological role of PrP(C) remains elusive and controversial. PrP(C) is a cell-surface glycoprotein involved in multiple cellular functions at the plasma membrane, where it interacts with a myriad of partners and regulates several intracellular signal transduction cascades. However, little is known about the gene expression changes modulated by PrP(C) in animals and in cellular models. In this article, we present PrP(C)-dependent gene expression signature in N2a cells and its implication in the most overrepresented functions: cell cycle, cell growth and proliferation, and maintenance of cell shape. PrP(C) over-expression enhances cell proliferation and cell cycle re-entrance after serum stimulation, while PrP(C) silencing slows down cell cycle progression. In addition, MAP kinase and protein kinase B (AKT) pathway activation are under the regulation of PrP(C) in asynchronous cells and following mitogenic stimulation. These effects are due in part to the modulation of epidermal growth factor receptor (EGFR) by PrP(C) in the plasma membrane, where the two proteins interact in a multimeric complex. We also describe how PrP(C) over-expression modulates filopodia formation by Rho GTPase regulation mainly in an AKT-Cdc42-N-WASP-dependent pathway.

Nomophobia and the influence of time to REST among nursing students. A descriptive, correlational and predictive research.

The inclusion of the smartphone in society has brought many advantages, but also disadvantages, such as nomophobia, considered as a digital disease generated by the excessive use of the smartphone. The general objective of the research is to know and analyse the prevalence of nomophobia among nursing students and knowing the influence of time to rest. The research design is descriptive, correlational, transversal and predictive with a quantitative research methodology. The sample consist of nursing students from the University of Granada (N = 880) in Spain. A descriptive analysis has been carried out, as well as a bivariate correlation of Pearson, the student T test and a multiple linear regression. The results show an average level of nomophobia among nursing students. Although the students can recognize that the use of the smartphone can reduce their rest period, mainly focused on the hours they spend sleeping, there is no significant relationship between the two facts, confirming also that the levels of nomophobia are significantly more related in those who claim not to have problems in their rest due to the use of the mobile phone. A large percentage of them claim to rest less time due to excessive use of their smartphone.

Genome comparison of a nonpathogenic myxoma virus field strain with its ancestor, the virulent Lausanne strain.

One of the best-studied examples of host-virus coevolution is the release of myxoma virus (MV) for biological control of European rabbits in Australia and Europe. To investigate the genetic basis of MV adaptation to its new host, we sequenced the genome of 6918, an attenuated Spanish field strain, and compared it with that of Lausanne, the strain originally released in Europe in 1952. Although isolated 43 years apart, the genomes were highly conserved (99.95% identical). Only 32 of the 159 MV predicted proteins revealed amino acid changes. Four genes (M009L, M036L, M135R, and M148R) in 6918 were disrupted by frameshift mutations.

Amino acids and acylcarnitine production by Chlorella vulgaris and Chlorella sorokiniana microalgae from wastewater culture.

BACKGROUND: Microalgae are a widely distributed group of prokaryotic and eukaryotic photosynthetic microorganisms that use a number of substances present in wastewater to produce a variety of biotechnological and nutritional biomolecules. METHODS: Production ofamino acids and acylcarnitine by Chlorella vulgaris and Chlorella sorokiniana was determined after 13 d of culture in wastewater, under various culture conditions. Wastewater was collected from "La Encantada" stream, located in Saltillo, Coahuila, Mexico. Microalgae was cultured at 23°C and natural day light, including the use of the following conditions: (1) extra light (12:12 light:dark cycles, 1,380 lumens), (2) agitation (130 rpm), and (3) both conditions, until exponential phase. Supernatant products were then analyzed by liquid chromatograph coupled to mass spectrometry. In addition, metabolomic profiles related to growing conditions were evaluated. RESULTS: Amino acids and acylcarnitine production by C. sorokiniana and C. vulgaris resulted in higher Ala and Leu concentrations by C. vulgaris compared with control, where control produced Gly and Pro in higher amounts compared with C. sorokiniana. Tyr, Phe, Val, and Cit were detected in lower amounts under light and shaking culture conditions. High concentrations of C0 acylcarnitines were produced by both microalgae compared with control, where C. sorokiniana production was independent of culture conditions, whereas C. vulgaris one was stimulated by shaking. C4 production was higher by C. sorokiniana compared with control. Furthermore, C4, C6DC, C14:1, C14:2, and C18:1OH production by microalga was low in all culture conditions. CONCLUSION: Microalgae produced essential amino acids and nutritionally important carnitines from wastewater. In addition, C. sorokiniana biomass has higher potential as animal nutrient supplement, as compared with that of C. vulgaris.

Enhanced Error Decoding from Error-Related Potentials using Convolutional Neural Networks.

Error-related potentials are considered an important neuro-correlate for monitoring human intentionality in decision-making, human-human, or human-machine interaction scenarios. Multiple methods have been proposed in order to improve the recognition of human intentions. Moreover, current brain-computer interfaces are limited in the identification of human errors by manual tuning of parameters (e.g., feature/channel selection), thus selecting fronto-central channels as discriminative features within-subject. In this paper, we propose the inclusion of error-related potential activity as a generalized two-dimensional feature set and a Convolutional Neural Network for classification of EEG-based human error detection. We evaluate this pipeline using the BNCI2020 - Monitoring Error-Related Potential dataset obtaining a maximum error detection accuracy of 79.8% in a within-session 10-fold cross-validation modality, and outperforming current state of the art.

Placebo-controlled crossover assessment of mecasermin for the treatment of Rett syndrome.

Objective: To measure the efficacy of mecasermin (recombinant human insulin-like growth factor 1, rhIGF-1), for treating symptoms of Rett syndrome (RTT) in a pediatric population using a double-blind crossover study design. Methods: Thirty girls with classic RTT in postregression stage were randomly assigned to placebo or rhIGF-1 in treatment period 1 and crossed over to the opposite assignment for period 2 (both 20 weeks), separated by a 28-week washout period. The primary endpoints were as follows: Anxiety Depression and Mood Scale (ADAMS) Social Avoidance subscale, Rett Syndrome Behaviour Questionnaire (RSBQ) Fear/Anxiety subscale, Parent Target Symptom Visual Analog Scale (PTSVAS) top three concerns, Clinical Global Impression (CGI), Parent Global Impression (PGI), and the Kerr severity scale. Cardiorespiratory- and electroencephalography (EEG)-based biomarkers were also analyzed. Results: There were no significant differences between randomization groups. The majority of AEs were mild to moderate, although 12 episodes of serious AEs occurred. The Kerr severity scale, ADAMS Depressed Mood subscale, Visual Analog Scale Hyperventilation, and delta average power change scores significantly increased, implying worsening of symptoms. Electroencephalography (EEG) parameters also deteriorated. A secondary analysis of subjects who were not involved in a placebo recall confirmed most of these findings. However, it also revealed improvements on a measure of stereotypic behavior and another of social communication. Interpretation: As in the phase 1 trial, rhIGF-1 was safe; however, the drug did not reveal significant improvement, and some parameters worsened.

Coenzyme Q and protein/lipid oxidation in a BSE-infected transgenic mouse model.

Oxidative stress and antioxidants play an important role in neurodegenerative diseases. However, the exact participation of antioxidants in the evolution of prion diseases is still largely unknown. The aim of this study was to assess brain levels of coenzyme Q (CoQ), an endogenous lipophilic antioxidant, and the antioxidant/pro-oxidant status by determining oxidative damage to proteins and lipids after intracerebral bovine spongiform encephalopathy (BSE) infection of transgenic mice expressing bovine prion protein (PrP). Our results indicate that, whereas the ratio between the two CoQ homologues present in mice (CoQ(9) and CoQ(10)) is not altered by prion infection during the course of the disease, significant increases in total CoQ(9) and CoQ(10) were observed in BSE-infected mice 150 days after inoculation. This time point coincided with the first manifestation of PrP(Sc) deposition in nervous tissue. In addition, CoQ(9) and CoQ(10) levels, neuropathological alterations, and PrP(Sc) deposition in nervous tissues underwent further increases as the illness progressed. Lipid and protein oxidation were observed only at the final stage of the disease after clinical signs had appeared. These findings indicate upregulation of CoQ(9)- and CoQ(10)-dependent antioxidant systems in response to the increased oxidative stress induced by prion infection in nervous tissue. However, the induction of these endogenous antioxidant systems seems to be insufficient to prevent the development of the illness.

Transmission and Replication of Prions.

Transmissible spongiform encephalopathies (TSEs) are a group of progressive, invariably fatal diseases that affect the nervous system of many mammals including humans. The key molecular event in the pathogenesis of TSEs is the conversion of the cellular prion protein PrPC into a disease-associated isoform PrPSc. The "protein-only hypothesis" argues that PrPSc itself is the infectious agent. In effect, PrPSc can adopt several structures that represent different prion strains. The interspecies transmission of TSEs is difficult because of differences between the host and donor primary PrP sequence. However, transmission is not impossible as this occurred when bovine spongiform encephalopathy spread to humans causing variant Creutzfeldt-Jakob disease (vCJD). This event determined a need for a thorough understanding of prion replication and transmission so that we could be one step ahead of further threats for human health. This chapter focuses on these concepts and on new insights gained into prion propagation mechanisms.

Cell expression of a four extra octarepeat mutated PrPC modifies cell structure and cell cycle regulation.

RK13 cell lines generated to express bovine PrP(C) with a four extra octarepeat insertional mutation (Bo-10ORPrP(C)) show partially insoluble PrP(C) and lower rates of cell growth when compared to either the same cells expressing wild type Bo-6ORPrP(C) or the original RK13 cell line. The expression of Bo-10ORPrP(C) in cell cultures was also associated with changes in cell size and reorganization of the actin cytoskeleton. This last process was reversed by Clostridium difficile toxin-B, a specific inhibitor of small GTPase proteins. Further, in clones expressing Bo-10ORPrP(C), increased proportions of cells at cell cycle stage G2/M were observed. Proteasome inhibitors caused a further expansion of G2/M-stage cells that was more marked in cell lines expressing Bo-10ORPrP(C) than those expressing Bo-6ORPrP(C), while this effect was minimal or null in the original RK13 cell line. Hence, the presence of Bo-10ORPrP(C) in RK13 cells promotes cell cycle arrest at G2/M, and the effect is amplified by proteasome inhibition. These findings suggest a role for PrP(C) in cell morphology and cell cycle regulation, and open new avenues for understanding the mechanisms underlying PrP mutation-associated diseases.

Comparison of three monoclonal antibodies for use in immunohistochemical detection of bovine spongiform encephalopathy protease-resistant prion protein.

Confirmatory diagnosis of prion diseases in humans and animals relies on the histopathological examination and immunodetection of the protease-resistant isoform of prion protein (PrPres). The generation of novel PrP-specific monoclonal antibodies (MAbs) has greatly improved diagnostic methodology and basic research on prion diseases as well. In this study, the performance of 3 different PrP-specific MAbs in recognizing brain PrPres deposits from cows affected with bovine spongiform encephalopathy (BSE) was compared by using a standard immunohistochemical technique under different pretreatment conditions. All antibodies showed similar reactivity after denaturing treatment. However, greater differences were found among them after proteinase K treatment, even in the absence of a denaturing step. In fact, 1 MAb (2A11) was able to react with PrPres deposits in the absence of a denaturing step, yielding the strongest signal and confirming the usefulness of MAb 2A11 in immunohistochemistry for the diagnosis of BSE.

Subclinical bovine spongiform encephalopathy infection in transgenic mice expressing porcine prion protein.

The bovine-porcine species barrier to bovine spongiform encephalopathy (BSE) infection was explored by generating transgenic mouse lines expressing the porcine prion protein (PrP) gene. All of the porcine transgenic (poTg) mice showed clinical signs of BSE after intracerebral inoculation with a high-titer BSE inoculum. The protease-resistant PrP (PrP(res)) was detected in 14% (3 of 22) of the BSE-infected poTg mice by immunohistochemical or immunoblot analysis. Despite being able to infect 42% (5 of 12) of control mice, a low-dose BSE inoculum failed to penetrate the species barrier in our poTg mouse model. The findings of these infectivity studies suggest that there is a strong species barrier between cows and pigs. However, after second-passage infection of poTg mice using brain homogenates of BSE-inoculated mice scoring negative for the incoming prion protein as inoculum, it was possible to detect the presence of the infectious agent. Thus, porcine-adapted BSE inocula were efficient at infecting poTg mice, giving rise to an incubation period substantially reduced from 300 to 177 d after inoculation and to the presence of PrP(res) in 100% (21 of 21) of the mice. We were therefore able to conclude that initial exposure to the bovine prion may lead to subclinical infection such that brain homogenates from poTg mice classified as uninfected on the basis of the absence of PrP(res) are infectious when used to reinoculate poTg mice. Collectively, our findings suggest that these poTg mice could be used as a sensitive bioassay model for prion detection in pigs.

Further characterisation of transmissible spongiform encephalopathy phenotypes after inoculation of cattle with two temporally separated sources of sheep scrapie from Great Britain.

BACKGROUND: The infectious agent responsible for the bovine spongiform encephalopathy (BSE) epidemic in Great Britain is a transmissible spongiform encephalopathy (TSE) strain with uniform properties but the origin of this strain remains unknown. Based on the hypothesis that classical BSE may have been caused by a TSE strain present in sheep, cattle were inoculated intracerebrally with two different pools of brains from scrapie-affected sheep sourced prior to and during the BSE epidemic to investigate resulting disease phenotypes and characterise their causal agents by transmission to rodents. RESULTS: As reported in 2006, intracerebral inoculation of cattle with pre-1975 and post-1990 scrapie brain pools produced two distinct disease phenotypes, which were unlike classical BSE. Subsequent to that report none of the remaining cattle, culled at 10 years post inoculation, developed a TSE. Retrospective Western immunoblot examination of the brains from TSE cases inoculated with the pre-1975 scrapie pool revealed a molecular profile similar to L-type BSE. The inoculation of transgenic mice expressing the bovine, ovine, porcine, murine or human prion protein gene and bank voles with brains from scrapie-affected cattle did not detect classical or atypical BSE strains but identified two previously characterised scrapie strains of sheep. CONCLUSIONS: Characterisation of the causal agents of disease resulting from exposure of cattle to naturally occurring scrapie agents sourced in Great Britain did not reveal evidence of classical or atypical BSE, but did identify two distinct previously recognised strains of scrapie. Although scrapie was still recognizable upon cattle passage there were irreconcilable discrepancies between the results of biological strain typing approaches and molecular profiling methods, suggesting that the latter may not be appropriate for the identification and differentiation of atypical, particularly L-type, BSE agents from cattle experimentally infected with a potential mixture of classical scrapie strains from sheep sources.

Early malnutrition and metabolic abnormalities later in life.

Early malnutrition has been significantly associated with chronic diseases later in life. The finding of metabolic abnormalities in individuals with malnutrition in fetal life and early in postnatal life may have important public health implications in developing countries, although only a few studies have examined the relationship between body weight at the first year of life and later metabolic abnormalities. We assessed the effects of malnutrition during the first year of life (independent of birth weight) on several metabolic parameters in young men. The areas under the curves of glucose and insulin were significantly higher in 52 individuals with a history of malnutrition during the first year of life (cases) than in controls. An insulin sensitivity index was derived and was lower in cases than in controls. In cases, the insulin sensitivity index worsened as body mass index (BMI) increased within the normal range of BMI. A euglycemic insulin clamp as well as an abdominal CT scan were performed in 26 cases and 27 controls. Higher levels of abdominal adipose tissue were more detrimental to insulin sensitivity in previously malnourished individuals. This study suggests that metabolic programming early in life is not restricted to intrauterine growth. We conclude that early malnutrition in extrauterine life independent of birth weight has an adverse effect on insulin sensitivity, glucose tolerance, and lipid profile in young men. This appears to worsen as body mass index increases, even within "normal" limits.

Proteinase K enhanced immunoreactivity of the prion protein-specific monoclonal antibody 2A11.

Here, we report the development and further characterisation of a novel PrP-specific monoclonal antibody: 2A11. By Western blot analysis, 2A11 reacts with PrPC from a variety of species including cow, sheep, pig, hamster, rabbit, cat, dog, deer and mouse but fails to react with human, chicken and turtle PrP. Reactivity to PrPC in Western blot was found to be dependent on the redox state of the protein since binding of mAb 2A11 to its epitope was more effective in reducing conditions. 2A11 binding site was mapped within a region comprised by residues 171-179 (six octarepeats bovine PrP notation; 163-171 for the ovine PrP notation). Interestingly, in immunohistochemistry (IHC) analysis, immunoreactivity was greatly enhanced after proteinase K (PK) sample treatment, while little or no reaction was observed in non-PK-treated BSE samples and samples from healthy animals. Quantitative differences in reactivity to BSE prions after PK treatment were also observed, to a lesser extent, by Western blot analysis. Since definitive diagnosis of prion diseases rely on IHC assays of proteinase K-treated samples, the use of mAb 2A11 might contribute to reduce the occurrence of false positive detection due to incomplete proteinase K digestion.

Comparison of mRNA expression levels of selected genes in the brain stem of cattle naturally infected with classical and atypical BSE.

Since 2004 cases of atypical bovine spongiform encephalopathy (BSE) in older cattle are recorded on the basis of aberrant glycoprofiles of prion protein resistant to proteolysis (PrP(res)). The nature of those types of PrP(res) is still not fully understood but the epidemiological data indicate that their occurrence is rare. Hitherto, most BSE cases were studied on the basis of the features of pathological form of prion protein (PrP(Sc)) or lesions observed in the gray matter of the brain. Here we propose the gene expression profiling as a method to characterize and distinguish BSE types. Thus, the aim of the study was to compare the activity of some genes which are known to play a role in the pathogenesis of transmissible spongiform encephalopathies (TSEs). Significant differences in the expression level of the selected genes in the brain stem were observed for 7 out of 11 genes tested when the results for BSE affected and healthy control animals were compared. Significant up-regulation of caspase 3, Bax and 14-3-3 protein encoding genes was apparent in the obex of all BSE affected cattle regardless of the prion type. Significant and unique to BSE H-type up-regulation was detected in prion and SOD1 genes, while BSE C-type was characterized by higher Bcl-2 and Fyn gene expression levels in respect to other BSE types and control animals. Different gene expression profiles of bovine brains infected with classical and atypical BSE indicate possible different pathogenesis or origin of the disease.