Subcellular proteomics for determining iron-limited remodeling of plastids in the model diatom Thalassiosira pseudonana (Bacillariophyta).

Diatoms are important primary producers in the world's oceans, yet their growth is constrained in large regions by low bioavailable iron (Fe). Low-Fe stress-induced limitation of primary production is due to requirements for Fe in components of essential metabolic pathways including photosynthesis and other chloroplast plastid functions. Studies have shown that under low-Fe stress, diatoms alter plastid-specific processes, including components of electron transport. These physiological changes suggest changes of protein content and in protein abundances within the diatom plastid. While in silico predictions provide putative information on plastid-localized proteins, knowledge of diatom plastid proteins remains limited in comparison to well-studied model photosynthetic organisms. To address this, we employed shotgun proteomics to investigate the proteome of subcellular plastid-enriched fractions from Thalassiosira pseudonana to gain a better understanding of how the plastid proteome is remodeled in response to Fe limitation. Using mass spectrometry-based peptide identification and quantification, we analyzed T. pseudonana grown under Fe-replete and -limiting conditions. Through these analyses, we inferred the relative quantities of each protein, revealing that Fe limitation regulates major metabolic pathways in the plastid, including the Calvin cycle. Additionally, we observed changes in the expression of light-harvesting proteins. In silico localization predictions of proteins identified in this plastid-enriched proteome allowed for an in-depth comparison of theoretical versus observed plastid-localization, providing evidence for the potential of additional protein import pathways into the diatom plastid.

Alexandrium on the Alaskan Beaufort Sea shelf: Impact of upwelling in a warming Arctic.

The harmful algal genus Alexandrium has characteristically been found in temperate and subtropical regions; however recent evidence suggests global warming may be expanding its range into high latitude waters. Alexandrium cysts have previously been documented in the Chukchi Sea and we hypothesize that Alexandrium may be expanding further into the Arctic due to distribution by the Beaufort shelfbreak jet. Here we document the presence of Alexandrium catenella along the Alaskan Beaufort Sea shelf, marking an expansion of its known range. The observations of A. catenella were made using three different methods: FlowCAM imaging, 18S eukaryotic sequencing, and real-time quantitative PCR. Four occupations of a shelf/slope transect spanned the evolution of a strong wind-driven upwelling event over a 5-day period. A nearby mooring provided the physical context for the event, revealing that enhanced easterly winds reversed the Beaufort shelfbreak jet to the west and induced upwelling of colder, denser water onto the outer shelf. A. catenella sequences dominated the surface phytoplankton community at the onset of the upwelling event. This signal vanished during and after the event, likely due to a combination of alongstream advection, cross-stream advection, and wind mixing. These results suggest contrasting physical processes that are both subject to global warming amplification, delivery of warm waters via the Beaufort shelfbreak jet and upwelling, may control the proliferation of this potential harmful alga into the Arctic.

Presynaptic inhibition decreases when estrogen level rises.

The objective was to determine estrogen's influence on control of a skeletal muscle through measurements of motorneuron excitability (H:M ratio) and presynaptic inhibition (PI). Estrogen serum concentrations were measured at menses and ovulation of female subjects and compared to male controls. Data were analyzed from 12 women and 13 men reporting no history of knee ligament injury. Women reported regular menstrual cycles and no hormone-based contraceptive use for the previous year. Women were tested at menses (Time1) and ovulation (Time2). Men were tested twice, approximately 14 days apart. Analysis indicated no difference in the H:M ratio between the sexes at either time point. A significant difference for the sexes was detected in the magnitude of estrogen change (∆EST) between observations. At Time1, the male and female estrogen concentrations were not different; however, they were different at Time2, primarily due to the large rise observed in the women. A significant difference between the sexes was also seen in the magnitude of change for PI (∆PI) between observations. As with EST, the levels of PI between the sexes at Time1 were not different; however, a difference existed at Time 2. Estrogen interacts with GABA at several nervous system locations affecting inhibition of synaptic transmission. This is the first study to investigate changes in PI of a skeletal muscle between times of low and high estrogen. Improving the understanding of estrogen's influence on skeletal muscles may provide answers to why noncontact anterior cruciate ligament injuries of the knee occur more frequently in women.

Modelling the effect of hydration on skin conductivity.

BACKGROUND: Electrical signals are recorded from and sent into the body via the skin in a number of applications. In practice, skin is often hydrated with liquids having different conductivities so a model was produced in order to determine the relationship between skin impedance and conductivity. METHODS: A model representing the skin was subjected to a variety of electrical signals. The parts of the model representing the stratum corneum were given different conductivities to represent different levels of hydration. RESULTS: The overall impedance and conductivity of the cells did not vary at frequencies below 40 kHz. Above 40 kHz, levels of increased conductivity caused the overall impedance to decrease. CONCLUSION: The variation in impedance with conductivity between 5 and 50 mSm-1 can be modelled quadratically while variation in impedance with conductivity between 5 and 5000 mSm-1 can be modelled with a double exponential decay.

Preferential depletion of zinc within Costa Rica upwelling dome creates conditions for zinc co-limitation of primary production.

The Costa Rica Dome (CRD) is a wind-driven feature characterized by high primary production and an unusual cyanobacterial bloom in surface waters. It is not clear whether this bloom arises from top-down or bottom-up processes. Several studies have argued that trace metal geochemistry within the CRD contributes to the composition of the phytoplankton assemblages, since cyanobacteria and eukaryotic phytoplankton have different transition metal requirements. Here, we report that total dissolved zinc (Zn) is significantly depleted relative to phosphate (P) and silicate (Si) within the upper water column of the CRD compared with other oceanic systems, and this may create conditions favorable for cyanobacteria, which have lower Zn requirements than their eukaryotic competitors. Shipboard grow-out experiments revealed that while Si was a limiting factor under our experimental conditions, additions of Si and either iron (Fe) or Zn led to higher biomass than Si additions alone. The addition of Fe and Zn alone did not lead to significant enhancements. Our results suggest that the depletion of Zn relative to P in upwelled waters may create conditions in the near-surface waters that favor phytoplankton with low Zn requirements, including cyanobacteria.

Antagonism of serotonin receptor 1B decreases viability and promotes apoptosis in the COS canine osteosarcoma cell line.

Serotonin receptor 1B (5HTR1B) traditionally exhibits anti-proliferative activity in osteoblasts. We examined the expression and function of 5HTR1B in the COS canine osteosarcoma cell line and normal canine osteoblasts. Equal levels of 5HTR1B gene and protein expression were found between normal and malignant osteoblasts. Treatment with serotonin enhanced viability of osteosarcoma cells but not normal osteoblasts. Challenge with the 5HTR1B agonist anpirtoline caused no change in cell viability. Rather incubation with the specific receptor antagonist SB224289 caused reduction in osteoblast viability, with this effect more substantial in osteosarcoma cells. Investigation of this inhibitory activity showed 5HTR1B antagonism induces apoptosis in malignant cells. Evaluation of phosphorylated levels of CREB and ERK, transcriptional regulators associated with serotonin receptor signalling in osteoblasts, revealed aberrant 5HTR1B signalling in COS. Our results confirm the presence of 5HTR1B in a canine osteosarcoma cell line and highlight this receptor as a possible novel therapeutic target.

Recovering ultraclean lignins of controlled molecular weight from Kraft black-liquor lignins.

By operating in a region of liquid-liquid equilibrium, hot acetic acid-water mixtures can be used to simultaneously clean, fractionate, and solvate Kraft black-liquor lignins. Lignin-rich liquid phases of controlled molecular weight with key metals contents reduced to <50 ppm are obtained without a washing step.

Genetic indicators of iron limitation in wild populations of Thalassiosira oceanica from the northeast Pacific Ocean.

Assessing the iron (Fe) nutritional status of natural diatom populations has proven challenging as physiological and molecular responses can differ in diatoms of the same genus. We evaluated expression of genes encoding flavodoxin (FLDA1) and an Fe-starvation induced protein (ISIP3) as indicators of Fe limitation in the marine diatom Thalassiosira oceanica. The specificity of the response to Fe limitation was tested in cultures grown under Fe- and macronutrient-deficient conditions, as well as throughout the diurnal light cycle. Both genes showed a robust and specific response to Fe limitation in laboratory cultures and were detected in small volume samples collected from the northeast Pacific, demonstrating the sensitivity of this method. Overall, FLDA1 and ISIP3 expression was inversely related to Fe concentrations and offered insight into the Fe nutritional health of T. oceanica in the field. As T. oceanica is a species tolerant to low Fe, indications of Fe limitation in T. oceanica populations may serve as a proxy for severe Fe stress in the overall diatom community. At two shallow coastal locations, FLD1A and ISIP3 expression revealed Fe stress in areas where dissolved Fe concentrations were high, demonstrating that this approach may be powerful for identifying regions where Fe supply may not be biologically available.

A new Monte Carlo program for simulating light transport through Port Wine Stain skin.

A new Monte Carlo program is presented for simulating light transport through clinically normal skin and skin containing Port Wine Stain (PWS) vessels. The program consists of an eight-layer mathematical skin model constructed from optical coefficients described previously. A simulation including diffuse illumination at the surface and subsequent light transport through the model is carried out using a radiative transfer theory ray-tracing technique. Total reflectance values over 39 wavelengths are scored by the addition of simulated light returning to the surface within a specified region and surface reflections (calculated using Fresnel's equations). These reflectance values are compared to measurements from individual participants, and characteristics of the model are adjusted until adequate agreement is produced between simulated and measured skin reflectance curves. The absorption and scattering coefficients of the epidermis are adjusted through changes in the simulated concentrations and mean diameters of epidermal melanosomes to reproduce non-lesional skin colour. Pseudo-cylindrical horizontal vessels are added to the skin model, and their simulated mean depths, diameters and number densities are adjusted to reproduce measured PWS skin colour. Accurate reproductions of colour measurement data are produced by the program, resulting in realistic predictions of melanin and PWS blood vessel parameters. Using a modest personal computer, the simulation currently requires an average of five and a half days to complete.

Thalassiosira spp. community composition shifts in response to chemical and physical forcing in the northeast Pacific Ocean.

Diatoms are genetically diverse unicellular photosynthetic eukaryotes that are key primary producers in the ocean. Many of the over 100 extant diatom species in the cosmopolitan genus Thalassiosira are difficult to distinguish in mixed populations using light microscopy. Here, we examine shifts in Thalassiosira spp. composition along a coastal to open ocean transect that encountered a 3-month-old Haida eddy in the northeast Pacific Ocean. To quantify shifts in Thalassiosira species composition, we developed a targeted automated ribosomal intergenic spacer analysis (ARISA) method to identify Thalassiosira spp. in environmental samples. As many specific fragment lengths are indicative of individual Thalassiosira spp., the ARISA method is a useful screening tool to identify changes in the relative abundance and distribution of specific species. The method also enabled us to assess changes in Thalassiosira community composition in response to chemical and physical forcing. Thalassiosira spp. community composition in the core of a 3-month-old Haida eddy remained largely (>80%) similar over a 2-week period, despite moving 24 km southwestward. Shifts in Thalassiosira species correlated with changes in dissolved iron (Fe) and temperature throughout the sampling period. Simultaneously tracking community composition and relative abundance of Thalassiosira species within the physical and chemical context they occurred allowed us to identify quantitative linkages between environmental conditions and community response.

Measuring scapular kinematics during arm lowering using the acromion marker cluster.

The aim of the present study was to examine the acromion marker cluster (AMC) method of measuring scapular kinematics during the arm lowering, eccentric, phase. Twenty six participants completed arm elevation and lowering in the sagittal, frontal and scapular plane. The participants held their arm at 30° increments while the orientation of the scapula was recorded using an AMC and a scapular locator (SL). There were no significant differences between the AMC and SL during the lowering phase for sagittal and scapular plane arm movements. The AMC significantly underestimated upward rotation (max RMSE = 6.0°), and significantly overestimated posterior tilt (max RMSE = 7.2°) during arm lowering in the frontal plane. The reported root mean square errors, however, were within the ranges observed during the elevation phase and reported in previous literature. The AMC therefore provides a reasonable description of scapular kinematics during the arm lowering phase.

Oestrogen induces rhythmic expression of the Kisspeptin-1 receptor GPR54 in hypothalamic gonadotrophin-releasing hormone-secreting GT1-7 cells.

Oestrogen-stimulated preovulatory gonadotrophin surges are temporally regulated in a way that remains not fully understood. Mammalian ovulation requires surges of gonadotrophin-releasing hormone (GnRH), released from specialised neurones in the hypothalamus. Surge regulation is mediated by ovarian oestrogen (17 ß-oestradiol; E(2) ) feedback-acting as a negative signal until the early afternoon of the pro-oestrous phase, at which point it stimulates robust increases in GnRH release. Multiple lines of evidence suggest a role for the circadian clock in surge generation, although the presence of endogenous oscillators in several neuronal populations throughout the mediobasal hypothalamus complicates an elucidation of the underlying mechanisms of circadian regulation. In the present study, we propose that endogenous oscillators within GnRH neurones are modulated by oestrogen to elicit GnRH surge secretion. One mechanism by which this may occur is through the up-regulation of receptors of known stimulators of GnRH, such as kisspeptin's cognate receptor, GPR54. Through analysis of mRNA and protein abundance patterns, we found that high levels of E(2) elicit circadian expression profiles of GPR54 in vitro, and that disruption of endogenous GnRH oscillators of the clock dampens this effect. Additionally, although kisspeptin administration to GT1-7 cells does not result in surge-level secretion, we observed increased GnRH secretion from GT1-7 cells treated with positive feedback levels of E(2) . These results in this in vitro neuronal model system suggest a possible mechanism whereby receptor expression levels, and thus GnRH sensitivity to kisspeptin, may change dramatically over the pro-oestrous day. In this way, elevated ovarian E(2) may increase kisspeptidergic tone at the same time as increasing GnRH neuronal sensitivity to this neuropeptide for maximal surge release.

Making sense of artificial hands.

A review of sensors for artificial hands is presented in terms of their range, specifications and characteristics. There is a growing need for sensors due to the development of prosthetic hands that have multiple degrees of freedom requiring finger coordination into different postures. The sensing of force, position (angle), object-slip and temperature allows for the control of these hands automatically and frees the user from cognitive burden. To make the best possible use of individual sensing elements, future controllers will need to combine data from different types of sensor. They may also have an integral power supply using a small battery or harvest energy from their environment and transmit data wirelessly.

Characterization of putative iron responsive genes as species-specific indicators of iron stress in thalassiosiroid diatoms.

Iron (Fe) availability restricts diatom growth and primary production in large areas of the oceans. It is a challenge to assess the bulk Fe nutritional health of natural diatom populations, since species can differ in their physiological and molecular responses to Fe limitation. We assayed expression of selected genes in diatoms from the Thalassiosira genus to assess their potential utility as species-specific molecular markers to indicate Fe status in natural diatom assemblages. In this study, we compared the expression of the photosynthetic genes encoding ferredoxin (a Fe-requiring protein) and flavodoxin (a Fe-free protein) in culture experiments with Fe replete and Fe stressed Thalassiosira pseudonana (CCMP 1335) isolated from coastal waters and Thalassiosira weissflogii (CCMP 1010) isolated from the open ocean. In T. pseudonana, expression of flavodoxin and ferredoxin genes were not sensitive to Fe status but were found to display diel periodicities. In T. weissflogii, expression of flavodoxin was highly responsive to iron levels and was only detectable when cultures were Fe limited. Flavodoxin genes have been duplicated in most diatoms with available genome data and we show that T. pseudonana has lost its copy related to the Fe-responsive copy in T. weissflogii. We also examined the expression of genes for a putative high affinity, copper (Cu)-dependent Fe uptake system in T. pseudonana. Our results indicate that genes encoding putative Cu transporters, a multi-Cu oxidase, and a Fe reductase are not linked to Fe status. The expression of a second putative Fe reductase increased in Fe limited cultures, but this gene was also highly expressed in Fe replete cultures, indicating it may not be a useful marker in the field. Our findings highlight that Fe metabolism may differ among diatoms even within a genus and show a need to validate responses in different species as part of the development pipeline for genetic markers of Fe status in field populations.

Constant angular velocity of the wrist during the lifting of a sphere.

The primary objective of the experiments was to investigate the wrist motion of a person while they were carrying out a prehensile task from a clinical hand function test. A six-camera movement system was used to observe the wrist motion of 10 participants. A very light sphere and a heavy sphere were used in the experiments to study any mass effects. While seated at a table, a participant moved a sphere over a small obstacle using their dominant hand. The participants were observed to move their wrist at a constant angular velocity. This phenomenon has not been reported previously. Theoretically, the muscles of the wrist provide an impulse of force at the start of the rotation while the forearm maintains a constant vertical force on a sphere. Light-heavy mean differences for the velocities, absolute velocities, angles and times taken showed no significant differences (p = 0.05).

A molecular assessment of the iron stress response in the two phylogenetic clades of Trichodesmium.

Trichodesmium spp. play key roles in global carbon and nitrogen budgets and thus defining what controls their productivity is important for understanding climate change. While iron availability has been shown to be an important chemical factor for controlling both growth and nitrogen fixation rates in Trichodesmium, all culture experiments to date have focused solely on representatives from one clade of Trichodesmium. Genomic sequence analysis determined that the Trichodesmium erythraeum (IMS101) genome contains many of the archetypical genes involved in the prokaryotic iron stress response. Focusing on three of these genes, isiB, idiA and feoB, we found that all three showed an iron stress response in axenic T. erythraeum (IMS101), and their sequences were well conserved across four species in our Trichodesmium culture collection [consisting of two T. erythraeum strains (IMS101 and GBRTRLI101), two Trichodesmium tenue strains (Z-1 and H9-4), Trichodesmium thiebautii and Trichodesmium spiralis]. With clade-specific quantitative PCR (qPCR) primers for one of these genes, isiB, we found that high isiB expression at low Fe levels corresponded to specific reductions in N(2) fixation rates in both major phylogenetic clades of Trichodesmium (the T. erythraeum clade and T. tenue clade). With regard to the two clades, the most significant difference determined was temperature optima, while more subtle differences in growth, N(2) fixation rate and gene expression responses to Fe stress were also observed. However the apparent conservation of the Fe stress response in the Trichodesmium genus suggests that it is an important adaptation for their niche in the oligotrophic ocean.

Shoulder and elbow muscle activity during fully supported trajectory tracking in people who have had a stroke.

An inability to perform tasks involving reaching is a common problem for stroke patients. This paper provides an insight into mechanisms associated with recovery of upper limb function by examining how stroke participants' upper limb muscle activation patterns differ from those of neurologically intact participants, and how they change in response to an intervention. In this study, five chronic stroke participants undertook nine tracking tasks in which trajectory (orientation and length), speed and resistance to movement were varied. During these tasks, EMG signals were recorded from triceps, biceps, anterior deltoid, upper, middle and lower trapezius and pectoralis major. Data collection was performed in sessions both before, and after, an intervention in which participants performed a similar range of tracking tasks with the addition of responsive electrical stimulation applied to their triceps muscle. The intervention consisted of eighteen one hour treatment sessions, with two participants attending an additional seven sessions. During all sessions, each participant's arm was supported by a hinged arm-holder which constrained their hand to move in a two dimensional plane. Analysis of the pre intervention EMG data showed that timing and amplitude of peak EMG activity for all stroke participants differed from neurologically intact participants. Analysis of post intervention EMG data revealed that statistically significant changes in these quantities had occurred towards those of neurologically intact participants.

Surface EMG pattern analysis of the wrist muscles at different speeds of contraction.

The aim of this study is to investigate the characteristics of surface electromyographic signals, particularly in pattern analysis. The data were collected from the wrist muscles (flexor carpi ulnaris and extensor carpi radialis) of 20 healthy participants. The study focuses on the movement of the wrist muscles at different frequencies. Participants were asked to contract their muscles at four different speeds (60, 90 and 120 cycles a minute and maximum speed) during wrist flexion and extension, co-contraction and isometric contraction. In this work, moving approximate entropy, mean absolute value and kurtosis are evaluated from the surface electromyographic signals at the four speeds. Moving approximate entropy and kurtosis analysis show that there are significant differences at three states of contraction; start, middle and end. It is shown that there are more regular data in a surface electromyographic signal at the beginning and end of a muscle contraction with low regularity during the middle part.

Feasibility of iterative learning control mediated by functional electrical stimulation for reaching after stroke.

BACKGROUND: An inability to perform tasks involving reaching is a common problem following stroke. Evidence supports the use of robotic therapy and functional electrical stimulation (FES) to reduce upper limb impairments, but current systems may not encourage maximal voluntary contribution from the participant because assistance is not responsive to performance. OBJECTIVE: This study aimed to investigate whether iterative learning control (ILC) mediated by FES is a feasible intervention in upper limb stroke rehabilitation. METHODS: Five hemiparetic participants with reduced upper limb function who were at least 6 months poststroke were recruited from the community. No participants withdrew. INTERVENTION: Participants undertook supported tracking tasks using 27 different trajectories augmented by responsive FES to their triceps brachii muscle, with their hand movement constrained in a 2-dimensional plane by a robot. Eighteen 1-hour treatment sessions were used with 2 participants receiving an additional 7 treatment sessions. OUTCOME MEASURES: The primary functional outcome measure was the Action Research Arm Test (ARAT). Impairment measures included the upper limb Fugl-Meyer Assessment (FMA), tests of motor control (tracking accuracy), and isometric force. RESULTS: Compliance was excellent and there were no adverse events. Statistically significant improvements were measured (P

Modulation of gonadotrophin-releasing hormone secretion by an endogenous circadian clock.

The mechanisms mediating positive feedback effects of oestradiol on pre-ovulatory gonadotrophin releasing-hormone (GnRH) surge generation in female mammals, although well-explored, are still incompletely understood. In addition to binding to and signalling through classical nuclear receptor-mediated pathways in afferent hypothalamic neurones, recent evidence suggests that ovarian steroids may use membrane-bound receptors or nonclassical signalling pathways to directly influence cell function leading to the generation of GnRH surge secretion. We review recent investigations into the role of the endogenous molecular circadian clock on modulation of GnRH gene expression and neuropeptide secretion, and will explore potential molecular mechanisms by which ovarian steroids may directly induce secretory changes at the level of the GnRH neurone, examining closely whether circadian clock gene oscillations may be involved.