Benefits of Automated Crystallization Plate Tracking, Imaging, and Analysis

SummaryWe describe the design of a database and software for managing and organizing protein crystallization data. We also outline the considerations behind the design of a fast web interface linking protein production data, crystallization images, and automated image analysis. The database and associated interfaces underpin the Oxford Protein Production Facility (OPPF) crystallization laboratory, collecting, in a routine and automatic manner, up to 100,000 images per day. Over 17 million separate images are currently held in this database. We discuss the substantial scientific benefits automated tracking, imaging, and analysis of crystallizations offers to the structural biologist: analysis of the time course of the trial and easy analysis of trials with related crystallization conditions. Features of this system address requirements common to many crystallographic laboratories that are currently setting up (semi-)automated crystallization imaging systems

An overview of harms associated with β-lactam antimicrobials: where do the carbapenems fit in?

The US Institute of Medicine's focus on patient safety has motivated hospital administrators to facilitate a culture of safety. As a result, subcommittees of the pharmacy and therapeutics committee have emerged in many hospitals to focus on adverse events and patient safety. Antimicrobial harms have gained the attention of practicing clinicians and hospital formulary committees, because they top the list of drugs that are associated with adverse events and because of certain serious harms that have ultimately led to the withdrawal of some antimicrobial agents. In the near future, several antimicrobials in the late phase of development will become available for clinical use (ceftobiprole, ceftaroline, and telavancin), and others (doripenem and dalbavancin) have recently joined the armamentarium. Because new antimicrobials will become part of the treatment armamentarium, it is important to discuss our current understanding of antimicrobial harms in general. Although not thought of as traditional adverse events, Clostridium difficile infection and development of resistance during therapy are adverse events that occur as a result of antimicrobial exposure and therefore are discussed. In addition, a distillation of our current understanding of β-lactam specific adverse events will be provided. Finally, new methods of administration are being evaluated that may influence peak concentration-related antimicrobial adverse events

Antipsychotics and Torsadogenic Risk: Signals Emerging from the US FDA Adverse Event Reporting System Database

Background: Drug-induced torsades de pointes (TdP) and related clinical entities represent a current regulatory and clinical burden. Objective: As part of the FP7 ARITMO (Arrhythmogenic Potential of Drugs) project, we explored the publicly available US FDA Adverse Event Reporting System (FAERS) database to detect signals of torsadogenicity for antipsychotics (APs). Methods: Four groups of events in decreasing order of drug-attributable risk were identified: (1) TdP, (2) QT-interval abnormalities, (3) ventricular fibrillation/tachycardia, and (4) sudden cardiac death. The reporting odds ratio (ROR) with 95 % confidence interval (CI) was calculated through a cumulative analysis from group 1 to 4. For groups 1+2, ROR was adjusted for age, gender, and concomitant drugs (e.g., antiarrhythmics) and stratified for AZCERT drugs, lists I and II (http://www.azcert.org, as of June 2011). A potential signal of torsadogenicity was defined if a drug met all the following criteria: (a) four or more cases in group 1+2; (b) significant ROR in group 1+2 that persists through the cumulative approach; (c) significant adjusted ROR for group 1+2 in the stratum without AZCERT drugs; (d) not included in AZCERT lists (as of June 2011). Results: Over the 7-year period, 37 APs were reported in 4,794 cases of arrhythmia: 140 (group 1), 883 (group 2), 1,651 (group 3), and 2,120 (group 4). Based on our criteria, the following potential signals of torsadogenicity were found: amisulpride (25 cases; adjusted ROR in the stratum without AZCERT drugs = 43.94, 95 % CI 22.82-84.60), cyamemazine (11; 15.48, 6.87-34.91), and olanzapine (189; 7.74, 6.45-9.30). Conclusions: This pharmacovigilance analysis on the FAERS found 3 potential signals of torsadogenicity for drugs previously unknown for this risk

Delayed Goblet Cell Hyperplasia, Acetylcholine Receptor Expression, and Worm Expulsion in SMC-Specific IL-4Rα–Deficient Mice

Interleukin 4 receptor α (IL-4Rα) is essential for effective clearance of gastrointestinal nematode infections. Smooth muscle cells are considered to play a role in the type 2 immune response–driven expulsion of gastrointestinal nematodes. Previous studies have shown in vitro that signal transducer and activator of transcription 6 signaling in response to parasitic nematode infection significantly increases smooth muscle cell contractility. Inhibition of the IL-4Rα pathway inhibits this response. How this response manifests itself in vivo is unknown. In this study, smooth muscle cell IL-4Rα–deficient mice (SM-MHC(Cre)IL-4Rα(−/lox)) were generated and characterized to uncover any role for IL-4/IL-13 in this non–immune cell type in response to Nippostrongylus brasiliensis infection. IL-4Rα was absent from α-actin–positive smooth muscle cells, while other cell types showed normal IL-4Rα expression, thus demonstrating efficient cell-type–specific deletion of the IL-4Rα gene. N. brasiliensis–infected SM-MHC(Cre)IL-4Rα(−/lox) mice showed delayed ability to resolve infection with significantly prolonged fecal egg recovery and delayed worm expulsion. The delayed expulsion was related to a delayed intestinal goblet cell hyperplasia, reduced T helper 2 cytokine production in the mesenteric lymph node, and reduced M3 muscarinic receptor expression during infection. Together, these results demonstrate that in vivo IL-4Rα–responsive smooth muscle cells are beneficial for N. brasiliensis expulsion by coordinating T helper 2 cytokine responses, goblet hyperplasia, and acetylcholine responsiveness, which drive smooth muscle cell contractions

Dynamic geospatial modeling of mycotoxin contamination of corn in Illinois: unveiling critical factors and predictive insights with machine learning

Mycotoxin contamination of corn is a pervasive problem that negatively impacts human and animal health and causes economic losses to the agricultural industry worldwide. Historical aflatoxin (AFL) and fumonisin (FUM) mycotoxin contamination data of corn, daily weather data, satellite data, dynamic geospatial soil properties, and land usage parameters were modeled to identify factors significantly contributing to the outbreaks of mycotoxin contamination of corn grown in Illinois (IL), AFL >20 ppb, and FUM >5 ppm. Two methods were used: a gradient boosting machine (GBM) and a neural network (NN). Both the GBM and NN models were dynamic at a state-county geospatial level because they used GPS coordinates of the counties linked to soil properties. GBM identified temperature and precipitation prior to sowing as significant influential factors contributing to high AFL and FUM contamination. AFL-GBM showed that a higher aflatoxin risk index (ARI) in January, March, July, and November led to higher AFL contamination in the southern regions of IL. Higher values of corn-specific normalized difference vegetation index (NDVI) in July led to lower AFL contamination in Central and Southern IL, while higher wheat-specific NDVI values in February led to higher AFL. FUM-GBM showed that temperature in July and October, precipitation in February, and NDVI values in March are positively correlated with high contamination throughout IL. Furthermore, the dynamic geospatial models showed that soil characteristics were correlated with AFL and FUM contamination. Greater calcium carbonate content in soil was negatively correlated with AFL contamination, which was noticeable in Southern IL. Greater soil moisture and available water-holding capacity throughout Southern IL were positively correlated with high FUM contamination. The higher clay percentage in the northeastern areas of IL negatively correlated with FUM contamination. NN models showed high class-specific performance for 1-year predictive validation for AFL (73%) and FUM (85%), highlighting their accuracy for annual mycotoxin prediction. Our models revealed that soil, NDVI, year-specific weekly average precipitation, and temperature were the most important factors that correlated with mycotoxin contamination. These findings serve as reliable guidelines for future modeling efforts to identify novel data inputs for the prediction of AFL and FUM outbreaks and potential farm-level management practices

Calpain Cleavage of Brain Glutamic Acid Decarboxylase 65 Is Pathological and Impairs GABA Neurotransmission

Previously, we have shown that the GABA synthesizing enzyme, L-glutamic acid decarboxylase 65 (GAD65) is cleaved to form its truncated form (tGAD65) which is 2–3 times more active than the full length form (fGAD65). The enzyme responsible for cleavage was later identified as calpain. Calpain is known to cleave its substrates either under a transient physiological stimulus or upon a sustained pathological insult. However, the precise role of calpain cleavage of fGAD65 is poorly understood. In this communication, we examined the cleavage of fGAD65 under diverse pathological conditions including rats under ischemia/reperfusion insult as well as rat brain synaptosomes and primary neuronal cultures subjected to excessive stimulation with high concentration of KCl. We have shown that the formation of tGAD65 progressively increases with increasing stimulus concentration both in rat brain synaptosomes and primary rat embryo cultures. More importantly, direct cleavage of synaptic vesicle - associated fGAD65 by calpain was demonstrated and the resulting tGAD65 bearing the active site of the enzyme was detached from the synaptic vesicles. Vesicular GABA transport of the newly synthesized GABA was found to be reduced in calpain treated SVs. Furthermore, we also observed that the levels of tGAD65 in the focal cerebral ischemic rat brain tissue increased corresponding to the elevation of local glutamate as indicated by microdialysis. Moreover, the levels of tGAD65 was also proportional to the degree of cell death when the primary neuronal cultures were exposed to high KCl. Based on these observations, we conclude that calpain-mediated cleavage of fGAD65 is pathological, presumably due to decrease in the activity of synaptic vesicle - associated fGAD65 resulting in a decrease in the GABA synthesis - packaging coupling process leading to reduced GABA neurotransmission

NeuroGrid: recording action potentials from the surface of the brain.

Recording from neural networks at the resolution of action potentials is critical for understanding how information is processed in the brain. Here, we address this challenge by developing an organic material-based, ultraconformable, biocompatible and scalable neural interface array (the 'NeuroGrid') that can record both local field potentials(LFPs) and action potentials from superficial cortical neurons without penetrating the brain surface. Spikes with features of interneurons and pyramidal cells were simultaneously acquired by multiple neighboring electrodes of the NeuroGrid, allowing for the isolation of putative single neurons in rats. Spiking activity demonstrated consistent phase modulation by ongoing brain oscillations and was stable in recordings exceeding 1 week's duration. We also recorded LFP-modulated spiking activity intraoperatively in patients undergoing epilepsy surgery. The NeuroGrid constitutes an effective method for large-scale, stable recording of neuronal spikes in concert with local population synaptic activity, enhancing comprehension of neural processes across spatiotemporal scales and potentially facilitating diagnosis and therapy for brain disorders