Attenuation of urokinase activity during experimental ischaemia protects the cerebral barrier from damage through regulation of matrix metalloproteinase-2 and NAD(P)H oxidase

Ischaemic injury impairs the integrity of the blood-brain barrier (BBB). In this study, we investigated the molecular causes of this defect with regard to the putative correlations among NAD(P)H oxidase, plasminogen-plasmin system components, and matrix metalloproteinases. Hence, the activities of NAD(P)H oxidase, matrix metalloproteinase-2, urokinase-type plasminogen activator (uPA), and tissue-type plasminogen activator (tPA), and superoxide anion levels, were assessed in human brain microvascular endothelial cells (HBMECs) exposed to oxygen-glucose deprivation (OGD) alone or OGD followed by reperfusion (OGD + R). The integrity of an in vitro model of BBB comprising HBMECs and astrocytes was studied by measuring transendothelial electrical resistance and the paracellular flux of albumin. OGD with or without reperfusion (OGD ± R) radically perturbed barrier function while concurrently enhancing uPA, tPA and NAD(P)H oxidase activities and superoxide anion release in HBMECs. Pharmacological inactivation of NAD(P)H oxidase attenuated OGD ± R-mediated BBB damage through modulation of matrix metalloproteinase-2 and tPA, but not uPA activity. Overactivation of NAD(P)H oxidase in HBMECs via cDNA electroporation of its p22-phox subunit confirmed the involvement of tPA in oxidase-mediated BBB disruption. Interestingly, blockade of uPA or uPA receptor preserved normal BBB function by neutralizing both NAD(P)H oxidase and matrix metalloproteinase-2 activities. Hence, selective targeting of uPA after ischaemic strokes may protect cerebral barrier integrity and function by concomitantly attenuating basement membrane degradation and oxidative stress.

PKC-β exacerbates in vitro brain barrier damage in hyperglycemic settings via regulation of RhoA/Rho-kinase/MLC2 pathway

Stroke patients with hyperglycemia (HG) develop higher volumes of brain edema emerging from disruption of blood-brain barrier (BBB). This study explored whether inductions of protein kinase C-β (PKC-β) and RhoA/Rho-kinase/myosin-regulatory light chain-2 (MLC2) pathway may account for HG-induced barrier damage using an in vitro model of human BBB comprising human brain microvascular endothelial cells (HBMEC) and astrocytes. Hyperglycemia (25 mmol/L D-glucose) markedly increased RhoA/Rho-kinase protein expressions (in-cell westerns), MLC2 phosphorylation (immunoblotting), and PKC-β (PepTag assay) and RhoA (Rhotekin-binding assay) activities in HBMEC while concurrently reducing the expression of tight junction protein occludin. Hyperglycemia-evoked in vitro barrier dysfunction, confirmed by decreases in transendothelial electrical resistance and concomitant increases in paracellular flux of Evan's blue-labeled albumin, was accompanied by malformations of actin cytoskeleton and tight junctions. Suppression of RhoA and Rho-kinase activities by anti-RhoA immunoglobulin G (IgG) electroporation and Y-27632, respectively prevented morphologic changes and restored plasma membrane localization of occludin. Normalization of glucose levels and silencing PKC-β activity neutralized the effects of HG on occludin and RhoA/Rho-kinase/MLC2 expression, localization, and activity and consequently improved in vitro barrier integrity and function. These results suggest that HG-induced exacerbation of the BBB breakdown after an ischemic stroke is mediated in large part by activation of PKC-β.

Numerical prediction of the low-velocity impact damage and compression after impact strength of composite laminates

Low-velocity impact damage can drastically reduce the residual mechanical properties of the composite structure even when there is barely visible impact damage. The ability to computationally predict the extent of damage and compression after impact (CAI) strength of a composite structure can potentially lead to the exploration of a larger design space without incurring significant development time and cost penalties. A three-dimensional damage model, to predict both low-velocity impact damage and compression after impact CAI strength of composite laminates, has been developed and implemented as a user material subroutine in the commercial finite element package, ABAQUS/Explicit. The virtual tests were executed in two steps, one to capture the impact damage and the other to predict the CAI strength. The observed intra-laminar damage features, delamination damage area as well as residual strength are discussed. It is shown that the predicted results for impact damage and CAI strength correlated well with experimental testing.

Investigations into the Control of Biofilm Damage with Relevance to Built Heritage (BIODAM)

This publication describes the results of a 3 year EC-funded R&D project (BIODAM) which investigated the effects of biological colonisation on heritage surfaces and evaluated of novel, low toxicity treatments for their ability to control of biofilms and for their compatibility with conservation products.

Mitochondrial DNA Damage Following Isolated Muscle Exercise: A Preliminary Investigation Into HO-1 Gene Expression

PURPOSE: This preliminary investigation was designed to test the hypothesis that high intensity single-leg exercise can cause extensive cell DNA damage, which subsequently may affect the expression of the HO-1 gene. METHODS: Six (n=6) apparently healthy male participants (age 27 + 7 yrs, stature 174 + 12 cm, body mass 79 + 4 kg and BMI 24 + 4 kg/m2) completed 100 isolated and continuous maximal concentric contractions (minimum force = 200 N, speed of contraction = 60°/sec) of the rectus femoris muscle. Using a spring-loaded and reusable Magnum biopsy gun with a 16-gauge core disposable biopsy needle, skeletal muscle micro biopsy tissue samples were extracted at rest and following exercise. mRNA gene expression was determined via two-step quantitative real-time PCR using GAPDH as a reference gene. RESULTS: The average muscle force production was 379 + 179 N. High intensity exercise increased mitochondrial 8-OHdG concentration (P < 0.05 vs. rest) with a concomitant decrease in total antioxidant capacity (P < 0.05 vs. rest). Exercise also increased protein oxidation as quantified by protein carbonyl concentration (P < 0.05 vs. rest). HO-1 expression increased (> 2-fold change vs. rest) following exercise, and it is postulated that this change was not significant due to low subject numbers (P > 0.05). CONCLUSION: These preliminary findings tentatively suggest that maximal concentric muscle contractions can cause intracellular DNA damage with no apparent disruption to the expression of the antioxidant stress protein HO-1. Moreover, it is likely that cell oxidant stress is required to activate the signal transduction cascade related to the expression of HO-1. A large-scale study incorporating a greater subject number is warranted to fully elucidate this relationship.

The European Union funded NEOShield project: A global approach to near-Earth object impact threat mitigation

Although discussions are underway within the Action Team 14 of the United Nations COPUOS, there is currently no concerted international plan addressing the impact threat from near-Earth objects (NEOs) and how to organize, prepare and implement mitigation measures. We report on a new international project to address impact hazard mitigation issues, being the subject of a proposal submitted to the European Commission in response to the 2011 FP7 Call "Prevention of impacts from near-Earth objects on our planet". Our consortium consists of 13 research institutes, universities, and industrial partners from 6 countries and includes leading US and Russian space organizations. The primary aim of the project, NEOShield, is to investigate in detail the three most promising mitigation techniques: the kinetic impactor, blast deflection,and the gravity tractor, and devise feasible demonstration missions. Furthermore, we will investigate options for an international strategy for implementation when an actual impact threat arises. The NEOShield project was formally accepted by the European Commission on 17 November 2011 and funded with a total of 5.8 million Euros for a period of 3.5 years. The kick-off meeting took place at the DLR Institute of Planetary Research, Berlin, in January 2012. In this paper we present a brief overview of the planned scope of the project.

NEOShield: An European Project to Address Impact Hazard Mitigation Issues

The European Union NEOShield project is desribed.The primary aim of the project is to investigate in detail the three most promising mitigation techniques: the kinetic impactor, blast deflection, and the gravity tractor, and devise feasible demonstration missions.

Mechanistic Rationale to Target PTEN-Deficient Tumor Cells with Inhibitors of the DNA Damage Response Kinase ATM

Ataxia telangiectasia mutated (ATM) is an important signaling molecule in the DNA damage response (DDR). ATM loss of function can produce a synthetic lethal phenotype in combination with tumor-associated mutations in FA/BRCA pathway components. In this study, we took an siRNA screening strategy to identify other tumor suppressors that, when inhibited, similarly sensitized cells to ATM inhibition. In this manner, we determined that PTEN and ATM were synthetically lethal when jointly inhibited. PTEN-deficient cells exhibited elevated levels of reactive oxygen species, increased endogenous DNA damage, and constitutive ATM activation. ATM inhibition caused catastrophic DNA damage, mitotic cell cycle arrest, and apoptosis specifically in PTEN-deficient cells in comparison with wild-type cells. Antioxidants abrogated the increase in DNA damage and ATM activation in PTEN-deficient cells, suggesting a requirement for oxidative DNA damage in the mechanism of cell death. Lastly, the ATM inhibitor KU-60019 was specifically toxic to PTEN mutant cancer cells in tumor xenografts and reversible by reintroduction of wild-type PTEN. Together, our results offer a mechanistic rationale for clinical evaluation of ATM inhibitors in PTEN-deficient tumors.

Predicting low velocity impact damage and Compression-After-Impact (CAI) behaviour of composite laminates

Low-velocity impact damage can drastically reduce the residual strength of a composite structure even when the damage is barely visible. The ability to computationally predict the extent of damage and compression-after-impact (CAI) strength of a composite structure can potentially lead to the exploration of a larger design space without incurring significant time and cost penalties. A high-fidelity three-dimensional composite damage model, to predict both low-velocity impact damage and CAI strength of composite laminates, has been developed and implemented as a user material subroutine in the commercial finite element package, ABAQUS/Explicit. The intralaminar damage model component accounts for physically-based tensile and compressive failure mechanisms, of the fibres and matrix, when subjected to a three-dimensional stress state. Cohesive behaviour was employed to model the interlaminar failure between plies with a bi-linear traction–separation law for capturing damage onset and subsequent damage evolution. The virtual tests, set up in ABAQUS/Explicit, were executed in three steps, one to capture the impact damage, the second to stabilize the specimen by imposing new boundary conditions required for compression testing, and the third to predict the CAI strength. The observed intralaminar damage features, delamination damage area as well as residual strength are discussed. It is shown that the predicted results for impact damage and CAI strength correlated well with experimental testing without the need of model calibration which is often required with other damage models.

Modelling Damage in Thin-Walled Structures [Keynote speaker]

A high-fidelity composite damage model is presented and applied to predict low-velocity impact damage, compression after impact (CAI) strength and crushing of thin-walled composite structures. The simulated results correlated well with experimental testing in terms of overall force-displacement response, damage morphologies and energy dissipation. The predictive power of this model makes it suitable for use as part of a virtual testing methodology, reducing the reliance on physical testing.  

Numerical prediction of the low-velocity impact damage and compression after impact strength of composite laminates

Low-velocity impact damage can drastically reduce the residual mechanical properties of the composite structure even when there is barely visible impact damage. The ability to computationally predict the extent of damage and compression after impact (CAI) strength of a composite structure can potentially lead to the exploration of a larger design space without incurring significant development time and cost penalties. A three-dimensional damage model, to predict both low-velocity impact damage and compression after impact CAI strength of composite laminates, has been developed and implemented as a user material subroutine in the commercial finite element package, ABAQUS/Explicit. The virtual tests were executed in two steps, one to capture the impact damage and the other to predict the CAI strength. The observed intra-laminar damage features, delamination damage area as well as residual strength are discussed. It is shown that the predicted results for impact damage and CAI strength correlated well with experimental testing.

Ultrasound entropy may be a new non-invasive measure of pre-clinical vascular damage in young hypertensive patients

Background: The identification of pre-clinical microvascular damage in hypertension by non-invasive techniques has proved frustrating for clinicians. This proof of concept study investigated whether entropy, a novel summary measure for characterizing blood velocity waveforms, is altered in participants with hypertension and may therefore be useful in risk stratification.

Methods: Doppler ultrasound waveforms were obtained from the carotid and retrobulbar circulation in 42 participants with uncomplicated grade 1 hypertension (mean systolic/diastolic blood pressure (BP) 142/92 mmHg), and 26 healthy controls (mean systolic/diastolic BP 116/69 mmHg). Mean wavelet entropy was derived from flow-velocity data and compared with traditional haemodynamic measures of microvascular function, namely the resistive and pulsatility indices.

Results: Entropy, was significantly higher in control participants in the central retinal artery (CRA) (differential mean 0.11 (standard error 0.05 cms(-1)), CI 0.009 to 0.219, p 0.017) and ophthalmic artery (0.12 (0.05), CI 0.004 to 0.215, p 0.04). In comparison, the resistive index (0.12 (0.05), CI 0.005 to 0.226, p 0.029) and pulsatility index (0.96 (0.38), CI 0.19 to 1.72, p 0.015) showed significant differences between groups in the CRA alone. Regression analysis indicated that entropy was significantly influenced by age and systolic blood pressure (r values 0.4-0.6). None of the measures were significantly altered in the larger conduit vessel.

Conclusion: This is the first application of entropy to human blood velocity waveform analysis and shows that this new technique has the ability to discriminate health from early hypertensive disease, thereby promoting the early identification of cardiovascular disease in a young hypertensive population.

Interventions Targeting Child Undernutrition in Developing Countries May Be Undermined by Dietary Exposure to Aflatoxin

Child undernutrition, a form of malnutrition, is a major public health burden in developing countries. Supplementation interventions targeting the major micronutrient deficiencies have only reduced the burden of child undernutrition to a certain extent, indicating that there are other underlying determinants that need addressed. Aflatoxin exposure, which is also highly prevalent in developing countries, may be considered to be an aggravating factor for child undernutrition. Increasing evidence suggests that aflatoxin exposure can occur in any stage of life including in utero through a trans-placental pathway and in early childhood (through contaminated weaning food and family food). Early life exposure to aflatoxin is associated with adverse effects on low birth weight, stunting, immune suppression and liver function damage. The mechanisms underlying impaired growth and aflatoxin exposure are still unclear but intestinal function damage, reduced immune function and alteration in the insulin-like growth factor axis caused by liver damage, are suggested hypotheses. Given the fact that both aflatoxin and child undernutrition are common in sub-Saharan Africa, effective interventions aimed at reducing undernutrition cannot be satisfactorily achieved until the interactive relationship between aflatoxin and child undernutrition is clearly understood, and an aflatoxin mitigation strategy has taken effect in those vulnerable mothers and children.