ER Stress Induced by Tunicamycin Triggers α-Synuclein Oligomerization,Dopaminergic Neurons Death and Locomotor Impairment: a New Model of Parkinson’s Disease

Parkinson’s disease (PD) is a neurodegenerative disorder characterized by progressive death of dopaminergic neurons of the substantia nigra pars compacta (SNpc), leading to the major clinical abnormalities that characterize this disease. Although PD’s etiology is unknown, α-synuclein aggregation plays a pivotal role in PD pathogenesis, which could be associated to some pathological processes such as oxidative stress, endoplasmic reticulum (ER) stress, impaired protein degradation, and mitochondrial dysfunction. Increasing experimental evidence indicates that ER stress is involved in PD, however most of the described results employed cultured cell lines and genetically modified animal models. In this study, we developed a new ER stress rat model employing the well-known ER stressor tunicamycin (Tm). To evaluate if ER stress was able to induce PD features, we performed an intranigral injection of Tm (0.1 μg/cerebral hemisphere) and animals (male Wistar rats) were analyzed 7 days post injection. The classical 6-OHDA neurotoxin model (1 μg/cerebral hemisphere) was used as an established positive control for PD. We show that Tm injection induced locomotor impairment, dopaminergic neurons death, and activation of astroglia. In addition, we observed an extensive α-synuclein oligomerization in SNpc of Tm-injected animals when compared with DMSO-injected controls. Finally, both Tm and 6-OHDA treated animals presented increased levels of ER stress markers. Taken together, these findings show for the first time that the ER stressor Tm recapitulates some of the phenotypic characteristics observed in rodent models of PD, reinforcing the concept that ER stress could be an important contributor to the pathophysiology of PD. Therefore, we propose the intranigral Tm injection as a new ER stress-based model for the study of PD in vivo.     

Antenatal breast expression in women with diabetes: outcomes from a retrospective cohort study

Background

Women with diabetes are sometimes advised to express breast milk antenatally to prepare for breastfeeding and to store colostrum for infant feeding in preventing or treating hypoglycaemia after the birth. The acceptability, risks and benefits of this practice have not been evaluated. This was aimed to investigate the pattern of antenatal breast expression uptake and its relationship with birth outcomes in women with diabetes.

Methods

This was part of a two year retrospective cohort study of pregnant women with diabetes (type 1, 2 and gestational diabetes) who gave birth during 2001–2003 in Derby Hospitals NHS Foundation Trust (n = 94). The information on the practice of antenatal breastfeeding expression and birth outcomes was collected via self-administered questionnaires and by examining maternity records.

Results

Thirty-seven percent of women (35/94) recalled that they were advised to express antenatally and 17% did (16/94). The mean gestational age at birth for women who hand-expressed was lower than that for those who did not (mean difference (MD) (95% confidence intervals (CI)): -1.2 (?2.4 to 0.04), p = 0.06). A higher proportion of babies from the antenatal expression group were admitted to special care baby units (SCBU) (MD (95% CI): 21% (?3.9 to 46.3).

Conclusions

Less than half the women who stated that they were advised to express, did so. There seems to be an indication that antenatal breast milk expression and lower gestational age at birth are associated. The trend of a higher rate of SCBU admission for babies from the breast milk expression group compared to those who did not express antenatally is of concern. An appropriately-powered randomised controlled trial is needed to determine the safety of this practice and its acceptability to women and health professionals before it can be recommended for implementation in practice.

     

Alternative splicing and protein function

Background  

Alternative splicing is a major mechanism of generating protein diversity in higher eukaryotes. Although at least half, and probably more, of mammalian genes are alternatively spliced, it was not clear, whether the frequency of alternative splicing is the same in different functional categories. The problem is obscured by uneven coverage of genes by ESTs and a large number of artifacts in the EST data.     

Detection of African horse sickness virus in Culicoides imicola pools using RT‐qPCR

African horse sickness (AHS) is an infectious, non‐contagious arthropod‐borne disease of equids, caused by the African horse sickness virus (AHSV), an orbivirus of the Reoviridae family. It is endemic in sub‐Saharan Africa and thought to be the most lethal viral disease of horses. This study focused on detection of AHSV in Culicoides imicola (Diptera: Ceratopogonidae) pools by the application of a RT‐qPCR. Midges were fed on AHSV‐infected blood. A single blood‐engorged female was allocated to pools of unfed nulliparous female midges. Pool sizes varied from 1 to 200. RNA was extracted and prepared for RT‐qPCR. The virus was successfully detected and the optimal pool size for the limit of detection of the virus was determined at a range between 1 to 25. Results from this investigation highlight the need for a standardized protocol for AHSV investigation in Culicoides midges especially for comparison among different studies and for the determination of infection rate.     

Enhanced glutamate,IP3 and cAMP activity in the cerebral cortex of Unilateral 6-hydroxydopamine induced Parkinson's rats: Effect of 5-HT,GABA and bone marrow cell supplementation

Parkinson's disease is characterized by progressive cell death in the substantia nigra pars compacta, which leads to dopamine depletion in the striatum and indirectly to cortical dysfunction. Increased glutamatergic transmission in the basal ganglia is implicated in the pathophysiology of Parkinson's disease and glutamate receptor mediated excitotoxicity has been suggested to be one of the possible causes of the neuronal degeneration. In the present study, the effects of serotonin, gamma-aminobutyric acid and bone marrow cells infused intranigrally to substantia nigra individually and in combination on unilateral 6-hydroxydopamine induced Parkinson's rat model was analyzed. Scatchard analysis of total glutamate and NMDA receptor binding parameters showed a significant increase in B_max (P < 0.001) in the cerebral cortex of 6-hydroxydopamine infused rat compared to control. Real Time PCR amplification of NMDA2B, mGluR5, bax, and ubiquitin carboxy-terminal hydrolase were up regulated in cerebral cortex of 6-hydroxydopamine infused rats compared to control. Gene expression studies of GLAST, ά-Synuclien and Cyclic AMP response element-binding protein showed a significant (P < 0.001) down regulation in 6-OHDA infused rats compared to control. Behavioural studies were carried out to confirm the biochemical and molecular studies. Serotonin and GABA along with bone marrow cells in combination showed reversal of glutamate receptors and behaviour abnormality shown in the Parkinson's rat model. The therapeutic significance in Parkinson's disease is of prominence.     

Ratio-dependent parasitism with Trissolcus basalis (Wollaston) (Hymenoptera: Scelionidae) on egg rafts of Nezara viridula (Linnaeus) (Hemiptera: Pentatomidae): effect of experimental variables and compatibility of 'ratio' and 'Holling' models

Abstract Egg rafts of Nezara viridula were exposed to the parasitoid wasp Trissolcus basalis in experimental arenas to establish the relationship of the rates of attack and parasitism to various combinations of arena size, parasitoid density, host density and parasitoid-to-host ratio. Arena sizes were varied in the ratio 1:9:63, with the largest having a search area of 1.44 m². Parasitoid and host densities were varied over a 27-fold range. The parasitoid-to-host ratios used were 1:1, 3:1 and 6:1. Finding time was related to a constant factor (flight propensity), rather than to the difficulty of finding (density of hosts). Initial attack rates were therefore related only to parasitoid numbers (or density), even at the lower densities and ratios. Parasitism rates (a function of attack rate per host) were thus also strongly related to parasitoid to host ratio, regardless of densities used and arena sizes. Even reducing host density, while keeping time and parasitoid density constant, increased the parasitism rate. A ratio model for parasitism rate was therefore compatible with the data but the more explicit Holling 'disc' equation was also compatible because handling time was sufficiently large to make it sensitive to the ratio of parasitoids to hosts for the densities used. We conclude that the two models would predict different results if the density of host egg rafts was in a range below one per square metre.     

Algorithms and software for support of gene identification experiments

MOTIVATION: Gene annotation is the final goal of gene prediction algorithms. However, these algorithms frequently make mistakes and therefore the use of gene predictions for sequence annotation is hardly possible. As a result, biologists are forced to conduct time-consuming gene identification experiments by designing appropriate PCR primers to test cDNA libraries or applying RT-PCR, exon trapping/amplification, or other techniques. This process frequently amounts to 'guessing' PCR primers on top of unreliable gene predictions and frequently leads to wasting of experimental efforts. RESULTS: The present paper proposes a simple and reliable algorithm for experimental gene identification which bypasses the unreliable gene prediction step. Studies of the performance of the algorithm on a sample of human genes indicate that an experimental protocol based on the algorithm's predictions achieves an accurate gene identification with relatively few PCR primers. Predictions of PCR primers may be used for exon amplification in preliminary mutation analysis during an attempt to identify a gene responsible for a disease. We propose a simple approach to find a short region from a genomic sequence that with high probability overlaps with some exon of the gene. The algorithm is enhanced to find one or more segments that are probably contained in the translated region of the gene and can be used as PCR primers to select appropriate clones in cDNA libraries by selective amplification. The algorithm is further extended to locate a set of PCR primers that uniformly cover all translated regions and can be used for RT-PCR and further sequencing of (unknown) mRNA.