Remote detection of plant physiological responses to TNT soil contamination

Our study was aimed at understanding physiological responses to trinitrotoluene (TNT) soil contamination, and using optical methods to detect TNT-induced stress in a woody plant prior to visible changes. Myrica cerifera plants were potted in soil concentrations of TNT ranging from 30–500 mg kg^?1. Physiological measurements were significantly affected by TNT exposure at all treatment levels, and photosynthetic decline likely resulted from metabolic impairment rather than stomatal closure as the experiment progressed. Several reflectance indices were able to detect TNT-induced stress before any changes in chlorophyll concentrations occurred. The most sensitive index was the simple ratio R₇₆₁/R₇₅₇ which is linked to fluorescence in-filling of the 0₂ atmospheric absorption. Changes at R₇₄₀/R₈₅₀ and R₇₃₅/R₈₅₀ may be attributed to both fluorescence and structural characteristics of leaf anatomy in the near infrared region. This could have been influenced by transformation and conjugation of TNT metabolites with other compounds. chlorophyll index (CHL) or in the water band index (WBI₉₇₀), which are indices typically associated with drought stress, and may provide a means of separating stress due to explosives. Further studies need to be conducted with a combination of stressors (TNT and natural) to determine if responses are in fact generalized or if any of these changes are separable from natural stress.     

Early activation of wheat polyamine biosynthesis during Fusarium head blight implicates putrescine as an inducer of trichothecene mycotoxin production

Background  

The fungal pathogen Fusarium graminearum causes Fusarium Head Blight (FHB) disease on wheat which can lead to trichothecene mycotoxin (e.g. deoxynivalenol, DON) contamination of grain, harmful to mammalian health. DON is produced at low levels under standard culture conditions when compared to plant infection but specific polyamines (e.g. putrescine and agmatine) and amino acids (e.g. arginine and ornithine) are potent inducers of DON by F. graminearum in axenic culture. Currently, host factors that promote mycotoxin synthesis during FHB are unknown, but plant derived polyamines could contribute to DON induction in infected heads. However, the temporal and spatial accumulation of polyamines and amino acids in relation to that of DON has not been studied.     

Improving the clinical assessment of consciousness with advances in electrophysiological and neuroimaging techniques

In clinical neurology, a comprehensive understanding of consciousness has been regarded as an abstract concept - best left to philosophers. However, times are changing and the need to clinically assess consciousness is increasingly becoming a real-world, practical challenge. Current methods for evaluating altered levels of consciousness are highly reliant on either behavioural measures or anatomical imaging. While these methods have some utility, estimates of misdiagnosis are worrisome (as high as 43%) - clearly this is a major clinical problem. The solution must involve objective, physiologically based measures that do not rely on behaviour. This paper reviews recent advances in physiologically based measures that enable better evaluation of consciousness states (coma, vegetative state, minimally conscious state, and locked in syndrome). Based on the evidence to-date, electroencephalographic and neuroimaging based assessments of consciousness provide valuable information for evaluation of residual function, formation of differential diagnoses, and estimation of prognosis.     

Enrichment and Site Mapping of O-Linked N-Acetylglucosamine by a Combination of Chemical/Enzymatic Tagging, Photochemical Cleavage, and Electron Transfer Dissociation Mass Spectrometry

Numerous cellular processes are regulated by the reversible addition of either phosphate or O-linked β-N-acetylglucosamine (O-GlcNAc) to nuclear and cytoplasmic proteins. Although sensitive methods exist for the enrichment and identification of protein phosphorylation sites, those for the enrichment of O-GlcNAc-containing peptides are lacking. Reported here is highly efficient methodology for the enrichment and characterization of O-GlcNAc sites from complex samples. In this method, O-GlcNAc-modified peptides are tagged with a novel biotinylation reagent, enriched by affinity chromatography, released from the solid support by photochemical cleavage, and analyzed by electron transfer dissociation mass spectrometry. Using this strategy, eight O-GlcNAc sites were mapped from a tau-enriched sample from rat brain. Sites of GlcNAcylation were characterized on important neuronal proteins such as tau, synucleins, and methyl CpG-binding protein 2.Numerous cytoplasmic and nuclear proteins are post-translationally modified with O-linked β-N-acetylglucosamine (O-GlcNAc).¹ GlcNAcylation is involved in almost all aspects of cellular metabolism (1) and is highly dependent on the nutrient status of the cell (2). The O-GlcNAc modification rivals phosphorylation in both abundance and protein distribution. Recent studies indicate that signaling pathways can be regulated by the interplay of these two modifications at the same or proximal sites on numerous protein substrates (3).Current understanding of the functions of O-GlcNAc and of the function of O-GlcNAcylation and its relationship to phosphorylation is severely hampered by the difficulties in detecting this labile monosaccharide modification. Problems associated with the identification of O-GlcNAc sites include the following. (a) O-GlcNAc is quickly removed by hydrolases during cell lysis. (b) Like phosphorylation, O-GlcNAc is usually present in less than stoichiometric amounts at given sites on protein substrates. (c) O-GlcNAc is readily lost as an oxonium ion during conventional peptide sequence analysis by collision-activated dissociation (CAD) (supplemental Fig. 1). (d) Modified and unmodified forms of the peptide often co-elute during reverse phase HPLC (supplemental Fig. 2), and the preferential ionization of the unmodified peptide suppresses the signal observed for the corresponding O-GlcNAc-modified peptide (supplemental Fig. 2, b and c).Several attempts have been made to enrich samples for O-GlcNAc-modified proteins and peptides. Immunoaffinity purification of O-GlcNAc-modified peptides with an antibody (CTD 110.6) has been largely unsuccessful because of low binding avidity (4). Long, wheat germ agglutinin lectin columns (∼39 ft) provide some enrichment but also bind strongly to complex glycans (5). A mutant galactosyltransferase (GalT1) has been used to label GlcNAcylated proteins with a ketone-containing galactose analog (6). Following proteolytic digestion, O-GlcNAc-modified peptides were biotinylated with hydrazine chemistry, isolated on a column packed with avidin beads, eluted with free biotin, and sequenced by ETD mass spectrometry. Failure to elute peptides with high efficiency from the avidin column and an inability to direct the fragmentation to the peptide backbone limit the usefulness of this approach. Reported here is an enrichment methodology that (a) is highly specific for O-GlcNAc-modified peptides, (b) provides for efficient release of the captured peptides from an affinity support, and (c) facilitates complete characterization of the released peptides by ETD mass spectrometry.     

Calcitriol enhances gemcitabine antitumor activity in vitro and in vivo by promoting apoptosis in a human pancreatic carcinoma model system

Gemcitabine is the standard care chemotherapeutic agent to treat pancreatic cancer. Previously we demonstrated that calcitriol (1, 25-dihydroxycholecalciferol) has significant anti-proliferative effects in vitro and in vivo in multiple tumor models and enhances the activity of a variety of chemotherapeutic agents. We therefore investigated whether calcitriol could potentiate the cytotoxic activity of gemcitabine in the human pancreatic cancer Capan-1 model system. Isobologram analysis revealed that calcitriol and gemcitabine had synergistic antiproliferative effect over a wide range of drug concentrations. Calcitriol did not reduce the cytidine deaminase activity in Capan-1 tumors nor in the livers of Capan-1 tumor bearing mice. Calcitriol and gemcitabine combination promoted apoptosis in Capan-1 cells compared with either agent alone. The combination treatment also increased the activation of caspases-8, -9, -6 and -3 in Capan-1 cells. This result was confirmed by substrate-based caspase activity assay. Akt phosphorylation was reduced by calcitriol and gemcitabine combination treatment compared to single agent treatment. However, ERK1/2 phosphorylation was not modulated by either agent alone or by the combination. Tumor regrowth delay studies showed that calcitriol in combination with gemcitabine resulted in a significant reduction of Capan-1 tumor volume compared to single agent treatment. Our study suggests that calcitriol and gemcitabine in combination promotes caspase-dependent apoptosis, which may contribute to increased anti-tumor activity compared to either agent alone.Key words: calcitriol, gemcitabine, pancreatic carcinoma, apoptosis, Akt, ERK1/2     

Bisphenol A: developmental toxicity from early prenatal exposure

Bisphenol A (BPA) exposure has been documented in pregnant women, but consequences for development are not yet widely studied in human populations. This review presents research on the consequences for offspring of BPA exposure during pregnancy. Extensive work in laboratory rodents has evaluated survival and growth of the conceptus, interference with embryonic programs of development, morphological sex differentiation, sex differentiation of the brain and behavior, immune responsiveness, and mechanism of action. Sensitive measures include RAR, aryl hydrocarbon receptor, and Hox A10 gene expression, anogenital distance, sex differentiation of affective and exploratory behavior, and immune hyperresponsiveness. Many BPA effects are reported at low doses (10–50 µg/kg d range) by the oral route of administration. At high doses (>500,000 µg/kg d) fetal viability is compromised. Much of the work has centered around the implications of the estrogenic actions of this agent. Some work related to thyroid mechanism of action has also been explored. BPA research has actively integrated current knowledge of developmental biology, concepts of endocrine disruption, and toxicological research to provide a basis for human health risk assessment. Birth Defects Res (Part B) 89:441–466, 2010. © 2010 Wiley‐Liss, Inc.     

Community Case Management of Fever Due to Malaria and Pneumonia in Children Under Five in Zambia: A Cluster Randomized Controlled Trial

Background

Pneumonia and malaria, two of the leading causes of morbidity and mortality among children under five in Zambia, often have overlapping clinical manifestations. Zambia is piloting the use of artemether-lumefantrine (AL) by community health workers (CHWs) to treat uncomplicated malaria. Valid concerns about potential overuse of AL could be addressed by the use of malaria rapid diagnostics employed at the community level. Currently, CHWs in Zambia evaluate and treat children with suspected malaria in rural areas, but they refer children with suspected pneumonia to the nearest health facility. This study was designed to assess the effectiveness and feasibility of using CHWs to manage nonsevere pneumonia and uncomplicated malaria with the aid of rapid diagnostic tests (RDTs).

Methods and Findings

Community health posts staffed by CHWs were matched and randomly allocated to intervention and control arms. Children between the ages of 6 months and 5 years were managed according to the study protocol, as follows. Intervention CHWs performed RDTs, treated test-positive children with AL, and treated those with nonsevere pneumonia (increased respiratory rate) with amoxicillin. Control CHWs did not perform RDTs, treated all febrile children with AL, and referred those with signs of pneumonia to the health facility, as per Ministry of Health policy. The primary outcomes were the use of AL in children with fever and early and appropriate treatment with antibiotics for nonsevere pneumonia. A total of 3,125 children with fever and/or difficult/fast breathing were managed over a 12-month period. In the intervention arm, 27.5% (265/963) of children with fever received AL compared to 99.1% (2066/2084) of control children (risk ratio 0.23, 95% confidence interval 0.14–0.38). For children classified with nonsevere pneumonia, 68.2% (247/362) in the intervention arm and 13.3% (22/203) in the control arm received early and appropriate treatment (risk ratio 5.32, 95% confidence interval 2.19–8.94). There were two deaths in the intervention and one in the control arm.

Conclusions

The potential for CHWs to use RDTs, AL, and amoxicillin to manage both malaria and pneumonia at the community level is promising and might reduce overuse of AL, as well as provide early and appropriate treatment to children with nonsevere pneumonia.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00513500","term_id":"NCT00513500"}}NCT00513500 Please see later in the article for the Editors'' Summary     

Non-centered spike-triggered covariance analysis reveals neurotrophin-3 as a developmental regulator of receptive field properties of ON-OFF retinal ganglion cells

The functional separation of ON and OFF pathways, one of the fundamental features of the visual system, starts in the retina. During postnatal development, some retinal ganglion cells (RGCs) whose dendrites arborize in both ON and OFF sublaminae of the inner plexiform layer transform into RGCs with dendrites that monostratify in either the ON or OFF sublamina, acquiring final dendritic morphology in a subtype-dependent manner. Little is known about how the receptive field (RF) properties of ON, OFF, and ON-OFF RGCs mature during this time because of the lack of a reliable and efficient method to classify RGCs into these subtypes. To address this deficiency, we developed an innovative variant of Spike Triggered Covariance (STC) analysis, which we term Spike Triggered Covariance - Non-Centered (STC-NC) analysis. Using a multi-electrode array (MEA), we recorded the responses of a large population of mouse RGCs to a Gaussian white noise stimulus. As expected, the Spike-Triggered Average (STA) fails to identify responses driven by symmetric static nonlinearities such as those that underlie ON-OFF center RGC behavior. The STC-NC technique, in contrast, provides an efficient means to identify ON-OFF responses and quantify their RF center sizes accurately. Using this new tool, we find that RGCs gradually develop sensitivity to focal stimulation after eye opening, that the percentage of ON-OFF center cells decreases with age, and that RF centers of ON and ON-OFF cells become smaller. Importantly, we demonstrate for the first time that neurotrophin-3 (NT-3) regulates the development of physiological properties of ON-OFF center RGCs. Overexpression of NT-3 leads to the precocious maturation of RGC responsiveness and accelerates the developmental decrease of RF center size in ON-OFF cells. In summary, our study introduces STC-NC analysis which successfully identifies subtype RGCs and demonstrates how RF development relates to a neurotrophic driver in the retina.