Lipid-modified azurin of Neisseria meningitidis is a copper protein localized on the outer membrane surface and not regulated by FNR

The laz gene of Neisseria meningitidis is predicted to encode a lipid-modified azurin (Laz). Laz is very similar to azurin, a periplasmic protein, which belongs to the copper-containing proteins in the cupredoxin superfamily. In other bacteria, azurin is an electron donor to nitrite reductase, an important enzyme in the denitrifying process. It is not known whether Laz could function as an electron transfer protein in this important pathogen. Laz protein was heterologously expressed in Escherichia coli and purified. Electrospray mass spectrometry indicated that the Laz protein contains one copper ion. Laz was shown to be redox-active in the presence of its redox center copper ion. When oxidized, Laz exhibits an intense blue colour and absorbs visible light around 626 nm. The absorption is lost when exposed to diethyldithiocarbamate, a copper chelating agent. Polyclonal antibodies were raised against purified Laz for detecting expression of Laz under different growth conditions and to determine the orientation of Laz on the outer membrane. The expression of Laz under microaerobic and microaerobic denitrifying conditions was slightly higher than that under aerobic conditions. However, the expression of Laz was similar between the wild type strain and an fnr mutant, suggesting that Fumarate/Nitrate reduction regulator (FNR) does not regulate the expression of Laz despite the presence of a partial FNR box upstream of the laz gene. We propose that some Laz protein is exposed on the outer membrane surface of N. meningitidis as the αLaz antibodies can increase killing by complement in a capsule deficient N. meningitidis strain, in a dose-dependent fashion.

Interlaminar mechanical properties of carbon fiber reinforced plastic laminates modified with graphene oxide interleaf

By taking the advantage of the excellent mechanical properties and high specific surface area of graphene oxide (GO) sheets, we develop a simple and effective strategy to improve the interlaminar mechanical properties of carbon fiber reinforced plastic (CFRP) laminates. With the incorporation of graphene oxide reinforced epoxy interleaf into the interface of CFRP laminates, the Mode-I fracture toughness and resistance were greatly increased. The experimental results of double cantilever beam (DCB) tests demonstrated that, with 2 g/m2 addition of GO, the Mode-I fracture toughness and resistance of the specimen increase by 170.8% and 108.0%, respectively, compared to those of the plain specimen. The improvement mechanisms were investigated by the observation of fracture surface with scanning electron microscopies. Moreover, finite element analyses were performed based on the cohesive zone model to verify the experimental fracture toughness and to predict the interfacial tensile strength of CFRP laminates.

Nelson's (1976) Modified Card Sorting Test: A review

We review studies of Nelson's (1976) Modified Card Sorting Test (MCST) that have examined the performance of subjects with frontal lobe dysfunction. Six studies investigated the performance of normal controls and patients with frontal lobe dysfunction, whereas four studies compared the performance of frontal and nonfrontal patients. One further study compared the performance of amnesic patients both on the MCST and on the original Wisconsin Card Sorting Test (WCST). Evidence regarding the MCST's differential sensitivity to frontal lobe dysfunction is weak, as is the evidence regarding the equivalence of the MCST and WCST. It is likely that the MCST is an altogether different test from the standard version. In the absence of proper normative data for the MCST, we provide a table of scores derived from the control groups of various studies. Given the paucity of evidence, further research is required before the MCST can be recommended for use as a marker of frontal lobe dysfunction.

Dynamics of gray matter loss in Alzheimer's disease

We detected and mapped a dynamically spreading wave of gray matter loss in the brains of patients with Alzheimer's disease (AD). The loss pattern was visualized in four dimensions as it spread over time from temporal and limbic cortices into frontal and occipital brain regions, sparing sensorimotor cortices. The shifting deficits were asymmetric (left hemisphere > right hemisphere) and correlated with progressively declining cognitive status (p < 0.0006). Novel brain mapping methods allowed us to visualize dynamic patterns of atrophy in 52 high-resolution magnetic resonance image scans of 12 patients with AD (age 68.4 ± 1.9 years) and 14 elderly matched controls (age 71.4 ± 0.9 years) scanned longitudinally (two scans; interscan interval 2.1 ± 0.4 years). A cortical pattern matching technique encoded changes in brain shape and tissue distribution across subjects and time. Cortical atrophy occurred in a well defined sequence as the disease progressed, mirroring the sequence of neurofibrillary tangle accumulation observed in cross sections at autopsy. Advancing deficits were visualized as dynamic maps that change over time. Frontal regions, spared early in the disease, showed pervasive deficits later (< 15% loss). The maps distinguished different phases of AD and differentiated AD from normal aging. Local gray matter loss rates (5.3 ± 2.3% per year in AD v 0.9 ± 0.9% per year in controls) were faster in the left hemisphere (p < 0.029) than the right. Transient barriers to disease progression appeared at limbic/frontal boundaries. This degenerative sequence, observed in vivo as it developed, provides the first quantitative, dynamic visualization of cortical atrophic rates in normal elderly populations and in those with dementia.

Mapping cortical change in Alzheimer's disease, brain development, and schizophrenia

This paper describes algorithms that can identify patterns of brain structure and function associated with Alzheimer's disease, schizophrenia, normal aging, and abnormal brain development based on imaging data collected in large human populations. Extraordinary information can be discovered with these techniques: dynamic brain maps reveal how the brain grows in childhood, how it changes in disease, and how it responds to medication. Genetic brain maps can reveal genetic influences on brain structure, shedding light on the nature-nurture debate, and the mechanisms underlying inherited neurobehavioral disorders. Recently, we created time-lapse movies of brain structure for a variety of diseases. These identify complex, shifting patterns of brain structural deficits, revealing where, and at what rate, the path of brain deterioration in illness deviates from normal. Statistical criteria can then identify situations in which these changes are abnormally accelerated, or when medication or other interventions slow them. In this paper, we focus on describing our approaches to map structural changes in the cortex. These methods have already been used to reveal the profile of brain anomalies in studies of dementia, epilepsy, depression, childhood- and adult-onset schizophrenia, bipolar disorder, attention-deficit/hyperactivity disorder, fetal alcohol syndrome, Tourette syndrome, Williams syndrome, and in methamphetamine abusers. Specifically, we describe an image analysis pipeline known as cortical pattern matching that helps compare and pool cortical data over time and across subjects. Statistics are then defined to identify brain structural differences between groups, including localized alterations in cortical thickness, gray matter density (GMD), and asymmetries in cortical organization. Subtle features, not seen in individual brain scans, often emerge when population-based brain data are averaged in this way. Illustrative examples are presented to show the profound effects of development and various diseases on the human cortex. Dynamically spreading waves of gray matter loss are tracked in dementia and schizophrenia, and these sequences are related to normally occurring changes in healthy subjects of various ages.

Pancreatic islet basement membrane loss and remodeling after mouse islet isolation and transplantation: Impact for allograft rejection

The isolation of islets by collagenase digestion can cause damage and impact the efficiency of islet engraftment and function. In this study, we assessed the basement membranes (BMs) of mouse pancreatic islets as a molecular biomarker for islet integrity, damage after isolation, and islet repair in vitro as well as in the absence or presence of an immune response after transplantation. Immunofluorescence staining of BM matrix proteins and the endothelial cell marker platelet endothelial cell adhesion molecule-1 (PECAM-1) was performed on pancreatic islets in situ, isolated islets, islets cultured for 4 days, and islet grafts at 3-10 days posttransplantation. Flow cytometry was used to investigate the expression of BM matrix proteins in isolated islet β-cells. The islet BM, consisting of collagen type IV and components of Engelbreth-Holm-Swarm (EHS) tumor laminin 111, laminin α2, nidogen-2, and perlecan in pancreatic islets in situ, was completely lost during islet isolation. It was not reestablished during culture for 4 days. Peri- and intraislet BM restoration was identified after islet isotransplantation and coincided with the migration pattern of PECAM-1(+) vascular endothelial cells (VECs). After islet allotransplantation, the restoration of VEC-derived peri-islet BMs was initiated but did not lead to the formation of the intraislet vasculature. Instead, an abnormally enlarged peri-islet vasculature developed, coinciding with islet allograft rejection. The islet BM is a sensitive biomarker of islet damage resulting from enzymatic isolation and of islet repair after transplantation. After transplantation, remodeling of both peri- and intraislet BMs restores β-cell-matrix attachment, a recognized requirement for β-cell survival, for isografts but not for allografts. Preventing isolation-induced islet BM damage would be expected to preserve the intrinsic barrier function of islet BMs, thereby influencing both the effector mechanisms required for allograft rejection and the antirejection strategies needed for allograft survival.

An N-ethyl-n-nitrosourea induced corticotropin-releasing hormone promoter mutation provides a mouse model for endogenous glucocorticoid excess

Cushing's syndrome, which is characterized by excessive circulating glucocorticoid concentrations, maybe due to ACTH-dependent or -independent causes that include anterior pituitary and adrenal cortical tumors, respectively. ACTH secretion is stimulated by CRH, and we report a mouse model for Cushing's syndrome due to an N-ethyl-N-nitrosourea (ENU) induced Crh mutation at -120 bp of the promoter region, which significantly increased luciferase reporter activity and was thus a gain-of-function mutation. Crh -120/+ mice, when compared with wild-type littermates, had obesity, muscle wasting, thin skin, hair loss, and elevated plasma and urinary concentrations of corticosterone. In addition, Crh-120/+ mice had hyperglycemia, hyperfructosaminemia, hyperinsulinemia, hypercholesterolemia, hypertriglyceridemia, and hyperleptinemia but normal adiponectin. Crh -120/+ mice also had low bone mineral density, hypercalcemia, hypercalciuria, and decreased concentrations of plasma PTH and osteocalcin. Bone histomorphometry revealed Crh-120/+ mice to have significant reductions in mineralizing surface area, mineral apposition, bone formation rates, osteoblast number, and the percentage of corticoendosteal bone covered by osteoblasts, which was accompanied by an increase in adipocytes in the bone marrow. Thus, a mouse model for Cushing's syndrome has been established, and this will help in further elucidating the pathophysiological effects of glucocorticoid excess and in evaluating treatments for corticosteroid-induced osteoporosis.

Genetic disorders of the LRP5-Wnt signalling pathway affecting the skeleton

Osteoporosis is a common, increasingly prevalent and potentially debilitating condition of men and women. Genetic factors are major determinants of bone mass and the risk of fracture, but few genes have been definitively demonstrated to be involved. The identification of these factors will provide novel insights into the processes of bone formation and loss and thus the pathogenesis of osteoporosis, enabling the rational development of novel therapies. In this article, we present the extensive genetic and functional data indicating that the LRP5 gene and the Wnt signalling pathway are key players in bone formation and the risk of osteoporosis, and that LRP5 signalling is essential for normal morphology, developmental processes and bone health.

Influence of LRP5 polymorphisms on normal variation in BMD

Genetic studies based on cohorts with rare and extreme bone phenotypes have shown that the LRP5 gene is an important genetic modulator of BMD. Using family-based and case-control approaches, this study examines the role of the LRP5 gene in determining normal population variation of BMD and describes significant association and suggestive linkage between LRP5 gene polymorphisms and BMD in >900 individuals with a broad range of BMD. Introduction: Osteoporosis is a common, highly heritable condition determined by complex interactions of genetic and environmental etiologies. Genetic factors alone can account for 50-80% of the interindividual variation in BMD. Mutations in the LRP5 gene on chromosome 11q12-13 have been associated with rare syndromes characterized by extremely low or high BMD, but little is known about the contribution of this gene to the development of osteoporosis and determination of BMD in a normal population. Materials and Methods: To examine the entire spectrum of low to high BMD, 152 osteoporotic probands, their families (597 individuals), and 160 women with elevated BMD (T score > 2.5) were recruited. BMD at the lumbar spine, femoral neck, and hip were measured in each subject using DXA. Results: PAGE sequencing of the LRP5 gene revealed 10 single nucleotide polymorphisms (SNPs), 8 of which had allele frequencies of >5%, in exons 8, 9, 10, 15, and 18 and in introns 6, 7, and 21. Within families, a strong association was observed between an SNP at nucleotide C171346A in intron 21 and total hip BMD (p < 1 × 10-5 in men only, p = 0.0019 in both men and women). This association was also observed in comparisons of osteoporotic probands and unrelated elevated BMD in women (p = 0.03), along with associations with markers in exons 8 (C135242T, p = 0.007) and 9 (C141759T, p = 0.02). Haplotypes composed of two to three of the SNPs G121513A, C135242T, G138351A, and C141759T were strongly associated with BMD when comparing osteoporotic probands and high BMD cases (p < 0.003). An SNP at nucleotide C165215T in exon 18 was linked to BMD at the lumbar spine, femoral neck, and total hip (parametric LOD scores = 2.8, 2.5, and 2.2 and nonparametric LOD scores = 0.3, 1.1, and 2.2, respectively) but was not genetically associated with BMD variation. Conclusion: These results show that common LRP5 polymorphisms contribute to the determination of BMD in the general population.

Effect of decellularization on the load-bearing characteristics of articular cartilage matrix

The application of decellularized extracellular matrices to aid tissue regeneration in reconstructive surgery and regenerative medicine has been promising. Several decellularization protocols for removing cellular materials from natural tissues such as heart valves are currently in use. This paper evaluates the feasibility of potential extension of this methodology relative to the desirable properties of load bearing joint tissues such as stiffness, porosity and ability to recover adequately after deformation to facilitate physiological function. Two decellularization protocols, namely: Trypsin and Triton X-100 were evaluated against their effects on bovine articular cartilage, using biomechanical, biochemical and microstructural techniques. These analyses revealed that decellularization with trypsin resulted in severe loss of mechanical stiffness including deleterious collapse of the collagen architecture which in turn significantly compromised the porosity of the construct. In contrast, triton X-100 detergent treatment yielded samples that retain mechanical stiffness relative to that of the normal intact cartilage sample, but the resulting construct contained ruminant cellular constituents. We conclude that both of these common decellularization protocols are inadequate for producing constructs that can serve as effective replacement and scaffolds to regenerate articular joint tissue.

Serum immunoreactive pancreatic lipase and cationic trypsinogen for the assessment of exocrine pancreatic function in older patients with cystic fibrosis

Indirect and qualitative tests of pancreatic function are commonly used to screen patients with cystic fibrosis for pancreatic insufficiency. In an attempt to develop a more quantitative assessment, we compared the usefulness of measuring serum pancreatic lipase using a newly developed enzyme-linked immunosorbent immunoassay with that of cationic trypsinogen using a radioimmunoassay in the assessment of exocrine pancreatic function in patients with cystic fibrosis. Previously, we have shown neither lipase nor trypsinogen to be of use in assessing pancreatic function prior to 5 years of age because the majority of patients with cystic fibrosis in early infancy have elevated serum levels regardless of pancreatic function. Therefore, we studied 77 patients with cystic fibrosis older than 5 years of age, 41 with steatorrhea and 36 without steatorrhea. In addition, 28 of 77 patients consented to undergo a quantitative pancreatic stimulation test. There was a significant difference between the steatorrheic and nonsteatorrheic patients with the steatorrheic group having lower lipase and trypsinogen values than the nonsteatorrheic group (P < .001). Sensitivities and specificities in detecting steatorrhea were 95% and 86%, respectively, for lipase and 93% and 92%, respectively, for trypsinogen. No correlations were found between the serum levels of lipase and trypsinogen and their respective duodenal concentrations because of abnormally high serum levels of both enzymes found in some nonsteatorrheic patients. We conclude from this study that both serum lipase and trypsinogen levels accurately detect steatorrhea in patients with cystic fibrosis who are older than 5 years but are imprecise indicators of specific pancreatic exocrine function above the level needed for normal fat absorption.

Effect of fragmentation, habitat loss and within-patch habitat characteristics on ant assemblages in semi-arid woodlands of eastern Australia.

The reliability of ants as bioindicators of ecosystem condition is dependent on the consistency of their response to localised habitat characteristics, which may be modified by larger-scale effects of habitat fragmentation and loss. We assessed the relative contribution of habitat fragmentation, habitat loss and within-patch habitat characteristics in determining ant assemblages in semi-arid woodland in Queensland, Australia. Species and functional group abundance were recorded using pitfall traps across 20 woodland patches in landscapes that exhibited a range of fragmentation states. Of fragmentation measures, changes in patch area and patch edge contrast exerted the greatest influence on species assemblages, after accounting for differences in habitat loss. However, 35% of fragmentation effects on species were confounded by the effects of habitat characteristics and habitat loss. Within-patch habitat characteristics explained more than twice the amount of species variation attributable to fragmentation and four times the variation explained by habitat loss. The study indicates that within-patch habitat characteristics are the predominant drivers of ant composition. We suggest that caution should be exercised in interpreting the independent effects of habitat fragmentation and loss on ant assemblages without jointly considering localised habitat attributes and associated joint effects.

Transmission loss analysis of rectangular expansion chamber with arbitrary location of inlet/outlet by means of Green's functions

Transmission loss of a rectangular expansion chamber, the inlet and outlet of which are situated at arbitrary locations of the chamber, i.e., the side wall or the face of the chamber, are analyzed here based on the Green's function of a rectangular cavity with homogeneous boundary conditions. The rectangular chamber Green's function is expressed in terms of a finite number of rigid rectangular cavity mode shapes. The inlet and outlet ports are modeled as uniform velocity pistons. If the size of the piston is small compared to wavelength, then the plane wave excitation is a valid assumption. The velocity potential inside the chamber is expressed by superimposing the velocity potentials of two different configurations. The first configuration is a piston source at the inlet port and a rigid termination at the outlet, and the second one is a piston at the outlet with a rigid termination at the inlet. Pressure inside the chamber is derived from velocity potentials using linear momentum equation. The average pressure acting on the pistons at the inlet and outlet locations is estimated by integrating the acoustic pressure over the piston area in the two constituent configurations. The transfer matrix is derived from the average pressure values and thence the transmission loss is calculated. The results are verified against those in the literature where use has been made of modal expansions and also numerical models (FEM fluid). The transfer matrix formulation for yielding wall rectangular chambers has been derived incorporating the structural–acoustic coupling. Parametric studies are conducted for different inlet and outlet configurations, and the various phenomena occurring in the TL curves that cannot be explained by the classical plane wave theory, are discussed.

Oscillatory brain activity in memory disorders

Neuronal oscillations are thought to underlie interactions between distinct brain regions required for normal memory functioning. This study aimed at elucidating the neuronal basis of memory abnormalities in neurodegenerative disorders. Magnetoencephalography (MEG) was used to measure oscillatory brain signals in patients with Alzheimer s disease (AD), a neurodegenerative disease causing progressive cognitive decline, and mild cognitive impairment (MCI), a disorder characterized by mild but clinically significant complaints of memory loss without apparent impairment in other cognitive domains. Furthermore, to help interpret our AD/MCI results and to develop more powerful oscillatory MEG paradigms for clinical memory studies, oscillatory neuronal activity underlying declarative memory, the function which is afflicted first in both AD and MCI, was investigated in a group of healthy subjects. An increased temporal-lobe contribution coinciding with parieto-occipital deficits in oscillatory activity was observed in AD patients: sources in the 6 12.5 Hz range were significantly stronger in the parieto-occipital and significantly weaker in the right temporal region in AD patients, as compared to MCI patients and healthy elderly subjects. Further, the auditory steady-state response, thought to represent both evoked and induced activity, was enhanced in AD patients, as compared to controls, possibly reflecting decreased inhibition in auditory processing and deficits in adaptation to repetitive stimulation with low relevance. Finally, the methodological study revealed that successful declarative encoding and retrieval is associated with increases in occipital gamma and right hemisphere theta power in healthy unmedicated subjects. This result suggests that investigation of neuronal oscillations during cognitive performance could potentially be used to investigate declarative memory deficits in AD patients. Taken together, the present results provide an insight on the role of brain oscillatory activity in memory function and memory disorders.

Prospective decrease in progesterone concentrations in female lightweight rowers during the competition season compared with the off-season: A controlled study examining weight loss and intensive exercise

The purpose of this study was to monitor ovarian hormone function response to intense exercise and body weight changes in female athletes. Ovarian hormone function was evaluated in 12 female lightweight rowers and 10 age-height-weight matched sedentary controls. Ovarian hormone function was assessed during consecutive competition season and off season, by measurement of peak and average alternative day overnight urinary oestrone glucuronide (E1G) and pregnanediol glucuronide (PdG) excretion. Competition season was associated with a 5.8 kg (9.3%) body weight loss in the lightweight rowers. Significantly lower competition season peak and average urinary excretion of PdG were found in the lightweight rowers compared with the controls. Lower competition season peak and average urinary excretion of E1G were also found in the lightweight rowers compared with the controls, but the difference did not reach significance. The number of rowing training hours was a significant determinant of peak PdG excretion in the rowers (R2 = 0.40; p<0.02). The seasonal suppression of PdG excretion was associated with degree of weight loss (R2 = 0.46; p<0.01). The competition related decrease in E1G and PdG excretion for the lightweight rowers was predominantly restored during the off season when exercise intensity and duration were decreased and body weight increased. These results showed a significant (p<0.05) reduction in progesterone metabolite excretion and a non-significant decrease in oestrone metabolite excretion associated with intensive competition season training loads and body weight reduction in female lightweight rowers.