Left Frontoparietal Control Network Connectivity Moderates the Effect of Amyloid on Cognitive Decline in Preclinical Alzheimer's Disease: The A4 Study.

BACKGROUND: Stronger resting-state functional connectivity of the default mode and frontoparietal control networks has been associated with cognitive resilience to Alzheimer's disease related pathology and neurodegeneration in smaller cohort studies. OBJECTIVES: We investigated whether these networks are associated with longitudinal CR to AD biomarkers of beta-amyloid (Aß). DESIGN: Longitudinal mixed. SETTING: The Anti-Amyloid Treatment in Asymptomatic Alzheimer's Disease (A4) study and its natural history observation arm, the Longitudinal Evaluation of Amyloid Risk and Neurodegeneration (LEARN) study. PARTICIPANTS: A sample of 1,021 cognitively unimpaired older adults (mean age = 71.2 years [SD = 4.7 years], 61% women, 42% APOEε4 carriers, 52% Aß positive). MEASUREMENTS: Global cognitive performance (Preclinical Alzheimer's Cognitive Composite) was assessed over an average 5.4 year follow-up period (SD = 2 years). Cortical Aß and functional connectivity (left and right frontoparietal control and default mode networks) were estimated from fMRI and PET, respectively, at baseline. Covariates included baseline age, APOEε4 carrier status, years of education, adjusted gray matter volume, head motion, study group, cumulative treatment exposure, and cognitive test version. RESULTS: Mixed effects models revealed that functional connectivity of the left frontoparietal control network moderated the negative effect of Aß on cognitive change (p = .025) such that stronger connectivity was associated with reduced Aß-related cognitive decline. CONCLUSIONS: Our results demonstrate a potential protective effect of functional connectivity in preclinical AD, such that stronger connectivity in this network is associated with slower Aß-related cognitive decline.

Synergism of small molecules targeting VDAC with sorafenib, regorafenib or lenvatinib on hepatocarcinoma cell proliferation and survival.

Voltage dependent anion channels (VDAC) in the outer mitochondrial membrane regulate the influx of metabolites that sustain mitochondrial metabolism and the efflux of ATP to the cytosol. Free tubulin and NADH close VDAC. The VDAC-binding small molecules X1 and SC18 modulate mitochondrial metabolism. X1 antagonizes the inhibitory effect of tubulin on VDAC. SC18 occupies an NADH-binding pocket in the inner wall of all VDAC isoforms. Here, we hypothesized that X1 and SC18 have a synergistic effect with sorafenib, regorafenib or lenvatinib to arrest proliferation and induce death in hepatocarcinoma cells. We used colony formation assays to determine cell proliferation, and a combination of calcein/propidium iodide, and trypan blue exclusion to assess cell death in the well differentiated Huh7 and the poorly differentiated SNU-449 cells. Synergism was assessed using the Chou-Talalay method. The inhibitory effect of X1, SC18, sorafenib, regorafenib and lenvatinib was concentration and time dependent. IC50s calculated from the inhibition of clonogenic capacity were lower than those determined from cell survival. At IC50s that inhibited cell proliferation, SC18 arrested cells in G0/G1. SC18 at 0.25-2 IC50s had a synergistic effect with sorafenib on clonogenic inhibition in Huh7 and SNU-449 cells, and with regorafenib or lenvatinib in SNU-449 cells. X1 or SC18 also had synergistic effects with sorafenib on promoting cell death at 0.5-2 IC50s for SC18 in Huh7 and SNU-449 cells. These results suggest that small molecules targeting VDAC represent a potential new class of drugs to treat liver cancer.

Thermal desorption effects on fragment ion production from multi-photon ionized uridine and selected analogues.

Experiments on neutral gas-phase nucleosides are often complicated by thermal lability. Previous mass spectrometry studies of nucleosides have identified enhanced relative production of nucleobase ions (e.g. uracil+ from uridine) as a function of desorption temperature to be the critical indicator of thermal decomposition. On this basis, the present multi-photon ionization (MPI) experiments demonstrate that laser-based thermal desorption is effective for producing uridine, 5-methyluridine, and 2'-deoxyuridine targets without thermal decomposition. Our experiments also revealed one notable thermal dependence: the relative production of the sugar ion C5H9O4 + from intact uridine increased substantially with the desorption laser power and this only occurred at MPI wavelengths below 250 nm (full range studied 222-265 nm). We argue that this effect can only be rationalized plausibly in terms of changing populations of different isomers, tautomers, or conformers in the target as a function of the thermal desorption conditions. Furthermore, the wavelength threshold behavior of this thermally-sensitive MPI channel indicates a critical dependence on neutral excited state dynamics between the absorption of the first and second photons. The experimental results are complemented by density functional theory (DFT) optimizations of the lowest-energy structure of uridine and two further conformers distinguished by different orientations of the hydroxymethyl group on the sugar part of the molecule. The energies of the transitions states between these three conformers are low compared with the energy required for decomposition.

Retrograde signaling by a mtDNA-encoded non-coding RNA preserves mitochondrial bioenergetics.

Alveolar epithelial type II (AETII) cells are important for lung epithelium maintenance and function. We demonstrate that AETII cells from mouse lungs exposed to cigarette smoke (CS) increase the levels of the mitochondria-encoded non-coding RNA, mito-RNA-805, generated by the control region of the mitochondrial genome. The protective effects of mito-ncR-805 are associated with positive regulation of mitochondrial energy metabolism, and respiration. Levels of mito-ncR-805 do not relate to steady-state transcription or replication of the mitochondrial genome. Instead, CS-exposure causes the redistribution of mito-ncR-805 from mitochondria to the nucleus, which correlated with the increased expression of nuclear-encoded genes involved in mitochondrial function. These studies reveal an unrecognized mitochondria stress associated retrograde signaling, and put forward the idea that mito-ncRNA-805 represents a subtype of small non coding RNAs that are regulated in a tissue- or cell-type specific manner to protect cells under physiological stress.

A new technique for probing chirality via photoelectron circular dichroism.

We present a proof-of-principle approach for discriminating chiral enantiomers based on the phenomenon of multiphoton photoelectron circular dichroism. A novel stereo detection setup was used to measure the number of photoelectrons emitted from chiral molecules in directions parallel or anti-parallel to the propagation of the ionising femtosecond laser pulses. In this study, we show how these asymmetries in the ketones camphor and fenchone depend upon the ellipticity of the laser pulses and the enantiomeric excess of the sample. By using a high repetition rate femtosecond laser, enantiomer excesses with uncertainties at the few-percent level could be measured in close to real-time. As the instrument is compact, and commercial turnkey femtosecond lasers are readily available, the development of a stand-alone chiral analysis instrument for a range of applications is now possible.

Dynamic damage in carbon-fibre composites.

The Taylor test is used to determine damage evolution in carbon-fibre composites across a range of strain rates. The hierarchy of damage across the scales is key in determining the suite of operating mechanisms and high-speed diagnostics are used to determine states during dynamic loading. Experiments record the test response as a function of the orientation of the cylinder cut from the engineered multi-ply composite with high-speed photography and post-mortem target examination. The ensuing damage occurs during the shock compression phase but three other tensile loading modes operate during the test and these are explored. Experiment has shown that ply orientations respond to two components of release; longitudinal and radial as well as the hoop stresses generated in inelastic flow at the impact surface. The test is a discriminant not only of damage thresholds but of local failure modes and their kinetics. This article is part of the themed issue 'Multiscale modelling of the structural integrity of composite materials'.

Pathophysiology of fecal incontinence differs between men and women: a case-matched study in 200 patients.

BACKGROUND: Fecal incontinence (FI) is a common and socially disabling condition with obstetric trauma considered the principal etiological factor. This study aimed to systematically evaluate symptom presentation and anorectal function in both females and males with FI. METHODS: One hundred males (M) and 100 age-matched females (F) with FI presenting between 2012 and 2014 were identified from a prospectively collected database. Comparison of clinical (history, symptom profile, and severity using validated questionnaires) and anorectal physiological (manometry, rectal sensory testing, endoanal ultrasonography, and evacuation proctography) data between M and F was performed. KEY RESULTS: Incidence of prior anal surgery (M: 28% vs F: 18%, p = 0.13) and abdominal surgery (M: 25% vs F: 26%, p = 0.90) was similar between sexes, but females had a higher incidence of previous pelvic surgery (M: 4% vs F: 47%, p < 0.001). Eighty-five females were parous and 75% reported history of traumatic vaginal delivery. There was a trend toward higher St Mark's incontinence scores in females (mean ± SD; M: 13 ± 4 vs F: 14 ± 5, p = 0.06). In men, structural sphincter abnormalities were uncommon (M: 37% vs F: 77%, p < 0.001), while impaired rectal sensation (M: 24% vs F: 7%, p = 0.001) and functional disturbances of evacuation (M: 36% vs F: 13%, p = 0.001) were more common than in women. No abnormality on all tests performed was observed in twice as many males (M: 18% vs F: 9%, p = 0.10). CONCLUSIONS & INFERENCES: Pathophysiological mechanisms of FI differ between sexes. Anal sphincter dysfunction was an uncommon finding in males, with impaired rectal sensation and functional disturbances of evacuation much more prominent than in the female cohort. These findings are likely to impact options for symptom management.

The design and implementation of a study to investigate the effectiveness of community vs hospital eye service follow-up for patients with neovascular age-related macular degeneration with quiescent disease.

IntroductionStandard treatment for neovascular age-related macular degeneration (nAMD) is intravitreal injections of anti-VEGF drugs. Following multiple injections, nAMD lesions often become quiescent but there is a high risk of reactivation, and regular review by hospital ophthalmologists is the norm. The present trial examines the feasibility of community optometrists making lesion reactivation decisions.MethodsThe Effectiveness of Community vs Hospital Eye Service (ECHoES) trial is a virtual trial; lesion reactivation decisions were made about vignettes that comprised clinical data, colour fundus photographs, and optical coherence tomograms displayed on a web-based platform. Participants were either hospital ophthalmologists or community optometrists. All participants were provided with webinar training on the disease, its management, and assessment of the retinal imaging outputs. In a balanced design, 96 participants each assessed 42 vignettes; a total of 288 vignettes were assessed seven times by each professional group.The primary outcome is a participant's judgement of lesion reactivation compared with a reference standard. Secondary outcomes are the frequency of sight threatening errors; judgements about specific lesion components; participant-rated confidence in their decisions about the primary outcome; cost effectiveness of follow-up by optometrists rather than ophthalmologists.DiscussionThis trial addresses an important question for the NHS, namely whether, with appropriate training, community optometrists can make retreatment decisions for patients with nAMD to the same standard as hospital ophthalmologists. The trial employed a novel approach as participation was entirely through a web-based application; the trial required very few resources compared with those that would have been needed for a conventional randomised controlled clinical trial.

The prediction of postoperative stroke or death in patients with preoperative atrial fibrillation undergoing non-cardiac surgery: a VISION sub-study.

BACKGROUND: The optimal means of pre-operative risk stratification in patients with atrial fibrillation (AF) is uncertain. OBJECTIVE: To examine the accuracy of AF thromboembolic risk models (the CHADS2, CHA2DS2-VASc, and R2CHADS2 scores) for predicting 30-day stroke and/or all-cause mortality after non-cardiac surgery in patients with preoperative AF, and to compare these risk scores with the Revised Cardiac Risk Index (RCRI). PATIENTS/METHODS: A multicentre (8 countries, 2007-2011) prospective cohort study of patients ≥ 45 years of age undergoing inpatient non-cardiac surgery, who were followed until 30 days after surgery. We calculated c-statistics for each risk prediction model and net reclassification improvements (NRIs) compared with the RCRI. RESULTS: The 961 patients with preoperative AF were at higher risk of any cardiovascular event in the 30 days postoperatively compared with the 13 001 patients without AF: 26.6% vs. 9.0%; adjusted odds ratio, 1.58; 95% confidence interval [CI], 1.33-1.88. All thromboembolic risk scores predicted postoperative death just as well as the RCRI (with c-indices between 0.67 and 0.72). Compared with the RCRI (which had a c-index of 0.64 for 30-day stroke/death), the CHADS2 (c-index, 0.67; NRI, 0.31; 95% CI, 0.02-0.61) significantly improved postoperative stroke/mortality risk prediction, largely due to improved discrimination of patients who did not subsequently have an event. CONCLUSIONS: In AF patients, the three thromboembolic risk scores performed similarly to the RCRI in predicting death within 30 days and the CHADS2 score was the best predictor of postoperative stroke/death regardless of type of surgery.

Health professionals' and service users' perspectives of shared care for monitoring wet age-related macular degeneration: a qualitative study alongside the ECHoES trial.

OBJECTIVES: To explore the views of eye health professionals and service users on shared community and hospital care for wet or neovascular age-related macular degeneration (nAMD). METHOD: Using maximum variation sampling, 5 focus groups and 10 interviews were conducted with 23 service users and 24 eye health professionals from across the UK (consisting of 8 optometrists, 6 ophthalmologists, 6 commissioners, 2 public health representatives and 2 clinical eye care advisors to local Clinical Commissioning Groups). Data were transcribed verbatim and analysed thematically using constant comparative techniques derived from grounded theory methodology. RESULTS: The needs and preferences of those with nAMD appear to be at odds with the current service being provided. There was enthusiasm among health professionals and service users about the possibility of shared care for nAMD as it was felt to have the potential to relieve hospital eye service burden and represent a more patient-centred option, but there were a number of perceived barriers to implementation. Some service users and ophthalmologists voiced concerns about optometrist competency and the potential for delays with referrals to secondary care if stable nAMD became active again. The health professionals were divided as to whether shared care was financially more efficient than the current model of care. Specialist training for optometrists, under the supervision of ophthalmologists, was deemed to be the most effective method of training and was perceived to have the potential to improve the communication and trust that shared care would require. CONCLUSIONS: While shared care is perceived to represent a promising model of nAMD care, voiced concerns suggest that there would need to be greater collaboration between ophthalmology and optometry, in terms of interprofessional trust and communication. TRIAL REGISTRATION NUMBER: ISRCTN07479761.

Clinical experience of scoring criteria for Familial Hypercholesterolaemia (FH) genetic testing in Wales.

BACKGROUND/OBJECTIVE: Familial Hypercholesterolaemia (FH) is caused by mutations in genes of the Low Density Lipoprotein (LDL) receptor pathway. A definitive diagnosis of FH can be made by the demonstration of a pathogenic mutation. The Wales FH service has developed scoring criteria to guide selection of patients for DNA testing, for those referred to clinics with hypercholesterolaemia. The criteria are based on a modification of the Dutch Lipid Clinic scoring criteria and utilise a combination of lipid values, physical signs, personal and family history of premature cardiovascular disease. They are intended to provide clinical guidance and enable resources to be targeted in a cost effective manner. METHODS: 623 patients who presented to lipid clinics across Wales had DNA testing following application of these criteria. RESULTS: The proportion of patients with a pathogenic mutation ranged from 4% in those scoring 5 or less up to 85% in those scoring 15 or more. LDL-cholesterol was the strongest discriminatory factor. Scores gained from physical signs, family history, coronary heart disease, and triglycerides also showed a gradient in mutation pick-up rate according to the score. CONCLUSION: These criteria provide a useful tool to guide selection of patients for DNA testing when applied by health professionals who have clinical experience of FH.

Reaction kinetics and targeting to cellular glutathione S-transferase of the glutathione peroxidase mimetic PhSeZnCl and its D,L-polylactide microparticle formulation.

Catalytic properties and cellular effects of the glutathione peroxidase (GPx)-mimetic compound PhSeZnCl or its d,l-lactide polymer microencapsulation form (M-PhSeZnCl) were investigated and compared with the prototypical Se-organic compounds ebselen and diselenide (PhSe)2. PhSeZnCl was confirmed to catalyze the ping-pong reaction of GPx with higher Vmax than ebselen and (PhSe)2, but the catalytic efficiency calculated for the cosubstrates glutathione (GSH) and H2O2, and particularly the high reactivity against thiols (lowest KM for GSH in the series of test molecules), suggested poor biological applicability of PhSeZnCl as a GPx mimetic. Cytotoxicity of PhSeZnCl was demonstrated in various cancer cell lines via increased reactive oxygen species (ROS) generation, depletion of intracellular thiols, and induction of apoptosis. Experiments carried out in GSH S-transferase P (GSTP)-overexpressing K562 human erythroleukemia cells and in GSTP1-1-knockout murine embryonic fibroblasts (MEFs) demonstrated that this cytosolic enzyme represents a preferential target of the redox disturbances produced by this Se-compound with a key role in controlling H2O2 generation and the perturbation of stress/survival kinase signaling. Microencapsulation was adopted as a strategy to control the thiol reactivity and oxidative stress effects of PhSeZnCl, then assessing applications alternative to anticancer. The uptake of this "depowered" GPx-mimetic formulation, which occurred through an endocytosis-like mechanism, resulted in a marked reduction of cytotoxicity. In MCF-7 cells transfected with different allelic variants of GSTP, M-PhSeZnCl lowered the burst of cellular ROS induced by the exposure to extracellular H2O2, and the extent of this effect changed between the GSTP variants. Microencapsulation is a straightforward strategy to mitigate the toxicity of thiol-reactive Se-organic drugs that enhanced the antioxidant and cellular protective effects of PhSeZnCl. A mechanistic linkage of these effects with the expression pattern and signaling properties of GSTP . This has overcome the GPx-mimetic paradigm proposed for Se-organic drugs with a more pragmatic concept of GSTP signaling modulators.

Segmentation and classification of capnograms: application in respiratory variability analysis.

Variability analysis of respiratory waveforms has been shown to provide key insights into respiratory physiology and has been used successfully to predict clinical outcomes. The current standard for quality assessment of the capnogram signal relies on a visual analysis performed by an expert in order to identify waveform artifacts. Automated processing of capnograms is desirable in order to extract clinically useful features over extended periods of time in a patient monitoring environment. However, the proper interpretation of capnogram derived features depends upon the quality of the underlying waveform. In addition, the comparison of capnogram datasets across studies requires a more practical approach than a visual analysis and selection of high-quality breath data. This paper describes a system that automatically extracts breath-by-breath features from capnograms and estimates the quality of individual breaths derived from them. Segmented capnogram breaths were presented to expert annotators, who labeled the individual physiological breaths into normal and multiple abnormal breath types. All abnormal breath types were aggregated into the abnormal class for the purpose of this manuscript, with respiratory variability analysis as the end-application. A database of 11,526 breaths from over 300 patients was created, comprising around 35% abnormal breaths. Several simple classifiers were trained through a stratified repeated ten-fold cross-validation and tested on an unseen portion of the labeled breath database, using a subset of 15 features derived from each breath curve. Decision Tree, K-Nearest Neighbors (KNN) and Naive Bayes classifiers were close in terms of performance (AUC of 90%, 89% and 88% respectively), while using 7, 4 and 5 breath features, respectively. When compared to airflow derived timings, the 95% confidence interval on the mean difference in interbreath intervals was ± 0.18 s. This breath classification system provides a fast and robust pre-processing of continuous respiratory waveforms, thereby ensuring reliable variability analysis of breath-by-breath parameter time series.

Protein Disulfide Isomerase Superfamily in Disease and the Regulation of Apoptosis.

Cellular homeostasis requires the balance of a multitude of signaling cascades that are contingent upon the essential proteins being properly synthesized, folded and delivered to appropriate subcellular locations. In eukaryotic cells the endoplasmic reticulum (ER) is a specialized organelle that is the central site of synthesis and folding of secretory, membrane and a number of organelletargeted proteins. The integrity of protein folding is enabled by the presence of ATP, Ca++, molecular chaperones, as well as an oxidizing redox environment. The imbalance between the load and capacity of protein folding results in a cellular condition known as ER stress. Failure of these pathways to restore ER homeostasis results in the activation of apoptotic pathways. Protein disulfide isomerases (PDI) compose a superfamily of oxidoreductases that have diverse sequences and are localized in the ER, nucleus, cytosol, mitochondria and cell membrane. The PDI superfamily has multiple functions including, acting as molecular chaperones, protein-binding partners, and hormone reservoirs. Recently, PDI family members have been implicated in the regulation of apoptotic signaling events. The complexities underlying the molecular mechanisms that define the switch from pro-survival to pro-death response are evidenced by recent studies that reveal the roles of specific chaperone proteins as integration points in signaling pathways that determine cell fate. The following review discusses the dual role of PDI in cell death and survival during ER stress.

A red tide of Alexandrium fundyense in the Gulf of Maine.

In early July 2009, an unusually high concentration of the toxic dinoflagellate Alexandrium fundyense occurred in the western Gulf of Maine, causing surface waters to appear reddish brown to the human eye. The discolored water appeared to be the southern terminus of a large-scale event that caused shellfish toxicity along the entire coast of Maine to the Canadian border. Rapid-response shipboard sampling efforts together with satellite data suggest the water discoloration in the western Gulf of Maine was a highly ephemeral feature of less than two weeks in duration. Flow cytometric analysis of surface samples from the red water indicated the population was undergoing sexual reproduction. Cyst fluxes downstream of the discolored water were the highest ever measured in the Gulf of Maine, and a large deposit of new cysts was observed that fall. Although the mechanisms causing this event remain unknown, its timing coincided with an anomalous period of downwelling-favorable winds that could have played a role in aggregating upward-swimming cells. Regardless of the underlying causes, this event highlights the importance of short-term episodic phenomena on regional population dynamics of A. fundyense.

Georges Bank: a leaky incubator of Alexandrium fundyense blooms.

A series of oceanographic surveys on Georges Bank document variability of populations of the toxic dinoflagellate Alexandrium fundyense on time scales ranging from synoptic to seasonal to interannual. Blooms of A. fundyense on Georges Bank can reach concentrations on the order of 104 cells l-1, and are generally bank-wide in extent. Georges Bank populations of A. fundyense appear to be quasi-independent of those in the adjacent coastal Gulf of Maine, insofar as they occupy a hydrographic niche that is colder and saltier than their coastal counterparts. In contrast to coastal populations that rely on abundant resting cysts for bloom initiation, very few cysts are present in the sediments on Georges Bank. Bloom dynamics must therefore be largely controlled by the balance between growth and mortality processes, which are at present largely unknown for this population. Based on correlations between cell abundance and nutrient distributions, ammonium appears to be an important source of nitrogen for A. fundyense blooms on Georges Bank.

Peroxiredoxin VI oxidation in cerebrospinal fluid correlates with traumatic brain injury outcome.

Traumatic brain injury (TBI) patients would benefit from the identification of reliable biomarkers to predict outcomes and treatment strategies. In our study, cerebrospinal fluid (CSF) from patients with severe TBI was evaluated for oxidant stress-mediated damage progression after hospital admission and subsequent ventriculostomy placement. Interestingly, substantial levels of peroxiredoxin VI (Prdx6), a major antioxidant enzyme normally found in astrocytes, were detected in CSF from control and TBI patients and were not associated with blood contamination. Functionally, Prdx6 and its associated binding partner glutathione S-transferase Pi (GSTP1-1, also detected in CSF) act in tandem to detoxify lipid peroxidation damage to membranes. We found Prdx6 was fully active in CSF of control patients but becomes significantly inactivated (oxidized) in TBI. Furthermore, significant and progressive oxidation of "buried" protein thiols in CSF of TBI patients (compared to those of nontrauma controls) was detected over a 24-h period after hospital admission, with increased oxidation correlating with severity of trauma. Conversely, recovery of Prdx6 activity after 24h indicated more favorable patient outcome. Not only is this the first report of an extracellular form of Prdx6 but also the first report of its detection at a substantial level in CSF. Taken together, our data suggest a meaningful correlation between TBI-initiated oxidation of Prdx6, its specific phospholipid hydroperoxide peroxidase activity, and severity of trauma outcome. Consequently, we propose that Prdx6 redox status detection has the potential to be a biomarker for TBI outcome and a future indicator of therapeutic efficacy.

Preliminary study of video imaging of blood vessels in tissues lining the gingival sulcus in periodontally healthy individuals.

BACKGROUND AND OBJECTIVE: The aim of this study was to obtain in vivo images of the microcirculation in tissues lining the gingival crevice in periodontally healthy volunteers and to assess the repeatability of the parameters measured. MATERIAL AND METHODS: Video microscopy images of the microcirculation of tissues lining the gingival crevice were obtained from 20 periodontally healthy volunteers. Images were obtained with a single 1 mm diameter 1 pitch gradient index lens with a high numerical aperture and with a plain glass lens and illumination with a green 525 nm light-emitting diode and recorded using a video microscope. RESULTS: The morphological features of the vessels (including vessel diameter, vessel density, loops, branches, dilated vessels) were similar to those described previously in other mammals. The Kappa values for the assessment of morphology of the vessels using the gradient index lens range from 0.83 for branching to 0.91 for dilated and using the glass lens 0.47 for branching and 0.38 for dilated. CONCLUSIONS: This novel system allowed for a consistent and repeatable assessment of the gingival microvasculature. However, there was some evidence of possible pressure artefacts in those cases where the measurements of separation between vessels exceeded 150 µm.

Nutrients and water masses in the Gulf of Maine - Georges Bank region: Variability and importance to blooms of the toxic dinoflagellate Alexandrium fundyense.

We report here the results of ten oceanographic survey cruises carried out in the Gulf of Maine - Georges Bank region of the Northwest Atlantic during the late spring to summer period in 2007, 2008 and 2010, for which we examine and characterize relationships among dissolved inorganic nutrient fields, water mass dynamics and cell densities of the toxic dinoflagellate Alexandrium fundyense. Nutrients are supplied to continental shelf waters of the Gulf of Maine - Georges Bank region by inflows of deep offshore water masses; once in the Gulf they are transported with the residual circulation and mix with surface waters, both in the Gulf and on the Bank. Those fluxes of offshore water masses and their nutrient loads are the major source of nutrients for phytoplankton production in the region, including annual blooms of A. fundyense in the Gulf and on Georges Bank. This much is already known. We suggest here that the locations and magnitude of A. fundyense blooms are controlled in part by variable nutrient fluxes to the interior Gulf of Maine from offshore, and, those interior Gulf of Maine waters are, in turn, the main nutrient source to Georges Bank, which are brought onto the Bank by tidal pumping on the Northern Flank. We present evidence that nitrate is the initial form of nitrogenous nutrient for A. fundyense blooms, but it is quickly depleted to limiting concentrations of less than 0.5 µM, at which time continued growth and maintenance of the population is likely fueled by recycled ammonium. We also show that phosphate may be the limiting nutrient over much of Georges Bank in summer, allowing recycled ammonium concentrations to increase. Our temperature-salinity analyses reveal spatial and temporal (seasonal and interannual) variability in the relative proportions of two deep source waters that enter the Gulf of Maine at depth through the Northeast Channel: Warm Slope Water (WSW) and Labrador Slope Water (LSW). Those two source waters are known to vary in their nutrient loads, with nitrate concentrations about 50% higher in WSW than LSW, for example, and as such the proportions of these two water masses to one another are important determinants of the overall nutrient loads in the interior Gulf. In addition to these deep slope water fluxes, we show evidence here of episodic fluxes of relatively fresh and low-nutrient shelf waters from the Nova Scotian Shelf, which enter the Gulf in pulses at depths between the surface and approximately 150 m, displacing deep slope waters, and consequently they significantly dilute the Gulf's interior waters, reducing nutrient concentrations and, in turn, affect the magnitude of A. fundyense blooms.

Allelic variants of glutathione S-transferase P1-1 differentially mediate the peroxidase function of peroxiredoxin VI and alter membrane lipid peroxidation.

The dual-functioning antioxidant enzyme peroxiredoxin VI (Prdx6) detoxifies lipid peroxides particularly in biological membranes, and its peroxidase function is activated by glutathione S-transferase Pi (GSTP). The GSTP gene is polymorphic in humans, with the wild-type GSTP1-1A (Ile105, Ala114) and three variants: GSTP1-1B (Ile105Val, Ala114), GSTP1-1C (Ile105Val, Ala114Val), and GSTP1-1D (Ile105, Ala114Val). The focus of this study was to determine the influence of these polymorphisms on Prdx6 peroxidase function. Using extracellular generation of OH radicals and fluorescence (DPPP dye) detection, we found a fast (~300 s) onset of lipid peroxidation in membranes of MCF-7 cells transfected with a catalytically inactive Y7F mutant of GSTP1-1 and either GSTP1-1B or GSTP1-1D. However, this effect was not detected in cells expressing either GSTP1-1A or GSTP1-1C. Imaging of DPPP-labeled MCF-7 cells showed fluorescence localized in the plasma membrane, but intensity was substantially diminished in the GSTP1-1A- and GSTP1-1C-expressing cells. Moreover, in the Y7F mutant of GSTP1-1A-, GSTP1-1B-, and GST1-1D-expressing cells ()OH generation resulted (after 36 h) in plasma membrane-permeability-related cell death, whereas GSTP1-1A- and GSTP1-1C-expressing cells had significantly better survival. We used FRET analyses to measure in vitro binding of purified GSTP1-1 allelic variant proteins to purified recombinant Prdx6. The affinities for Prdx6 binding to GSH-loaded GSTP1-1's either mirrored their observed peroxidase activities (using phospholipid hydroperoxide as a substrate), GSTP1-1A>GSTP1-1C (K(D)=51.0 vs 57.0 nM), or corresponded to inactivation, GSTP1-1B (GSTP1-1D) (K(D)=101.0 (94.0) nM). In silico modeling of the GSTP1-1-Prdx6 heterodimer revealed that the sites of GSTP1-1 polymorphism (Ile105 and Ala114) are in close proximity to the binding interface. Thus, there is a hierarchy of effectiveness for polymorphic variants of GSTP1-1 to regulate Prdx6 peroxidase function, a feature that may influence human population susceptibilities to oxidant stress.