Fiscal Year 2018 National Defense Authorization Act, Section 734, Weapon Systems Line of Inquiry: Overview and Blast Overpressure Tool-A Module for Human Body Blast Wave Exposure for Safer Weapons Training.

INTRODUCTION: Experiences by service members in recent conflicts and training environments illuminate concerns about the possible effects of blast overpressure (BOP) exposure on brain health. Section 734 of the National Defense Authorization Act for Fiscal Year (FY) 2018 (Public Law 115-91) requires that the Secretary of Defense conducts a longitudinal medical study on blast pressure exposure of members of the Armed Forces during combat and training, and the Assistant Secretary of Defense for Health Affairs was assigned responsibility for fulfilling requirements. The study's goal is to improve DoD's understanding of the impact of BOP exposure from weapon systems on service members' brain health and inform policy for risk mitigation, unit readiness, and health care decisions. This article focuses on the activities of the Weapon Systems Line of Inquiry (LOI) and the development of a prototype BOP Tool. MATERIALS AND METHODS: The DoD established the Section 734 Workgroup, which developed a program structure with five LOIs. The Weapon Systems LOI coordinated, collated, and analyzed information on BOP resulting from heavy weapons and blast events to inform strategies, and accounted for emerging research on health effects and performance. Ongoing research was leveraged to develop a BOP Tool as a standalone module and for integration into the Range Managers Toolkit. RESULTS: The effort identified opportunities for the DoD to improve the clarity of communications about BOP exposure, risk, and safety; establish methods to leverage emerging research; and develop a prototype BOP Tool to predict exposure loads when firing heavy weapons in training. CONCLUSIONS: It is recommended that the DoD revises requirements and policy to improve and standardize safety guidance throughout research, development, testing, and evaluation; acquisition; and training. The validated BOP Tool allows users to generate a scenario to predict BOP exposure and allows service members to modify them during planning for safer training.

Mathematical model of mechanobiology of acute and repeated synaptic injury and systemic biomarker kinetics.

Background: Blast induced Traumatic Brain Injury (bTBI) has become a signature casualty of military operations. Recently, military medics observed neurocognitive deficits in servicemen exposed to repeated low level blast (LLB) waves during military heavy weapons training. In spite of significant clinical and preclinical TBI research, current understanding of injury mechanisms and short- and long-term outcomes is limited. Mathematical models of bTBI biomechanics and mechanobiology of sensitive neuro-structures such as synapses may help in better understanding of injury mechanisms and in the development of improved diagnostics and neuroprotective strategies. Methods and results: In this work, we formulated a model of a single synaptic structure integrating the dynamics of the synaptic cell adhesion molecules (CAMs) with the deformation mechanics of the synaptic cleft. The model can resolve time scales ranging from milliseconds during the hyperacute phase of mechanical loading to minutes-hours acute/chronic phase of injury progression/repair. The model was used to simulate the synaptic injury responses caused by repeated blast loads. Conclusion: Our simulations demonstrated the importance of the number of exposures compared to the duration of recovery period between repeated loads on the synaptic injury responses. The paper recognizes current limitations of the model and identifies potential improvements.

Effects of Isolated and Combined Exposure of the Brain and Lungs to a Laser-Induced Shock Wave(s) on Physiological and Neurological Responses in Rats.

Blast-induced traumatic brain injury (bTBI) has been suggested to be caused by direct head exposure and by torso exposure to a shock wave (thoracic hypotheses). It is unclear, however, how torso exposure affects the brain in real time. This study applied a mild-impulse laser-induced shock wave(s) (LISW[s]) only to the brain (Group 1), lungs (Group 2), or to the brain and lungs (Group 3) in rats. Because LISWs are unaccompanied by a dynamic pressure in principle, the effects of acceleration can be excluded, allowing analysis of the pure primary mechanism (the effects of a shock wave). For all rat groups, real-time monitoring of the brain and systemic responses were conducted for up to 1 h post-exposure and motor function assessments for up to seven days post-exposure. As reported previously, brain exposure alone caused cortical spreading depolarization (CSD), followed by long-lasting hypoxemia and oligemia in the cortices (Group 1). It was found that even LISW application only to the lungs caused prolonged hypoxemia and mitochondrial dysfunction in the cortices (Group 2). Importantly, features of CSD and mitochondrial dysfunction were significantly exacerbated by combined exposure (Group 3) compared with those caused by brain exposure alone (Group 1). Motor dysfunction was observed in all exposure groups, but their time courses differed depending on the groups. Rats with brain exposure alone exhibited the most evident motor dysfunction at one day post-exposure, and after that, it did not change much for up to seven days post-exposure. Alternatively, two groups of rats with lung exposure (Group 2 and Group 3) exhibited continuously aggravated motor functions for up to seven days post-exposure, suggesting different mechanisms for motor dysfunction caused by brain exposure and that caused by lung exposure. As for the reported thoracic hypotheses, our observations seem to support the volumetric blood surge and vagovagal reflex. Overall, the results of this study indicate the importance of the torso guard to protect the brain and its function.

A Methodology to Compare Biomechanical Simulations With Clinical Brain Imaging Analysis Utilizing Two Blunt Impact Cases.

According to the US Defense and Veterans Brain Injury Center (DVBIC) and Centers for Disease Control and Prevention (CDC), mild traumatic brain injury (mTBI) is a common form of head injury. Medical imaging data provides clinical insight into tissue damage/injury and injury severity, and helps medical diagnosis. Computational modeling and simulation can predict the biomechanical characteristics of such injury, and are useful for development of protective equipment. Integration of techniques from computational biomechanics with medical data assessment modalities (e.g., magnetic resonance imaging or MRI) has not yet been used to predict injury, support early medical diagnosis, or assess effectiveness of personal protective equipment. This paper presents a methodology to map computational simulations with clinical data for interpreting blunt impact TBI utilizing two clinically different head injury case studies. MRI modalities, such as T1, T2, diffusion-weighted imaging (DWI) and apparent diffusion coefficient (ADC), were used for simulation comparisons. The two clinical cases have been reconstructed using finite element analysis to predict head biomechanics based on medical reports documented by a clinician. The findings are mapped to simulation results using image-based clinical analyses of head impact injuries, and modalities that could capture simulation results have been identified. In case 1, the MRI results showed lesions in the brain with skull indentation, while case 2 had lesions in both coup and contrecoup sides with no skull deformation. Simulation data analyses show that different biomechanical measures and thresholds are needed to explain different blunt impact injury modalities; specifically, strain rate threshold corresponds well with brain injury with skull indentation, while minimum pressure threshold corresponds well with coup-contrecoup injury; and DWI has been found to be the most appropriate modality for MRI data interpretation. As the findings from these two cases are substantiated with additional clinical studies, this methodology can be broadly applied as a tool to support injury assessment in head trauma events and to improve countermeasures (e.g., diagnostics and protective equipment design) to mitigate these injuries.

Fast-Running Tools for Personalized Monitoring of Blast Exposure in Military Training and Operations.

INTRODUCTION: During training and combat operations, military personnel may be exposed to repetitive low-level blast while using explosives to gain entry or by firing heavy weapon systems such as recoilless weapons and high-caliber sniper rifles. This repeated exposure, even within allowable limits, has been associated with cognitive deficits similar to that of accidental and sports concussion such as delayed verbal memory, visual-spatial memory, and executive function. This article presents a novel framework for accurate calculation of the human body blast exposure in military heavy weapon training scenarios using data from the free-field and warfighter wearable pressure sensors. MATERIALS AND METHODS: The CoBi human body model generator tools were used to reconstruct multiple training scenes with different weapon systems. The CoBi Blast tools were used to develop the weapon signature and estimate blast overpressure exposure. The authors have used data from the free-field and wearable pressure sensors to evaluate the framework. RESULTS: Carl-Gustav and 0.50 caliber sniper training scenarios were used to demonstrate and validate the developed framework. These simulations can calculate spatially and temporally resolved blast loads on the whole human body and on specific organs vulnerable to blast loads, such as head, face, and lungs. CONCLUSIONS: This framework has numerous advantages including easier model setup and shorter simulation times. The framework is an important step towards developing an advanced field-applicable technology to monitor low-level blast exposure during heavy weapon military training and combat scenarios.

Confinement-Enhanced Luminescence in Protein-Gold Nanoclusters.

Confinement has profound effects on protein functions. Nanoscale probes for confinement or excluded volume interactions could help us understand how these interactions influence protein functions. This work reports on the increased luminescence of BSA-gold nanoclusters when confined. Confinement of the BSA-gold nanoclusters occurred within reverse micelles (RMs), where the size of the RMs determined the degree of confinement. The confinement-enhanced luminescence is reversible, i.e., the emission returns to its original value following cyclic changes in RM size. Circular dichroism measurements show an increase in alpha-helical character of the BSA-stabilized nanoclusters with confinement, which could provide a mechanism for the increase in luminescence. The alpha-helical character of the native proteins also increases with confinement, suggesting that the protein-nanocluster might sense confinement in an analogous fashion as the proteins. When the RMs approach the size of the protein, the intensity becomes independent of alpha-helical character, suggesting a different mechanism for the luminescence increase.

Human Nontumorigenic Microglia Synthesize Strongly Fluorescent Au/Fe Nanoclusters, Retaining Bioavailability.

The ability for cells to self-synthesize metal-core nanoclusters (mcNCs) offers increased imaging and identification opportunities. To date, much work has been done illustrating the ability for human tumorigenic cell lines to synthesize mcNCs; however, this has not been illustrated for nontumorigenic cell lines. Here, we present the ability for human nontumorigenic microglial cells, which are the major immune cells in the central nervous system, to self-synthesize gold (Au) and iron (Fe) core nanoclusters, following exposures to metallic salts. We also show the ability for cells to internalize presynthesized Au and Fe mcNCs. Cellular fluorescence increased in most exposures and in a dose dependent manner in the case of Au salt. Scanning transmission electron microscopic imaging confirmed the presence of the metal within cells, while transmission electron microscopy images confirmed nanocluster structures and self-synthesis. Interestingly, self-synthesized nanoclusters were of similar size and internal structure as presynthesized mcNCs. Toxicity assessment of both salts and presynthesized NCs illustrated a lack of toxicity from Au salt and presynthesized NCs. However, Fe salt was generally more toxic and stressful to cells at similar concentrations.

Oxidative stress contributes to cerebral metabolomic profile changes in animal model of blast-induced traumatic brain injury.

INTRODUCTION: Blast-induced neurotrauma (BINT) has been recognized as the common mode of traumatic brain injury amongst military and civilian personnel due to an increased insurgent activity domestically and abroad. Previous studies from this laboratory have identified three major pathological events following BINT which include blood brain barrier disruption the earliest event, followed by oxidative stress and neuroinflammation as secondary events occurring a few hours following blast. OBJECTIVES: Our recent studies have also identified an increase in oxidative stress mediated by the activation of superoxide producing enzyme NADPH oxidase (NOX) in different brain regions at varying levels with neurons displaying higher oxidative stress (NOX activation) compared to any other neural cell. Since neurons have higher energy demands in brain and are more prone to oxidative damage, this study evaluated the effect of oxidative stress on blast-blast induced changes in metabolomics profiles in different brain regions. METHODS: Animals were exposed to mild/moderate blast injury (180 kPa) and examined the metabolites of energy metabolism, amino acid metabolism as well as the profiles of plasma membrane metabolites in different brain regions at different time points (24 h, 3 day and 7 day) after blast using 1H NMR spectroscopy. Effect of apocynin, an inhibitor of superoxide producing enzyme NADPH oxidase on cerebral metabalomics profiles was also examined. RESULTS: Several metabolomic profile changes were observed in frontal cortex and hippocampus with concomitant decrease in energy metabolism. In addition, glutamate/glutamine and other amino acid metabolism as well as metabolites involved in plasma membrane integrity were also altered. Hippocampus appears metabolically more vulnerable than the frontal cortex. A post-treatment of animals with apocynin, an inhibitor of NOX activation significantly prevented the changes in metabolite profiles. CONCLUSION: Together these studies indicate that blast injury reduces both cerebral energy and neurotransmitter amino acid metabolism and that oxidative stress contributes to these processes. Thus, strategies aimed at reducing oxidative stress can have a therapeutic benefit in mitigating metabolic changes following BINT.

Biomechanics of Blast TBI With Time-Resolved Consecutive Primary, Secondary, and Tertiary Loads.

Blast-induced traumatic brain injury (bTBI) has become a signature casualty of recent military operations. In spite of significant clinical and preclinical TBI research, current understanding of injury mechanisms and short- and long-term outcomes is limited. Mathematical models of bTBI biomechanics may help in better understanding of injury mechanisms and in the development of improved neuroprotective strategies. Until present, bTBI has been analyzed as a single event of a blast pressure wave propagating through the brain. In many bTBI events, the loads on the body and the head are spatially and temporarily distributed, involving the primary intracranial pressure wave, followed by the head rotation and then by head impact on the ground. In such cases, the brain microstructures may experience time/space distributed (consecutive) damage and recovery events. The paper presents a novel multiscale simulation framework that couples the body/brain scale biomechanics with micro-scale mechanobiology to study the effects of micro-damage to neuro-axonal structures. Our results show that the micro-mechanical responses of neuro-axonal structures occur sequentially in time with "damage" and "relaxation" periods in different parts of the brain. A new integrated computational framework is described coupling the brain-scale biomechanics with micro-mechanical damage to axonal and synaptic structures.

A Single Primary Blast-Induced Traumatic Brain Injury in a Rodent Model Causes Cell-Type Dependent Increase in Nicotinamide Adenine Dinucleotide Phosphate Oxidase Isoforms in Vulnerable Brain Regions.

Blast-induced traumatic brain injury (bTBI) is a leading cause of morbidity in soldiers on the battlefield and in training sites with long-term neurological and psychological pathologies. Previous studies from our laboratory demonstrated activation of oxidative stress pathways after blast injury, but their distribution among different brain regions and their impact on the pathogenesis of bTBI have not been explored. The present study examined the protein expression of two isoforms: nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 and 2 (NOX1, NOX2), corresponding superoxide production, a downstream event of NOX activation, and the extent of lipid peroxidation adducts of 4-hydroxynonenal (4HNE) to a range of proteins. Brain injury was evaluated 4 h after the shock-wave exposure, and immunofluorescence signal quantification was performed in different brain regions. Expression of NOX isoforms displayed a differential increase in various brain regions: in hippocampus and thalamus, there was the highest increase of NOX1, whereas in the frontal cortex, there was the highest increase of NOX2 expression. Cell-specific analysis of changes in NOX expression with respect to corresponding controls revealed that blast resulted in a higher increase of NOX1 and NOX 2 levels in neurons compared with astrocytes and microglia. Blast exposure also resulted in increased superoxide levels in different brain regions, and such changes were reflected in 4HNE protein adduct formation. Collectively, this study demonstrates that primary blast TBI induces upregulation of NADPH oxidase isoforms in different regions of the brain parenchyma and that neurons appear to be at higher risk for oxidative damage compared with other neural cells.

Combination Therapy for Multi-Target Manipulation of Secondary Brain Injury Mechanisms.

Traumatic brain injury (TBI) is a major healthcare problem that affects millions of people worldwide. Despite advances in understanding and developing preventative and treatment strategies using preclinical animal models, clinical trials to date have failed, and a 'magic bullet' for effectively treating TBI-induced damage does not exist. Thus, novel pharmacological strategies to effectively manipulate the complex and heterogeneous pathophysiology of secondary injury mechanisms are needed. Given that goal, this paper discusses the relevance and advantages of combination therapies (COMTs) for 'multi-target manipulation' of the secondary injury cascade by administering multiple drugs to achieve an optimal therapeutic window of opportunity (e.g., temporally broad window) and compares these regimens to monotherapies that manipulate a single target with a single drug at a given time. Furthermore, we posit that integrated mechanistic multiscale models that combine primary injury biomechanics, secondary injury mechanobiology/neurobiology, physiology, pharmacology and mathematical programming techniques could account for vast differences in the biological space and time scales and help to accelerate drug development, to optimize pharmacological COMT protocols and to improve treatment outcomes.

Rapid Development and Characterization of Chromosome Specific Translocation Line of Thinopyrum elongatum with Improved Dough Strength.

The protein content and its type are principal factors affecting wheat (Triticum aestivum) end product quality. Among the wheat proteins, glutenin proteins, especially, high molecular weight glutenin subunits (HMW-GS) are major determinants of processing quality. Wheat and its primary gene pool have limited variation in terms of HMW-GS alleles. Wild relatives of wheat are an important source of genetic variation. For improvement of wheat processing quality its wild relative Thinopyrum elongatum with significant potential was utilized. An attempt was made to replace Th. elongatum chromosome long arm (1EL) carrying HMW-GS genes related to high dough strength with chromosome 1AL of wheat with least or negative effect on dough strength while retaining the chromosomes 1DL and 1BL with a positive effect on bread making quality. To create chromosome specific translocation line [1EL(1AS)], double monosomic of chromosomes 1E and 1A were created and further crossed with different cultivars and homoeologous pairing suppressor mutant line PhI . The primary selection was based upon glutenin and gliadin protein profiles, followed by sequential genomic in situ hybridization (GISH) and fluorescent in situ hybridization (FISH). These steps significantly reduced time, efforts, and economic cost in the generation of translocation line. In order to assess the effect of translocation on wheat quality, background recovery was carried out by backcrossing with recurrent parent for several generations and then selfing while selecting in each generation. Good recovery of parent background indicated the development of almost near isogenic line (NIL). Morphologically also translocation line was similar to recipient cultivar N61 that was further confirmed by seed storage protein profiles, RP-HPLC and scanning electron microscopy. The processing quality characteristics of translocation line (BC4F6) indicated significant improvement in the gluten performance index (GPI), dough mixing properties, dough strength, and extensibility. Our work aims to address the challenge of limited genetic diversity especially at chromosome 1A HMW-GS locus. We report successful development of chromosome 1A specific translocation line of Th. elongatum in wheat with improved dough strength.

Military Blast Injury and Chronic Neurodegeneration: Research Presentations from the 2015 International State-of-the-Science Meeting.

Blast-related traumatic brain injury (TBI) is a signature injury of recent military conflicts, leading to increased Department of Defense (DoD) interest in its potential long-term effects, such as chronic traumatic encephalopathy (CTE). The DoD Blast Injury Research Program Coordinating Office convened the 2015 International State-of-the-Science Meeting to discuss the existing evidence regarding a causal relationship between TBI and CTE. Over the course of the meeting, experts across government, academia, and the sports community presented cutting edge research on the unique pathological characteristics of blast-related TBI, blast-related neurodegenerative mechanisms, risk factors for CTE, potential biomarkers for CTE, and treatment strategies for chronic neurodegeneration. The current paper summarizes these presentations. Although many advances have been made to address these topics, more research is needed to establish the existence of links between the long-term effects of single or multiple blast-related TBI and CTE.

The Biological Basis of Chronic Traumatic Encephalopathy following Blast Injury: A Literature Review.

The United States Department of Defense Blast Injury Research Program Coordinating Office organized the 2015 International State-of-the-Science meeting to explore links between blast-related head injury and the development of chronic traumatic encephalopathy (CTE). Before the meeting, the planning committee examined articles published between 2005 and October 2015 and prepared this literature review, which summarized broadly CTE research and addressed questions about the pathophysiological basis of CTE and its relationship to blast- and nonblast-related head injury. It served to inform participants objectively and help focus meeting discussion on identifying knowledge gaps and priority research areas. CTE is described generally as a progressive neurodegenerative disorder affecting persons exposed to head injury. Affected individuals have been participants primarily in contact sports and military personnel, some of whom were exposed to blast. The symptomatology of CTE overlaps with Alzheimer's disease and includes neurological and cognitive deficits, psychiatric and behavioral problems, and dementia. There are no validated diagnostic criteria, and neuropathological evidence of CTE has come exclusively from autopsy examination of subjects with histories of exposure to head injury. The perivascular accumulation of hyperphosphorylated tau (p-tau) at the depths of cortical sulci is thought to be unique to CTE and has been proposed as a diagnostic requirement, although the contribution of p-tau and other reported pathologies to the development of clinical symptoms of CTE are unknown. The literature on CTE is limited and is focused predominantly on head injuries unrelated to blast exposure (e.g., football players and boxers). In addition, comparative analyses of clinical case reports has been challenging because of small case numbers, selection biases, methodological differences, and lack of matched controls, particularly for blast-exposed individuals. Consequently, the existing literature is not sufficient to determine whether the development of CTE is associated with head injury frequency (e.g., single vs. multiple exposures) or head injury type (e.g., impact, nonimpact, blast-related). Moreover, the incidence and prevalence of CTE in at-risk populations is unknown. Future research priorities should include identifying additional risk factors, pursuing population-based longitudinal studies, and developing the ability to detect and diagnose CTE in living persons using validated criteria.

Validation of Laboratory Animal and Surrogate Human Models in Primary Blast Injury Studies.

Blast-induced neurotrauma has affected more than 300,000 service members. It is important to understand the effect of single and repeated shock-blast wave exposures on the neuropsychological behavior of soldiers, to offer them better protection, diagnostics, and treatment. Preclinical animal models and helmet design studies on human surrogate models have relied on the use of compression gas-driven shock tubes. Traditional shock tubes are so simple that if not carefully designed and operated, the test results can easily introduce detrimental artifacts clouding the conclusions. In this work, we present live-fire test results of an instrumented human surrogate head-neck model and compare with the data obtained in a carefully designed shock tube. We present various features incorporated in the shock tube design that led to better fidelity between live-fire and laboratory shock-blast conditions. The effect of specimen placement, choice of driver gas, pressure and volume of driver, end-plate conditions, and measurement techniques all determine the successful replication of live-fire loading conditions. These parameters become more important when conducting animal testing as the totality of loading will dictate the injury severity and type which ultimately will determine the mechanisms of blast-induced neurotrauma and hence their prevention and treatment strategies.

Computational comparison of a calcium-dependent jellyfish protein (apoaequorin) and calmodulin-cholesterol in short-term memory maintenance.

Memory reconsolidation and maintenance depend on calcium channels and on calcium/calmodulin-dependent kinases regulating protein turnover in the hippocampus. Ingestion of a jellyfish protein, apoaequorin, reportedly protects and/or improves verbal learning in adults and is currently widely advertised for use by the elderly. Apoaequorin is a member of the EF-hand calcium binding family of proteins that includes calmodulin. Calmodulin-1 (148 residues) differs from Apoaequorin (195 residues) in that it contains four rather than three Ca2+-binding sites and three rather than four cholesterol-binding (CRAC, CARC) domains. All three cholesterol-binding CARC domains in calmodulin have a high interaction affinity for cholesterol compared to only two high affinity CARC domains in apoaequorin. Both calmodulin and apoaequorin can form dimers with a potential of eight bound Ca2+ ions and six high affinity-bound cholesterol molecules in calmodulin with six bound Ca2+ ions and a mixed population of eight cholesterols bound to both CARC and CRAC domains in apoaqueorin. MEMSAT-SVM analysis indicates that both calmodulin and apoaqueorin have a pore-lining region. The Peptide-Cutter algorithm predicts that calmodulin-1 contains 11 trypsin-specific cleavage sites (compared to 21 in apoaqueorin), four of which are potentially blocked by cholesterol and three are within the Ca-binding domains and/or the pore-lining region. Three are clustered between the third and fourth Ca2+-binding sites. Only calmodulin pore-lining regions contain Ca2+ binding sites and as dimers may insert into the plasma membrane of neural cells and act as Ca2+ channels. In a dietary supplement, bound cholesterol may protect both apoaequorin and calmodulin from proteolysis in the gut as well as facilitate uptake across the blood-brain barrier. Our results suggest that a physiological calmodulin-cholesterol complex, not cholesterol-free jellyfish protein, may better serve as a dietary supplement to facilitate memory maintenance.

The role of receptor topology in the vitamin D3 uptake and Ca(2+) response systems.

The steroid hormone, vitamin D3, regulates gene transcription via at least two receptors and initiates putative rapid response systems at the plasma membrane. The vitamin D receptor (VDR) binds vitamin D3 and a second receptor, importin-4, imports the VDR-vitamin D3 complex into the nucleus via nuclear pores. Here we present evidence that the Homo sapiens VDR homodimer contains two transmembrane (TM) helices ((327)E - D(342)), two TM "half-helix" ((264)K N(276)), one or more large channels, and 16 cholesterol binding (CRAC/CARC) domains. The importin-4 monomer exhibits 3 pore-lining regions ((226)E - L(251); (768)V - G(783); (876)S - A(891)) and 16 CRAC/CARC domains. The MEMSAT algorithm indicates that VDR and importin-4 may not be restricted to cytoplasm and nucleus. VDR homodimer TM helix-topology predicts insertion into the plasma membrane, with two 84 residue C-terminal regions being extracellular. Similarly, MEMSAT predicts importin-4 insertion into the plasma membrane with 226 residue extracellular N-terminal regions and 96 residue C-terminal extracellular loops; with the pore-lining regions contributing gated Ca(2+) channels. The PoreWalker algorithm indicates that, of the 427 residues in each VDR monomer, 91 line the largest channel, including two vitamin D3 binding sites and residues from both the TM helix and "half-helix". Cholesterol-binding domains also extend into the channel within the ligand binding region. Programmed changes in bound cholesterol may regulate both membrane Ca(2+) response systems and vitamin D3 uptake as well as receptor internalization by the endomembrane system culminating in uptake of the vitamin D3-VDR-importin-4 complex into the nucleus.

Evolution of the α-Subunit of Na/K-ATPase from Paramecium to Homo sapiens: Invariance of Transmembrane Helix Topology.

Na/K-ATPase is a key plasma membrane enzyme involved in cell signaling, volume regulation, and maintenance of electrochemical gradients. The α-subunit, central to these functions, belongs to a large family of P-type ATPases. Differences in transmembrane (TM) helix topology, sequence homology, helix-helix contacts, cell signaling, and protein domains of Na/K-ATPase α-subunit were compared in fungi (Beauveria), unicellular organisms (Paramecia), primitive multicellular organisms (Hydra), and vertebrates (Xenopus, Homo sapiens), and correlated with evolution of physiological functions in the α-subunit. All α-subunits are of similar length, with groupings of four and six helices in the N- and C-terminal regions, respectively. Minimal homology was seen for protein domain patterns in Paramecium and Hydra, with high correlation between Hydra and vertebrates. Paramecium α-subunits display extensive disorder, with minimal helix contacts. Increases in helix contacts in Hydra approached vertebrates. Protein motifs known to be associated with membrane lipid rafts and cell signaling reveal significant positional shifts between Paramecium and Hydra vulgaris, indicating that regional membrane fluidity changes occur during evolution. Putative steroid binding sites overlapping TM-3 occurred in all species. Sites associated with G-protein-receptor stimulation occur both in vertebrates and amphibia but not in Hydra or Paramecia. The C-terminus moiety "KETYY," necessary for the Na(+) activation of pump phosphorylation, is not present in unicellular species indicating the absence of classical Na(+)/K(+)-pumps. The basic protein topology evolved earliest, followed by increases in protein domains and ordered helical arrays, correlated with appearance of α-subunit regions known to involve cell signaling, membrane recycling, and ion channel formation.

Herbicide options for effective weed management in dry direct-seeded rice under scented rice-wheat rotation of western Indo-Gangetic Plains.

Farmers' participatory field trials were conducted at Madhuban, and Taraori, the two participatory experimental sites/locations of the Cereal Systems Initiative for South Asia (CSISA), a collaborative project of IRRI and CIMMYT in Karnal district of Haryana, India, during Kharif (wet season) 2010 and 2011. This research aimed to evaluate preemergence (PRE) and postemergence (POST) herbicides for providing feasible and economically viable weed management options to farmers for predominant scented rice varieties. Treatments with pendimethalin PRE fb bispyribac-sodium + azimsulfuron POST had lower weed biomass at 45 days after sowing (DAS). At Madhuban, highest grain yield of scented basmati rice (3.43 t ha-1) was recorded with the sequential application of pendimethalin PRE fb bispyribac-sodium + azimsulfuron POST. However, at Taraori, yields were similar with pendimethalin or oxadiargyl PRE fb bispyribac-sodium and/or azimsulfuron POST. Applying oxadiargyl by mixing with sand onto flooded field was less effective than spray applications in non-flooded field. The benefit-cost ratio of rice crop was higher with herbicide treatments at both sites as compared with the non-treated weed-free check except single PRE and POST applications and sequential application of oxadiargyl PRE fb oxadiargyl PRE. In a separate experiment conducted at Nagla and Taraori sites, scented rice cultivars' ('CSR 30' and 'Pusa 1121') tolerance to three rates of azimsulfuron (15, 25, and 35 g ai ha-1) was evaluated over two years (2010 and 2011). CSR 30 (superfine, scented) was more sensitive to higher rates (35 g ai ha-1) of azimsulfuron as compared to Pusa 1121 (fine, scented). Crop injuries were 8 and 28% in case of CSR 30; 5 and 15% in Pusa 1121 when applied with azimsulfuron 25 and 35 g ai ha-1, respectively. Azimsulfuron applied at 35 g ai ha-1 reduced yield in both cultivars but in CSR 30 yield reduction was twofold (11.5%) as that of Pusa 1121 (5.2%).