Early keratocyte apoptosis after epithelial scrape injury in the human cornea

Animal studies in mice, rats, rabbits, pigs and hens demonstrated that anterior keratocytes undergo programmed cell death or apoptosis after corneal epithelial injury. Many other wound healing changes subsequently follow the keratocyte apoptosis response. This study evaluated early keratocyte apoptosis after corneal epithelial scrape injury in human eyes scheduled for enucleation for malignancy. Two eyes had corneal epithelial scrape 1 h prior to the enucleation and another eye served as a control and had no corneal scrape prior to enucleation. One additional eye was enucleated, washed with balanced salt solution, and then had the corneal epithelium scraped 1 h prior to processing for analysis. Apoptosis was identified by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and confirmed by transmission electron microscopy (TEM). Anterior keratocyte apoptosis was detected in the three corneas that had epithelial scrape injury, but not in the control unwounded cornea. This study confirmed that keratocyte apoptosis is also an early response to corneal epithelial injury in humans and showed that tears are not essential for keratocyte apoptosis to occur in response to epithelial injury. (C) 2009 Elsevier Ltd. All rights reserved.

DNA extraction and quantification from touch and scrape preparations obtained from autopsy liver cells

The objective of the present study was to develop a simplified low cost method for the collection and fixation of pediatric autopsy cells and to determine the quantitative and qualitative adequacy of extracted DNA. Touch and scrape preparations of pediatric liver cells were obtained from 15 cadavers at autopsy and fixed in 95% ethanol or 3:1 methanol:acetic acid. Material prepared by each fixation procedure was submitted to DNA extraction with the Wizard® genomic DNA purification kit for DNA quantification and five of the preparations were amplified by multiplex PCR (azoospermia factor genes). The amount of DNA extracted varied from 20 to 8,640 µg, with significant differences between fixation methods. Scrape preparation fixed in 95% ethanol provided larger amount of extracted DNA. However, the mean for all groups was higher than the quantity needed for PCR (50 ng) or Southern blot (500 ng). There were no qualitative differences among the different material and fixatives. The same results were also obtained for glass slides stored at room temperature for 6, 12, 18 and 24 months. We conclude that touch and scrape preparations fixed in 95% ethanol are a good source of DNA and present fewer limitations than cell culture, tissue paraffin embedding or freezing that require sterile material, culture medium, laboratory equipment and trained technicians. In addition, they are more practical and less labor intensive and can be obtained and stored for a long time at low cost.

(Table T1) Abundance of radiolarian skeletons in scrape samples of ODP Hole 178-1098B

Palmer Deep is a series of three glacially overdeepened basins on the Antarctic Peninsula shelf, ~20 km southwest of Anvers Island. Site 1098 (64°51.72'S, 64°12.48'W) was drilled in the shallowest basin, Basin I, at 1012 m water depth. The sediment recovered was primarily laminated, siliceous, biogenic, pelagic muds alternating with siliciclastic hemipelagic sediments (Barker, Camerlenghi, Acton, et al., 1999). Sedimentation rates of 0.1725 cm/yr in the upper 25 m and 0.7-0.80 cm/yr in the lower 25 m of the core have been estimated from 14C (Domack et al., 2001). The oldest datable sediments have an age of ~13 ka and were underlain by diamicton sediments of the last glacial maximum (Domack et al., 2001). The large-scale water-mass distribution and circulation in the vicinity of Palmer Deep is dominated by Circumpolar Deep Water (CDW) below 200 m (Hofmann et al., 1996). Palmer Deep is too far from the coast to be influenced by glacial meltwater and cold-tongue generation associated with it (Domack and Williams, 1990; Dixon and Domack, 1991). Circulation patterns in the Palmer Deep area are not well understood, but evidence suggests southward flow across Palmer Deep from Anvers Island to Renaud Island (Kock and Stein, 1978). The water south of Anvers Island is nearly open with loose pack ice from February through May. The area is covered with sea ice beginning in June (Gloersen et al., 1992; Leventer et al., 1996). Micropaleontologic data from the work of Leventer et al. (1996) on a 9-m piston core has revealed circulation and climate patterns for the past 3700 yr in the Palmer Deep. The benthic foraminifer assemblage is dominated by two taxa, Bulimina aculeata and Bolivina pseudopunctata, which are inversely related. High relative abundances of B. aculeata occur cyclically over a period of ~230 yr. The assemblage associated with high abundance of B. aculeata in Palmer Deep resembles that from the Bellingshausen shelf, which is associated with CDW. In addition to the faunal evidence, hydrographic data indicate incursions of CDW into Palmer Deep (Leventer et al., 1996). A distinctive diatom assemblage dominated by a single genus was associated with peaks in B. aculeata, whereas a few different assemblages were associated with lows in B. aculeata. Leventer et al. (1996) interpreted the variability in diatom assemblages as an indication of changes in productivity associated with changes in water column stability. Abelmann and Gowing (1997) studied the horizontal and vertical distributions of radiolarians in the Atlantic sector of the Southern Ocean. They show that the spatial distribution of radiolarian assemblages reflects hydrographic boundaries. In a transect from the subtropical Atlantic to polar Antarctic zones, radiolarians in the upper 1000 m of the water column occurred in distinct surface and deep-living assemblages related to water depth, temperature, salinity, and nutrient content. Living assemblages resembled those preserved in underlying surface sediments (Abelmann and Gowing, 1997). Circumantarctic coastal sediments from neritic environments contained a distinctive assemblage dominated by the Phormacantha hystrix/Plectacantha oikiskos group and Rhizoplegma boreale (Nishimura et al., 1997). Low diversity and species compositions distinguished the coastal sediments from the typical pelagic Antarctic assemblages. Factors that controlled the assemblages were water depth, proximity to the coast, occurrence of sea ice, and steepness of topography, rather than temperature and salinity. Nishimura et al. (1997) found a gradient of sorts from deep-water sites containing diverse assemblages typical of pelagic environments to coastal sites with low diversity assemblages dominated by P. hystrix/P. oikiskos group and R. boreale. In general, sites between these two extremes had increased proportions of the coastal assemblage with decreasing water depth (Nishimura et al., 1997). At a site near Hole 1098 (GC905), they showed that the relative abundance of the coastal assemblage increased downcore (Nishimura et al., 1997). The purpose of the research presented here was to make a cursory investigation into the radiolarian assemblages as possible paleoenvironmental indicators.

Multifractality in plasma edge electrostatic turbulence

Plasma edge turbulence in Tokamak Chauffage Alfven Bresilien (TCABR) [R. M. O. Galvao et al., Plasma Phys. Contr. Fusion 43, 1181 (2001)] is investigated for multifractal properties of the fluctuating floating electrostatic potential measured by Langmuir probes. The multifractality in this signal is characterized by the full multifractal spectra determined by applying the wavelet transform modulus maxima. In this work, the dependence of the multifractal spectrum with the radial position is presented. The multifractality degree inside the plasma increases with the radial position reaching a maximum near the plasma edge and becoming almost constant in the scrape-off layer. Comparisons between these results with those obtained for random test time series with the same Hurst exponents and data length statistically confirm the reported multifractal behavior. Moreover, the persistence of these signals, characterized by their Hurst exponent, present radial profile similar to the deterministic component estimated from analysis based on dynamical recurrences. (C) 2008 American Institute of Physics.

Bicoherence in electrostatic turbulence driven by high magnetohydrodynamic activity in Tokamak Chauffage Alfven Bresilien

During some discharges in Tokamak Chauffage Alfven Bresilien [R. M. O. Galvao et al., Plasma Phys. Controlled Fusion 43, 1181 (2001)] high magnetohydrodynamic activity may appear with a peaked frequency spectrum. Whenever this peak occurs, the ambient broadband electrostatic turbulence is remarkably modified, synchronizing into the dominant magnetic fluctuation frequency and presenting high bicoherence in the whole plasma edge with a maximum bicoherence inside the plasma. A phenomenological model is introduced to investigate this driven turbulence bicoherence, consisting of nonlinearly coupled phase-randomized drift modes with time-periodic external driving at the dominant magnetic fluctuation frequency. The bicoherence spectrum of this model can mimic features of the experimental results. (C) 2009 American Institute of Physics. [DOI: 10.1063/1.3099701]

Reduction of chaotic particle transport driven by drift waves in sheared flows

Investigations of chaotic particle transport by drift waves propagating in the edge plasma of tokamaks with poloidal zonal flow are described. For large aspect ratio tokamaks, the influence of radial electric field profiles on convective cells and transport barriers, created by the nonlinear interaction between the poloidal flow and resonant waves, is investigated. For equilibria with edge shear flow, particle transport is seen to be reduced when the electric field shear is reversed. The transport reduction is attributed to the robust invariant tori that occur in nontwist Hamiltonian systems. This mechanism is proposed as an explanation for the transport reduction in Tokamak Chauffage Alfven Bresilien [R. M. O. Galvao , Plasma Phys. Controlled Fusion 43, 1181 (2001)] for discharges with a biased electrode at the plasma edge.

Electrostatic turbulence driven by high magnetohydrodynamic activity in Tokamak Chauffage Alfven Bresilien

In Tokamak Chauffage Alfven Bresilien [R. M. O. Galvao , Plasma Phys. Controlled Fusion 43, 1181 (2001)], high magnetohydrodynamic (MHD) activity may appear spontaneously or during discharges with a voltage biased electrode inserted at the plasma edge. The turbulent electrostatic fluctuations, measured by Langmuir probes, are modulated by Mirnov oscillations presenting a dominant peak with a common frequency around 10 kHz. We report the occurrence of phase locking of the turbulent potential fluctuations driven by MHD activity at this frequency. Using wavelet cross-spectral analysis, we characterized the phase and frequency synchronization in the plasma edge region. We introduced an order parameter to characterize the radial dependence of the phase-locking intensity. (c) 2008 American Institute of Physics.

Corneal myofibroblast generation from bone marrow-derived cells

The purpose of this study was to determine whether bone marrow-derived cells can differentiate into myofibroblasts, as defined by alpha-smooth muscle actin (SMA) expression, that arise in the corneal stroma after irregular phototherapeutic keratectomy and whose presence within the cornea is associated with corneal stromal haze. C578L/6J-GFP chimeric mice were generated through bone marrow transplantation from donor mice that expressed enhanced green fluorescent protein (GFP) in a high proportion of their bone marrow-derived cells. Twenty-four GFP chimeric mice underwent haze-generating corneal epithelial scrape followed by irregular phototherapeutic keratectomy (PTK) with an excimer laser in one eye. Mice were euthanized at 2 weeks or 4 weeks after PTK and the treated and control contralateral eyes were removed and cryo-preserved for sectioning for immunocytochemistry. Double immunocytochemistry for GFP and myofibroblast marker alpha-smooth muscle actin (SMA) were performed and the number of SMA+GFP+, SMA+GFP, SMA-GFP+ and SMA GFP cells, as well as the number of DAPI+ cell nuclei, per 400x field of stroma was determined in the central, mid-peripheral and peri-limbal cornea. In this mouse model, there were no SMA+ cells and only a few GFP+ cells detected in unwounded control corneas. No SMA+ cells were detected in the stroma at two weeks after irregular PTK, even though there were numerous GFP+ cells present. At 4 weeks after irregular PTK, all corneas developed mild to moderately severe corneal haze. In each of the three regions of the corneas examined, there were on average more than 9x more SMA+GFP+ than SMA+GFP myofibroblasts. This difference was significant (p < 0.01). There were significantly more (p < 0.01) SMA GFP+ cells, which likely include inflammatory cells, than SMA+GFP+ or SMA+GFP cells, although SMA GFP cells represent the largest population of cells in the corneas. In this mouse model, the majority of myofibroblasts developed from bone marrow-derived cells. It is possible that all myofibroblasts in these animals developed from bone marrow-derived cells since mouse chimeras produced using this method had only 60-95% of bone marrow-derived cells that were GFP+ and it is not possible to achieve 100% chimerization. This model, therefore, cannot exclude the possibility of myofibroblasts also developed from keratocytes and/or corneal fibroblasts. (C) 2010 Elsevier Ltd. All rights reserved.

Topical interleukin-1 receptor antagonist inhibits inflammatory cell infiltration into the cornea

Interleukin (IL)-1 alpha and beta are important modulators of many functions of corneal epithelial and stromal cells that occur following injury to the cornea, including the influx of bone marrow-derived inflammatory cells into the stroma attracted by chemokines released from the stroma and epithelium. In this study, we examined the effect of topical soluble IL-1 receptor antagonist on bone marrow-derived cell influx following corneal epithelial scrape injury in a mouse model. C57BL/6 mice underwent corneal epithelial scrape followed by application of IL-1 receptor antagonist (Amgen, Thousand Oaks, CA) at a concentration of 20 mg/ml or vehicle for 24 h prior to immunocytochemical detection of marker CD11b-positive cells into the stroma. In two experiments, topical IL-1 receptor antagonist had a marked effect in blocking cell influx. For example, in experiment 1, topical IL-1 receptor antagonist markedly reduced detectible CD11b-positive cells into the corneal stroma at 24 It after epithelial injury compared with the vehicle control (3.5 +/- 0.5 (standard error of the mean) cells/400x field and 13.9 +/- 1.2 cells/400x field, respectively, p < 0.01). A second experiment with a different observer performing cell counting had the same result. Thus, the data demonstrate conclusively that topical IL-1 receptor antagonist markedly down-regulates CD-11b-positive monocytic cell appearance in the corneal stroma. Topical IL-1 receptor antagonist could be an effective adjuvant for clinical treatment of corneal conditions in which unwanted inflammation has a role in the pathophysiology of the disorder. (c) 2008 Elsevier Ltd. All rights reserved.

Doramectin as a treatment for canine and feline demodicosis

Twenty-three dogs and three cats with generalised demodicosis were treated with weekly subcutaneous injections of doramectin at a dose of 600mcg/kg body weight. All dogs and cats responded to treatment by going into remission. The median time until skin scrape results were negative was eight weeks (range five to 20 weeks). Ten dogs remained in remission after the first treatment, five were lost to follow up and seven needed a second course of doramectin or were maintained in remission by monthly injections. Time to when skin scrape results were negative for the cats was two to three weeks with one cat still in remission four years later. The other two cats were euthanased due to their underlying disease whilst only four and six months into remission. Weekly subcutaneous doramectin injections are useful in the treatment of generalised demodicosis in dogs and cats. No systemic side effects of the drug were seen, the injection was painless and no reactions were seen at the sites.

Connexins 43 and 26 are differentially increased after rat bladder outlet obstruction.

To evaluate the regulation of connexin expression by fluid pressure, we have studied the effects of elevated transmural urine pressure on Connexin43 (Cx43) and Cx26. We chose to focus on these two proteins out of the five connexins (Cx26, 43, 40, 37, and 45) which we found by RT-PCR to be expressed in the rat bladder, since in situ hybridization and immunofluorescence showed that Cx43 is the predominant connexin expressed by smooth muscle cells (SMC), whereas Cx26 is abundantly expressed only in the latter cell type. To evaluate whether these connexins are affected by changes in transmural urine pressure, we used a rat model of bladder outlet obstruction, in which a ligature is placed around the urethra. Under conditions of increased fluid pressure due to urine retention, we observed that the expression of both Cx43 and Cx26 increased at both transcript and protein levels, reaching a maximum 7-9 h after the ligature. Further analysis revealed that these changes were accounted for by a fourfold increase in Cx43 mRNA of SMC but not urothelial cell and by a fivefold increase in Cx26 mRNA of urothelium. Scrape-loading of propidium iodide showed that the latter change was paralleled by a twofold increase in coupling between urothelial cells. The data show that Cx43 and Cx26 are differentially regulated during bladder outlet obstruction and contribute to the response of the bladder wall to increased voiding pressure, possibly to control its elasticity.

Optimal Usage of De-Icing Chemicals when Scraping Ice, November 2003

One of the challenges that faces the winter maintainer is how much chemical to apply to the road under given conditions. Insufficient chemical can lead to the road surface becoming slick, and the road thus becoming unsafe. In all likelihood, additional applications will have to be made, requiring additional effort and use of resources. However, too much chemical can also be bad. While an excess of chemical will ensure (in most circumstances) that a safe road condition is achieved, it may also result in a substantial waste of chemical (with associated costs for this waste) and in ancillary damage to the road itself and to the surrounding environment. Ideally, one should apply what might be termed the “goldilocks” amount of chemical to the road: Not too much, and not too little, but just right. Of course the reality of winter maintenance makes achieving the “goldilocks” application rate somewhat of a fairy tale. In the midst of a severe storm, when conditions are poor and getting worse, the last thing on a plow operator’s mind is a minute adjustment in the amount of chemical being applied to the road. However, there may be considerable benefit and substantial savings to be achieved if chemical applications can be optimized to some degree, so that wastage is minimized without compromising safety. The goal of this study was to begin to develop such information through a series of laboratory studies in which the force needed to scrape ice from concrete blocks was measured, under a variety of chemical application conditions.

Field Measurements of Ice Scraping Loads on Front Mounted Plow Blades, HR-374, 1997

Removal of ice from roads is of the more challenging task in winter highway maintenance. The best mechanical method is to use a truck with underbody plow blade, but such equipment is not available to all agencies charged with winter maintenance operations. While counties and cities often use motor graders to scrape ice, it would be of great benefit if front mounted plows could be used effectively for ice removal. To reveal and understand the factors that influence the performance of these plows, measurement of the forces experienced by the plow blades during ice scraping is desirable. This study explores the possibility of using accelerometers to determine the forces on a front-mounted plow when scraping ice. The plow was modeled by using a dynamic approach. The forces on the plow were to be determined by the measurement of the accelerations of the plow. Field tests were conducted using an "as is" front-mounted plow instrumented with accelerometers. The results of the field tests indicate that in terms of ice removal, the front-mounted plow is not favorable equipment. The major problem in this study is that the front mounted plow was not able to cut ice, and therefore experienced no significant scraping forces. However, the use of accelerometers seems to be promising for analyzing the vibration problems of the front-mounted plow.

Field Measurements of Plow Loads During Ice Removal Operations, HR-334, 1993

One of the more severe winter hazards is ice or compacted snow on roadways. While three methods are typically used to combat ice (salting, sanding and scraping), relatively little effort has been applied to improve methods of scraping ice from roads. In this project, a new test facility has been developed, comprising a truck with an underbody blade, which has been instrumented such that the forces to scrape ice from a pavement can be measured. A test site has been used, which is not accessible to the public, and ice covers have been sprayed onto the pavement and subsequently scraped from it, while the scraping loads have been recorded. Three different cutting edges have been tested for their ice scraping efficiency. Two of the blades are standard (one with a carbide insert, the other without) while the third blade was designed under the SHRP H-204A project. Results from the tests allowed two parameters to be identified. The first is the scraping efficiency which is the ratio of vertical to horizontal force. The lower this ratio, the more efficiently ice is being removed. The second parameter is the scraping effectiveness, which is related (in some as yet unspecified manner) to the horizontal load. The higher the horizontal load, the more ice is being scraped. The ideal case is thus to have as high a horizontal load as possible, combined with the lowest possible vertical load. Results indicate that the SHRP blade removed ice more effectively than the other two blades under equivalent conditions, and furthermore, did so with greater efficiency and thus more control. Furthermore, blade angles close to 0 deg provide for the most efficient scraping for all three blades. The study has shown that field testing of plow blades is possible in controlled situations, and that blades can be evaluated using this system. The system is available for further tests as are deemed appropriate.

Fractal structures in nonlinear plasma physics

Fractal structures appear in many situations related to the dynamics of conservative as well as dissipative dynamical systems, being a manifestation of chaotic behaviour. In open area-preserving discrete dynamical systems we can find fractal structures in the form of fractal boundaries, associated to escape basins, and even possessing the more general property of Wada. Such systems appear in certain applications in plasma physics, like the magnetic field line behaviour in tokamaks with ergodic limiters. The main purpose of this paper is to show how such fractal structures have observable consequences in terms of the transport properties in the plasma edge of tokamaks, some of which have been experimentally verified. We emphasize the role of the fractal structures in the understanding of mesoscale phenomena in plasmas, such as electromagnetic turbulence.