Stimulation of MMP-11 (stromelysin-3) expression in mouse fibroblasts by cytokines, collagen and co-culture with human breast cancer cell lines

Background Matrix metalloproteinases (MMPs) are central to degradation of the extracellular matrix and basement membrane during both normal and carcinogenic tissue remodeling. MT1-MMP (MMP-14) and stromelysin-3 (MMP-11) are two members of the MMP family of proteolytic enzymes that have been specifically implicated in breast cancer progression. Expressed in stromal fibroblasts adjacent to epithelial tumour cells, the mechanism of MT1-MMP and MMP-11 induction remains unknown. Methods To investigate possible mechanisms of induction, we examined the effects of a number of plausible regulatory agents and treatments that may physiologically influence MMP expression during tumour progression. Thus NIH3T3 and primary mouse embryonic fibroblasts (MEFs) were: a) treated with the cytokines IL-1β, IL-2, IL-6, IL-8 and TGF-β for 3, 6, 12, 24, and 48 hours; b) grown on collagens I, IV and V; c) treated with fibronectin, con-A and matrigel; and d) co-cultured with a range of HBC (human breast cancer) cell lines of varied invasive and metastatic potential. Results Competitive quantitative RT-PCR indicated that MMP-11 expression was stimulated to a level greater than 100%, by 48 hour treatments of IL-1β, IL-2, TGF-β, fibronectin and collagen V. No other substantial changes in expression of MMP-11 or MT1-MMP in either tested fibroblast culture, under any treatment conditions, were observed. Conclusion We have demonstrated significant MMP-11 stimulation in mouse fibroblasts using cytokines, matrix constituents and HBC cell lines, and also some inhibition of MT1-MMP. Our data suggest that the regulation of these genes in the complex stromal-epithelial interactions that occur in human breast carcinoma, is influenced by several mechanisms.

Real - time prediction of rainfall induced instability of residual soil slopes associated with deep cracks

The early warning based on real-time prediction of rain-induced instability of natural residual slopes helps to minimise human casualties due to such slope failures. Slope instability prediction is complicated, as it is influenced by many factors, including soil properties, soil behaviour, slope geometry, and the location and size of deep cracks in the slope. These deep cracks can facilitate rainwater infiltration into the deep soil layers and reduce the unsaturated shear strength of residual soil. Subsequently, it can form a slip surface, triggering a landslide even in partially saturated soil slopes. Although past research has shown the effects of surface-cracks on soil stability, research examining the influence of deep-cracks on soil stability is very limited. This study aimed to develop methodologies for predicting the real-time rain-induced instability of natural residual soil slopes with deep cracks. The results can be used to warn against potential rain-induced slope failures. The literature review conducted on rain induced slope instability of unsaturated residual soil associated with soil crack, reveals that only limited studies have been done in the following areas related to this topic: - Methods for detecting deep cracks in residual soil slopes. - Practical application of unsaturated soil theory in slope stability analysis. - Mechanistic methods for real-time prediction of rain induced residual soil slope instability in critical slopes with deep cracks. Two natural residual soil slopes at Jombok Village, Ngantang City, Indonesia, which are located near a residential area, were investigated to obtain the parameters required for the stability analysis of the slope. A survey first identified all related field geometrical information including slope, roads, rivers, buildings, and boundaries of the slope. Second, the electrical resistivity tomography (ERT) method was used on the slope to identify the location and geometrical characteristics of deep cracks. The two ERT array models employed in this research are: Dipole-dipole and Azimuthal. Next, bore-hole tests were conducted at different locations in the slope to identify soil layers and to collect undisturbed soil samples for laboratory measurement of the soil parameters required for the stability analysis. At the same bore hole locations, Standard Penetration Test (SPT) was undertaken. Undisturbed soil samples taken from the bore-holes were tested in a laboratory to determine the variation of the following soil properties with the depth: - Classification and physical properties such as grain size distribution, atterberg limits, water content, dry density and specific gravity. - Saturated and unsaturated shear strength properties using direct shear apparatus. - Soil water characteristic curves (SWCC) using filter paper method. - Saturated hydraulic conductivity. The following three methods were used to detect and simulate the location and orientation of cracks in the investigated slope: (1) The electrical resistivity distribution of sub-soil obtained from ERT. (2) The profile of classification and physical properties of the soil, based on laboratory testing of soil samples collected from bore-holes and visual observations of the cracks on the slope surface. (3) The results of stress distribution obtained from 2D dynamic analysis of the slope using QUAKE/W software, together with the laboratory measured soil parameters and earthquake records of the area. It was assumed that the deep crack in the slope under investigation was generated by earthquakes. A good agreement was obtained when comparing the location and the orientation of the cracks detected by Method-1 and Method-2. However, the simulated cracks in Method-3 were not in good agreement with the output of Method-1 and Method-2. This may have been due to the material properties used and the assumptions made, for the analysis. From Method-1 and Method-2, it can be concluded that the ERT method can be used to detect the location and orientation of a crack in a soil slope, when the ERT is conducted in very dry or very wet soil conditions. In this study, the cracks detected by the ERT were used for stability analysis of the slope. The stability of the slope was determined using the factor of safety (FOS) of a critical slip surface obtained by SLOPE/W using the limit equilibrium method. Pore-water pressure values for the stability analysis were obtained by coupling the transient seepage analysis of the slope using finite element based software, called SEEP/W. A parametric study conducted on the stability of an investigated slope revealed that the existence of deep cracks and their location in the soil slope are critical for its stability. The following two steps are proposed to predict the rain-induced instability of a residual soil slope with cracks. (a) Step-1: The transient stability analysis of the slope is conducted from the date of the investigation (initial conditions are based on the investigation) to the preferred date (current date), using measured rainfall data. Then, the stability analyses are continued for the next 12 months using the predicted annual rainfall that will be based on the previous five years rainfall data for the area. (b) Step-2: The stability of the slope is calculated in real-time using real-time measured rainfall. In this calculation, rainfall is predicted for the next hour or 24 hours and the stability of the slope is calculated one hour or 24 hours in advance using real time rainfall data. If Step-1 analysis shows critical stability for the forthcoming year, it is recommended that Step-2 be used for more accurate warning against the future failure of the slope. In this research, the results of the application of the Step-1 on an investigated slope (Slope-1) showed that its stability was not approaching a critical value for year 2012 (until 31st December 2012) and therefore, the application of Step-2 was not necessary for the year 2012. A case study (Slope-2) was used to verify the applicability of the complete proposed predictive method. A landslide event at Slope-2 occurred on 31st October 2010. The transient seepage and stability analyses of the slope using data obtained from field tests such as Bore-hole, SPT, ERT and Laboratory tests, were conducted on 12th June 2010 following the Step-1 and found that the slope in critical condition on that current date. It was then showing that the application of the Step-2 could have predicted this failure by giving sufficient warning time.

The effect of varied test instructions on neuropsychological performance following mild traumatic brain injury : an investigation of ‘diagnosis threat’

Diagnosis threat is a psychosocial factor that has been proposed to contribute to poor outcomes following mild traumatic brain injury (mTBI). This threat is thought to impair the cognitive test performance of individuals with mTBI because of negative injury stereotypes. University students (N= 45, 62.2% female) with a history of mTBI were randomly allocated to a diagnosis threat (DT, n=15), reduced threat (DT-reduced, n=15) or neutral (n=15) group. The reduced threat condition invoked a positive stereotype (i.e., that people with mTBI can perform well on cognitive tests). All participants were given neutral instructions before they completed baseline tests of: a) objective cognitive function across a number of domains; b) psychological symptoms; and, c) PCS symptoms, including self-reported cognitive and emotional difficulties. Participants then received either neutral, DT or DT-reduced instructions, before repeating the tests. Results were analyzed using separate mixed model ANOVAs; one for each dependent measure. The only significant result was for the 2 X 3 ANOVA on an objective test of attention/working memory, Digit Span, p<.05, such that the DT-reduced group performed better than the other groups, which were not different from each other. Although not consistent with predictions or earlier DT studies, the absence of group differences on most tests fits with several recent DT findings. The results of this study suggest that it is timely to reconsider the role of DT as a unique contributor to poor mTBI outcome.

Comparison between electrically-evoked and voluntary wrist movements on sensorimotor and prefrontal cortical activation : A multi-channel time domain fNIRS study

Neuromuscular electrical stimulation (NMES) has been consistently demonstrated to improve skeletal muscle function in neurological populations with movement disorders, such as poststroke and incomplete spinal cord injury (Vanderthommen and Duchateau, 2007). Recent research has documented that rapid, supraspinal central nervous system reorganisation/neuroplastic mechanisms are also implicated during NMES (Chipchase et al., 2011). Functional neuroimaging studies have shown NMES to activate a network of sub-cortical and cortical brain regions, including the sensorimotor (SMC) and prefrontal (PFC) cortex (Blickenstorfer et al., 2009; Han et al., 2003; Muthalib et al., 2012). A relationship between increase in SMC activation with increasing NMES current intensity up to motor threshold has been previously reported using functional MRI (Smith et al., 2003). However, since clinical neurorehabilitation programmes commonly utilise NMES current intensities above the motor threshold and up to the maximum tolerated current intensity (MTI), limited research has determined the cortical correlates of increasing NMES current intensity at or above MTI (Muthalib et al., 2012). In our previous study (Muthalib et al., 2012), we assessed contralateral PFC activation using 1-channel functional near infrared spectroscopy (fNIRS) during NMES of the elbow flexors by increasing current intensity from motor threshold to greater than MTI and showed a linear relationship between NMES current intensity and the level of PFC activation. However, the relationship between NMES current intensity and activation of the motor cortical network, including SMC and PFC, has not been clarified. Moreover, it is of scientific and clinical relevance to know how NMES affects the central nervous system, especially in comparison to voluntary (VOL) muscle activation. Therefore, the aim of this study was to utilise multi-channel time domain fNIRS to compare SMC and PFC activation between VOL and NMESevoked wrist extension movements.

Brain-inspired sensor fusion for navigating robots

Current state of the art robot mapping and navigation systems produce impressive performance under a narrow range of robot platform, sensor and environmental conditions, in contrast to animals such as rats that produce “good enough” maps that enable them to function under an incredible range of situations. In this paper we present a rat-inspired featureless sensor-fusion system that assesses the usefulness of multiple sensor modalities based on their utility and coherence for place recognition, without knowledge as to the type of sensor. We demonstrate the system on a Pioneer robot in indoor and outdoor environments with abrupt lighting changes. Through dynamic weighting of the sensors, the system is able to perform correct place recognition and mapping where the static sensor weighting approach fails.

Is recovery driven by central or peripheral factors? A role for the brain in recovery following intermittent-sprint exercise

Prolonged intermittent-sprint exercise (i.e., team sports) induce disturbances in skeletal muscle structure and function that are associated with reduced contractile function, a cascade of inflammatory responses, perceptual soreness, and a delayed return to optimal physical performance. In this context, recovery from exercise-induced fatigue is traditionally treated from a peripheral viewpoint, with the regeneration of muscle physiology and other peripheral factors the target of recovery strategies. The direction of this research narrative on post-exercise recovery differs to the increasing emphasis on the complex interaction between both central and peripheral factors regulating exercise intensity during exercise performance. Given the role of the central nervous system (CNS) in motor-unit recruitment during exercise, it too may have an integral role in post-exercise recovery. Indeed, this hypothesis is indirectly supported by an apparent disconnect in time-course changes in physiological and biochemical markers resultant from exercise and the ensuing recovery of exercise performance. Equally, improvements in perceptual recovery, even withstanding the physiological state of recovery, may interact with both feed-forward/feed-back mechanisms to influence subsequent efforts. Considering the research interest afforded to recovery methodologies designed to hasten the return of homeostasis within the muscle, the limited focus on contributors to post-exercise recovery from CNS origins is somewhat surprising. Based on this context, the current review aims to outline the potential contributions of the brain to performance recovery after strenuous exercise.

Mechanical stimulation of the pro-angiogenic capacity of human fracture haematoma: Involvement of VEGF mechano-regulation

Compromised angiogenesis appears to be a major limitation in various suboptimal bone healing situations. Appropriate mechanical stimuli support blood vessel formation in vivo and improve healing outcomes. However, the mechanisms responsible for this association are unclear. To address this question, the paracrine angiogenic potential of early human fracture haematoma and its responsiveness to mechanical loading, as well as angiogenic growth factors involved, were investigated in vitro. Human haematomas were collected from healthy patients undergoing surgery within 72. h after bone fracture. The haematomas were embedded in a fibrin matrix, and cultured in a bioreactor resembling the in vivo conditions of the early phase of bone healing (20 compression, 1. Hz) over 3. days. Conditioned medium (CM) from the bioreactor was then analyzed. The matrices were also incubated in fresh medium for a further 24. h to evaluate the persistence of the effects. Growth factor (GF) concentrations were measured in the CM by ELISAs. In vitro tube formation assays were conducted on Matrigel with the HMEC-1 cell line, with or without inhibition of vascular endothelial growth factor receptor 2 (VEGFR2). Cell numbers were quantified using an MTS test. In vitro endothelial tube formation was enhanced by CM from haematomas, compared to fibrin controls. The angiogenesis regulators, vascular endothelial growth factor (VEGF) and transforming growth factor β1 (TGF-β1), were released into the haematoma CM, but not angiopoietins 1 or 2 (Ang1, 2), basic fibroblast growth factor (bFGF) or platelet-derived growth factor (PDGF). Mechanical stimulation of haematomas, but not fibrin controls, further increased the induction of tube formation by their CM. The mechanically stimulated haematoma matrices retained their elevated pro-angiogenic capacity for 24. h. The pro-angiogenic effect was cancelled by inhibition of VEGFR2 signalling. VEGF concentrations in CM tended to be elevated by mechanical stimulation; this was significant in haematomas from younger, but not from older patients. Other GFs were not mechanically regulated. In conclusion, the paracrine pro-angiogenic capacity of early human haematomas is enhanced by mechanical stimulation. This effect lasts even after removing the mechanical stimulus and appears to be VEGFR2-dependent.

The optimal duration and delay of first aid treatment for deep partial thickness burn injuries

Using our porcine model of deep dermal partial thickness burn injury, various durations (10min, 20min, 30min or 1h) and delays (immediate, 10min, 1h, 3h) of 15 degrees C running water first aid were applied to burns and compared to untreated controls. The subdermal temperatures were monitored during the treatment and wounds observed weekly for 6 weeks, for re-epithelialisation, wound surface area and cosmetic appearance. At 6 weeks after the burn, tissue biopsies were taken of the scar for histological analysis. Results showed that immediate application of cold running water for 20min duration is associated with an improvement in re-epithelialisation over the first 2 weeks post-burn and decreased scar tissue at 6 weeks. First aid application of cold water for as little as 10min duration or up to 1h delay still provides benefit.

A porcine deep dermal partial thickness burn model with hypertrophic scarring

We developed a reproducible model of deep dermal partial thickness burn injury in juvenile Large White pigs. The contact burn is created using water at 92 degrees C for 15s in a bottle with the bottom replaced with plastic wrap. The depth of injury was determined by a histopathologist who examined tissue sections 2 and 6 days after injury in a blinded manner. Upon creation, the circular wound area developed white eschar and a hyperaemic zone around the wound border. Animals were kept for 6 weeks or 99 days to examine the wound healing process. The wounds took between 3 and 5 weeks for complete re-epithelialisation. Most wounds developed contracted, purple, hypertrophic scars. On measurement, the thickness of the burned skin was approximately 1.8 times that of the control skin at week 6 and approximately 2.2 times thicker than control skin at 99 days after injury. We have developed various methods to assess healing wounds, including digital photographic analysis, depth of organising granulation tissue, immunohistochemistry, electron microscopy and tensiometry. Immunohistochemistry and electron microscopy showed that our porcine hypertrophic scar appears similar to human hypertrophic scarring. The development of this model allows us to test and compare different treatments on burn wounds.

Deep dermal burn injury results in scarless wound healing in the ovine fetus

Early to mid-term fetuses heal cutaneous incisional wounds without scars; however, fetal response to burn injury has not been ascertained. We present a fetal model of thermal injury and subsequent analysis of fetal and lamb response to burn injury. A reproducible deep dermal burn injury was created in the fetus by application of water at 66 degrees C for 7 seconds, and at 82 degrees C for 10 seconds to the lamb. Macroscopically, the area of fetal scald was undetectable from day 7 post injury, while all lamb scalds were readily identified and eventually healed with scarring. Using a five-point histopathology scoring system for alteration in tissue morphology, differences were detected between control and scalded skin at all stages in lamb postburn, but no difference was detected in the fetal model after day 7. There were also large differences in content of alpha-smooth muscle actin and transforming growth factor-beta1 between control and scalded lamb and these differences were statistically significant at day 14 (P < 0.01). This novel model of fetal and lamb response to deep dermal injury indicates that the fetus heals a deep burn injury in a scarless fashion. Further elucidation of this specific fetal process of burn injury repair may lead to improved outcome for patients with burn injury.

Genome-wide DNA methylation analysis of human brain tissue from schizophrenia patients

Recent studies suggest that genetic and environmental factors do not account for all the schizophrenia risk and epigenetics also plays a role in disease susceptibility. DNA methylation is a heritable epigenetic modification that can regulate gene expression. Genome-Wide DNA methylation analysis was performed on post-mortem human brain tissue from 24 patients with schizophrenia and 24 unaffected controls. DNA methylation was assessed at over 485 000 CpG sites using the Illumina Infinium Human Methylation450 Bead Chip. After adjusting for age and post-mortem interval (PMI), 4 641 probes corresponding to 2 929 unique genes were found to be differentially methylated. Of those genes, 1 291 were located in a CpG island and 817 were in a promoter region. These include NOS1, AKT1, DTNBP1, DNMT1, PPP3CC and SOX10 which have previously been associated with schizophrenia. More than 100 of these genes overlap with a previous DNA methylation study of peripheral blood from schizophrenia patients in which 27 000 CpG sites were analysed. Unsupervised clustering analysis of the top 3 000 most variable probes revealed two distinct groups with significantly more people with schizophrenia in cluster one compared to controls (p = 1.74x10-4). The first cluster was composed of 88% of patients with schizophrenia and only 12% controls while the second cluster was composed of 27% of patients with schizophrenia and 73% controls. These results strongly suggest that differential DNA methylation is important in schizophrenia etiology and add support for the use of DNA methylation profiles as a future prognostic indicator of schizophrenia.

The effect of varying diagnostic terminology within patient discharge information on expected mild traumatic brain injury outcome

This study aimed to determine if systematic variation of the diagnostic terminology embedded within written discharge information (i.e., concussion or mild traumatic brain injury, mTBI) would produce different expected symptoms and illness perceptions. We hypothesized that compared to concussion advice, mTBI advice would be associated with worse outcomes. Sixty-two volunteers with no history of brain injury or neurological disease were randomly allocated to one of two conditions in which they read a mTBI vignette followed by information that varied only by use of the embedded terms concussion (n = 28) or mTBI (n = 34). Both groups reported illness perceptions (timeline and consequences subscale of the Illness Perception Questionnaire-Revised) and expected Postconcussion Syndrome (PCS) symptoms 6 months post injury (Neurobehavioral Symptom Inventory, NSI). Statistically significant group differences due to terminology were found on selected NSI scores (i.e., total, cognitive and sensory symptom cluster scores (concussion > mTBI)), but there was no effect of terminology on illness perception. When embedded in discharge advice, diagnostic terminology affects some but not all expected outcomes. Given that such expectations are a known contributor to poor mTBI outcome, clinicians should consider the potential impact of varied terminology on their patients.

Spatial and developmental heterogeneity of calcium signaling in olfactory ensheathing cells

Olfactory ensheathing cells (OECs) are specialized glial cells in the mammalian olfactory system supporting growth of axons from the olfactory epithelium into the olfactory bulb. OECs in the olfactory bulb can be subdivided into OECs of the outer nerve layer and the inner nerve layer according to the expression of marker proteins and their location in the nerve layer. In the present study, we have used confocal calcium imaging of OECs in acute mouse brain slices and olfactory bulbs in toto to investigate physiological differences between OEC subpopulations. OECs in the outer nerve layer, but not the inner nerve layer, responded to glutamate, ATP, serotonin, dopamine, carbachol, and phenylephrine with increases in the cytosolic calcium concentration. The calcium responses consisted of a transient and a tonic component, the latter being mediated by store-operated calcium entry. Calcium measurements in OECs during the first three postnatal weeks revealed a downregulation of mGluR(1) and P2Y(1) receptor-mediated calcium signaling within the first 2 weeks, suggesting that the expression of these receptors is developmentally controlled. In addition, electrical stimulation of sensory axons evoked calcium signaling via mGluR(1) and P2Y(1) only in outer nerve layer OECs. Downregulation of the receptor-mediated calcium responses in postnatal animals is reflected by a decrease in amplitude of stimulation-evoked calcium transients in OECs from postnatal days 3 to 21. In summary, the results presented reveal striking differences in receptor responses during development and in axon-OEC communication between the two subpopulations of OECs in the olfactory bulb.

Investigation of brain derived neurotrophic factor (BDNF) gene variants in migraine

A number of observations have suggested that brain derived neurotrophic factor (BDNF) plays a role in migraine pathophysiology. This study investigates whether variants in the BDNF gene are associated with migraine in an Australian case-control population. Background. Brain derived neurotrophic factor (BDNF) has an important role in neural growth, development and survival in the central nervous system and is an important modulator of central and peripheral pain responses. Variants in BDNF, in particular the functional Val66Met polymorphism (rs6265), have been found to be associated with a number of psychiatric disorders, cognitive function and obesity. As BDNF has been found to be differentially expressed in a number of aspects related to migraine, we tested for association between single nucleotide polymorphisms (SNPs) in BDNF and migraine. Methods. Five SNPs in the BDNF locus (rs1519480, rs6265, rs712507, rs2049046 and rs12273363) were genotyped initially in a cohort of 277 migraine cases, including 172 diagnosed with migraine with aura (MA) and 105 with migraine without aura (MO), and 277 age- and sex-matched controls. Three of these SNPs (rs6265, rs2049046 and rs12273363) were subsequently genotyped in a second cohort of 580 migraineurs, including 473 diagnosed with MA and 105 with O, and 580 matched controls. Results. – BDNF SNPs rs1519480, rs6265, rs712507 and rs12273363 were not significantly associated with migraine. However, rs2049046 showed a significant association with migraine, and in particular, MA in the first cohort. In the second cohort, although an increase in the rs2049046 T-allele frequency was observed in migraine cases, and in both MA and MO subgroups, it was not significantly different from controls. Analysis of data combined from both cohorts for rs2049046 showed significant differences in the genotypic and allelic distributions for this marker in both migraine and the MA sub-group. Conclusion. This study confirmed previous studies that the functional BDNF SNP rs6265 (Val66Met) is not associated with migraine. However, we found that rs2049046, which resides at the 5’ end of 3 one the BDNF transcripts, may be associated with migraine, suggesting that further investigations of this SNP may be warranted.

Stimulation of osteogenic and angiogenic ability of cells on polymers by pulsed laser deposition of uniform akermanite-glass nanolayer

Polymer biomaterials have been widely used for bone replacement/regeneration because of their unique mechanical properties and workability. Their inherent low bioactivity makes them lack osseointegration with host bone tissue. For this reason, bioactive inorganic particles have been always incorporated into the matrix of polymers to improve their bioactivity. However, mixing inorganic particles with polymers always results in inhomogeneity of particle distribution in polymer matrix with limited bioactivity. This study sets out to apply the pulsed laser deposition (PLD) technique to prepare uniform akermanite (Ca2MgSi2O7, AKT) glass nanocoatings on the surface of two polymers (non-degradable polysulfone (PSU) and degradable polylactic acid (PDLLA)) in order to improve their surface osteogenic and angiogenic activity. The results show that a uniform nanolayer composed of amorphous AKT particles (∼30nm) of thickness 130nm forms on the surface of both PSU and PDLLA films with the PLD technique. The prepared AKT-PSU and AKT-PDLLA films significantly improved the surface roughness, hydrophilicity, hardness and apatite mineralization, compared with pure PSU and PDLLA, respectively. The prepared AKT nanocoatings distinctively enhance the alkaline phosphate (ALP) activity and bone-related gene expression (ALP, OCN, OPN and Col I) of bone-forming cells on both PSU and PDLLA films. Furthermore, AKT nanocoatings on two polymers improve the attachment, proliferation, VEGF secretion and expression of proangiogenic factors and their receptors of human umbilical vein endothelial cells (HUVEC). The results suggest that PLD-prepared bioceramic nanocoatings are very useful for enhancing the physicochemical, osteogenic and angiogenic properties of both degradable and non-degradable polymers for application in bone replacement/regeneration.