Myoglobin toxicity in proximal human kidney cells: Roles of Fe, Ca2+, H2O2, and terminal mitochondrial electron transport

Myoglobin toxicity in proximal human kidney cells: Roles of Fe, Ca2+, H2O2, and terminal mitochondrial electron transport. The purpose of this study was to gain direct insights into mechanisms by which myoglobin induces proximal tubular cell death. To avoid confounding systemic and hemodynamic influences, an in vitro model of myoglobin cytotoxicity was employed. Human proximal tubular (HK-2) cells were incubated with 10 mg/ml myoglobin, and after 24 hours the lethal cell injury was assessed (vital dye uptake; LDH release). The roles played by heme oxygenase (HO), cytochrome p450, free iron, intracellular Ca2+, nitric oxide, H2O2, hydroxyl radical (·OH), and mitochondrial electron transport were assessed. HO inhibition (Sn protoporphyrin) conferred almost complete protection against myoglobin cytotoxicity (92% vs. 22% cell viability). This benefit was fully reproduced by iron chelation therapy (deferoxamine). Conversely, divergent cytochrome p450 inhibitors (cimetidine, aminobenzotriazole, troleandomycin) were without effect. Catalase induced dose dependent cytoprotection, virtually complete, at a 5000 U/ml dose. Conversely, ·OH scavengers (benzoate, DMTU, mannitol), xanthine oxidase inhibition (oxypurinol), superoxide dismutase, and manipulators of nitric oxide expression (L-NAME, L-arginine) were without effect. Intracellular (but not extracellular) calcium chelation (BAPTA-AM) caused ∼50% reductions in myoglobin-induced cell death. The ability of Ca2+ (plus iron) to drive H2O2 production (phenol red assay) suggests one potential mechanism. Blockade of site 2 (antimycin) and site 3 (azide), but not site 1 (rotenone), mitochondrial electron transport significantly reduced myoglobin cytotoxicity. Inhibition of Na,K-ATPase driven respiration (ouabain) produced a similar protective effect. We conclude that: (1) HO-generated iron release initiates myoglobin toxicity in HK-2 cells; (2) myoglobin, rather than cytochrome p450, appears to be the more likely source of toxic iron release; (3) H2O2 generation, perhaps facilitated by intracellular Ca2+/iron, appears to play a critical role; and (4) cellular respiration/terminal mitochondrial electron transport ultimately helps mediate myoglobin's cytotoxic effect. Formation of poorly characterized toxic iron/H2O2-based reactive intermediates at this site seems likely to be involved

Decreased expression of mitochondrial-derived H2O2 and hydroxyl radical in cytoresistant proximal tubules

Decreased expression of mitochondrial-derived H2O2 and hydroxyl radical in cytoresistant proximal tubules. Increased production of reactive oxygen metabolites (ROM) can contribute to the initiation phase of nephrotoxic and ischemic acute renal failure (ARF). However, whether altered ROM expression also exists during the maintenance phase of ARF has not been adequately assessed. Since diverse forms of tubular injury can initiate a “cytoresistant state,” this study tested whether a down-regulation of ROM expression might develop in the aftermath of acute tubular damage, potentially limiting renal susceptibility to further attack. To test this hypothesis, rats were subjected to either mild myohemoglobinuria (glycerol injection) or bilateral ureteral obstruction and 24 hours later, cytoresistant proximal tubular segments (PTS) were isolated to assess ROM expression. PTS from sham operated rats were used to establish normal values. Both sets of cytoresistant PTS manifested ∼ 75% reductions in H2O2 levels, as assessed by the phenol red/horseradish peroxidase technique (P < 0.01 to 0.001). A 40% reduction in hydroxyl radical (˙OH) levels was also observed (salicylate trap method), thereby substantiating decreased oxidant stress in cytoresistant PTS. Catalase, glutathione peroxidase, and free iron levels were comparable in control and cytoresistant PTS, suggesting that decreased H2O2 production (such as by mitochondria) was the cause of the decreased oxidant stress. To test this latter hypothesis, H2O2 expression by control and cytoresistant PTS was assessed in the presence of respiratory chain inhibitors. Although site 1 and site 3 inhibition markedly suppressed H2O2 production in control PTS, they had no impact on H2O2 production in cytoresistant PTS, implying that production at these sites was already maximally suppressed. Correlates of the decreased mitochondrial H2O2 production were improvements in cell energetics (increased ATP/ADP ratios with Na ionophore treatment) and ∼ 40 to 90% increases in PTS/renal cortical glutathione content. We conclude that: (1) proximal tubule H2O2/˙OH expression can be down-regulated during the maintenance phase of ARF; (2) this seemingly reflects a decrease in mitochondrial ROM generation; and (3) the associated improvements in glutathione content and/or cellular energetics could conceivably contribute to a post-injury cytoresistant state

The Csk-binding protein PAG regulates PDGF-induced Src mitogenic signaling via GM1

Spatial regulation is an important feature of signal specificity elicited by cytoplasmic tyrosine kinases of the Src family (SRC family protein tyrosine kinases [SFK]). Cholesterol-enriched membrane domains, such as caveolae, regulate association of SFK with the platelet-derived growth factor receptor (PDGFR), which is needed for kinase activation and mitogenic signaling. PAG, a ubiquitously expressed member of the transmembrane adaptor protein family, is known to negatively regulate SFK signaling though binding to Csk. We report that PAG modulates PDGFR levels in caveolae and SFK mitogenic signaling through a Csk-independent mechanism. Regulation of SFK mitogenic activity by PAG requires the first N-terminal 97 aa (PAG-N), which include the extracellular and transmembrane domains, palmitoylation sites, and a short cytoplasmic sequence. We also show that PAG-N increases ganglioside GM1 levels at the cell surface and, thus, displaces PDGFR from caveolae, a process that requires the ganglioside-specific sialidase Neu-3. In conclusion, PAG regulates PDGFR membrane partitioning and SFK mitogenic signaling by modulating GM1 levels within caveolae independently from Csk

Ethyl 4-{1-[(2,4-dinitro­phen­yl)hydrazono]eth­yl}-5-(2-naphthyl­methoxy­meth­yl)isoxazole-3-carboxyl­ate

The title compound, C26H23N5O8, was prepared and its structure investigated to further develop a working hypothesis for the essential binding pharmacophore for ligands of the System Xc- transporter [Patel et al. (2004 ▶). Neuropharmacology, 46, 273–284]. The hydrazone group displays an E geometry and the isoxazole double bond and C=N group of the hydrazone are in an s-cis relationship. The secondary amino NH group forms an intra­molecular N—H⋯O hydrogen bond to a ring nitro group. There is a dihedral angle of 44.27 (5)° between the isoxazole plane and the hydrazone group plane

Savor the Cryosphere

This article provides concise documentation of the ongoing retreat of glaciers, along with the implications that the ice loss presents, as well as suggestions for geoscience educators to better convey this story to both students and citizens. We present the retreat of glaciers—the loss of ice—as emblematic of the recent, rapid contraction of the cryosphere. Satellites are useful for assessing the loss of ice across regions with the passage of time. Ground-based glaciology, particularly through the study of ice cores, can record the history of environmental conditions present during the existence of a glacier. Repeat photography vividly displays the rapid retreat of glaciers that is characteristic across the planet. This loss of ice has implications to rising sea level, greater susceptibility to dryness in places where people rely upon rivers delivering melt water resources, and to the destruction of natural environmental archives that were held within the ice. Warming of the atmosphere due to rising concentrations of greenhouse gases released by the combustion of fossil fuels is causing this retreat. We highlight multimedia productions that are useful for teaching this story effectively. As geoscience educators, we attempt to present the best scholarship as accurately and eloquently as we can, to address the core challenge of conveying the magnitude of anthropogenic impacts, while also encouraging optimistic determination on the part of students, coupled to an increasingly informed citizenry. We assert that understanding human perturbation of nature, then choosing to engage in thoughtful science-based decision-making, is a wise choice. This topic comprised “Savor the Cryosphere,” a Pardee Keynote Symposium at the 2015 Annual Meeting in Baltimore, Maryland, USA, for which the GSA recorded supporting interviews and a webinar

A Density-Based Approach for Leaf Area Index Assessment in a Complex Forest Environment Using a Terrestrial Laser Scanner

Forests are an important part natural ecosystems, by for example providing food, fiber, habitat, and biodiversity, all of which contribute to stable natural systems. Assessing and modeling the structure and characteristics of forests, e.g., Leaf Area Index (LAI), volume, biomass, etc., can lead to a better understanding and management of these resources. In recent years, Terrestrial Laser Scanning (TLS) has been recognized as a tool that addresses many of the limitations of manual and traditional forest data collection methods. In this study, we propose a density-based approach for estimating the LAI in a structurally-complex forest environment, which contains variable and diverse structural attributes, e.g., non-circular stem forms, dense canopy and below-canopy vegetation cover, and a diverse species composition. In addition, 242 TLS scans were collected using a portable low-cost scanner, the Compact Biomass Lidar (CBL), in the Hawaii Volcanoes National Park (HAVO), Hawaii Island, USA. LAI also was measured for 242 plots in the site, using an AccuPAR LP-80 ceptometer. The first step after cleaning the point cloud involved detecting the higher forest canopy in the light detection and ranging (lidar) point clouds, using normal change rate assessment. We then estimated Leaf Area Density (LAD), using a voxel-based approach, and divided the canopy point cloud into five layers in the Z (vertical) direction. These five layers subsequently were divided into voxels in the X direction, where the size of these voxels were obtained based on inter-quartile analysis and the number of points in each voxel. We hypothesized that the intensity returned to the lidar system from woody materials, like branches, would be higher than from leaves, due to the liquid water absorption feature of the leaves and higher reflectance for woody material at the 905 nm laser wavelength. We also differentiated between foliar and woody materials using edge detection in the images from projected point clouds and evaluated the density of these regions to support our hypothesis. Density of points, or the number of points divided by the volume of a grid, in a 3D grid size of 0.1 m, was calculated for each of the voxels. The grid size was determined by investigating the size of the branches in the lower portion of the canopy. Subsequently, we fitted a Kernel Density Estimator (KDE) to these values, with the threshold set based on half of the area under the curve in each of the density distributions. All the grids with a density below the threshold were labeled as leaves, while those grids above the threshold were identified as non-leaves. Finally, we modeled LAI using the point densities derived from the TLS point clouds and the listed analysis steps. This model resulted in an R 2 value of 0.88. We also estimated the LAI directly from lidar data using the point densities and calculating LAD, which is defined as the total one-sided leaf area per unit volume. LAI can be obtained as the sum of the LAD values in all the voxels. The accuracy of LAI estimation was 90%, with an RMSE value of 0.31, and an average overestimation of 9% in TLS-derived LAI, when compared to field-measured LAI. Algorithm performance mainly was affected by the vegetation density and complexity of the canopy structures. It is worth noting that, since the LAI values cannot be considered spatially independent throughout all the plots in this site, we performed semivariogram analysis on the field-measured LAI data. This analysis showed that the LAI values can be assumed to be independent in plots that are at least 30 m apart. As a result, we divided the data into six subsets in which the plots were 30 m spaced. The R 2 values for these subsets, based on modeling of the field-measured LAI using leaf point density values, ranged between 0.84–0.96. The results bode well for using this method for efficient, automatic, and accurate/precise estimation of LAI values in complex forest environments, using a low-cost, rapid-scan TLS

AmFm and lithium gap stars: Stellar evolution models with mass loss

A thorough study of the effects of mass loss on internal and surface abundances of A and F stars is carried out in order to constrain mass loss rates for these stars, as well as further elucidate some of the processes which compete with atomic diffusion. Self-consistent stellar evolution models of 1.3 to 2.5 M_sun stars including atomic diffusion and radiative accelerations for all species within the OPAL opacity database were computed with mass loss and compared to observations as well as previous calculations with turbulent mixing. Models with unseparated mass loss rates between 5 x 10^-14 and 10^-13 M_sun/yr reproduce observations for many cluster AmFm stars as well as Sirius A and o Leonis. These models also explain cool Fm stars, but not the Hyades lithium gap. Like turbulent mixing, these mass loss rates reduce surface abundance anomalies; however, their effects are very different with respect to internal abundances. For most of the main sequence lifetime of an A or F star, surface abundances in the presence of such mass loss depend on separation which takes place between log(Delta M/M_star)= -6 and -5. The current observational constraints do not allow us to conclude that mass loss is to be preferred over turbulent mixing (induced by rotation or otherwise) in order to explain the AmFm phenomenon. Internal concentration variations which could be detectable through asteroseismic tests should provide further information. If atomic diffusion coupled with mass loss are to explain the Hyades Li gap, the wind would need to be separated.Comment: 27 pages, 25 figures, accepted for publication in A&

Chemical composition of A and F dwarf members of the Coma Berenices open cluster

Abundances of 18 chemical elements have been derived for 11 A (normal and Am) and 11 F dwarfs members of the Coma Berenices open cluster in order to set constraints on evolutionary models including transport processes (radiative and turbulent diffusion)calculated with the Montreal code. A spectral synthesis iterative procedure has been applied to derive the abundances from selected high quality lines in high resolution high signal-to-noise echelle spectra obtained with ELODIE at the Observatoire de Haute Provence. The chemical pattern found for the A and F dwarfs in Coma Berenices is reminiscent of that found in the Hyades and the UMa moving group. In graphs representing the abundances [X/H] versus the effective temperature, the A stars often display abundances much more scattered around their mean values than the F stars do. Large star-to-star variations are detected for A stars in their abundances which we interpret as evidence of transport processes competing with radiative diffusion. The F stars have solar abundances for almost all elements except for Mg, Si, V and Ba. The derived abundances patterns, [X/H] versus atomic number, for the slow rotator HD108642 (A2m) and the moderately fast rotator HD106887 (A4m) were compared to the predictions of self consistent evolutionary model codes including radiative and different amounts of turbulent diffusion. None of the models reproduces entirely the overall shape of the abundance pattern. While part of the discrepancies between derived and predicted abundances may be accounted for by non-LTE effects, the inclusion of competing processes such as rotational mixing in the radiative zones of these stars seems necessary to improve the agreement between observed and predicted abundance patterns.Comment: 25 pages, 20 figure

Experience With the Cardiac Surgery Simulation Curriculum: Results of the Resident and Faculty Survey

BACKGROUND: The Cardiac Surgery Simulation Curriculum was developed at 8 institutions from 2010 to 2013. A total of 27 residents were trained by 18 faculty members. A survey was conducted to gain insight into the initial experience. METHODS: Residents and faculty were sent a 72- and 68-question survey, respectively. In addition to demographic information, participants reported their view of the overall impact of the curriculum. Focused investigation into each of the 6 modules was obtained. Participants evaluated the value of the specific simulators used. Institutional biases regarding implementation of the curriculum were evaluated. RESULTS: Twenty (74%) residents and 14 (78%) faculty responded. The majority (70%) of residents completed this training in their first and second year of traditional-track programs. The modules were well regarded with no respondents having an unfavorable view. Both residents and faculty found low, moderate, and high fidelity simulators to be extremely useful, with particular emphasis on utility of high fidelity components. The vast majority of residents (85%) and faculty (100%) felt more comfortable in the resident skill set and performance in the operating room. Simulation of rare adverse events allowed for development of multidisciplinary teams to address them. At most institutions, the conduct of this curriculum took precedence over clinical obligations (64%). CONCLUSIONS: The Cardiac Surgery Simulation Curriculum was implemented with robust adoption among the investigating centers. Both residents and faculty viewed the modules favorably. Using this curriculum, participants indicated an improvement in resident technical skills and were enthusiastic about training in adverse events and crisis management

The Grizzly, October 6, 1992

Amen, It\u27s Over!: Congratulations to the Sorority Pledge Classes of 1992 • New Party Policies • UC Grad Makes Scientific Breakthrough • Freedom of Press Forum • A Night to Remember • Berman To Exhibit Oriental Photographs • Homecoming Queen Nominees • A Need for RICO • Letter to the Editor • Men\u27s Cross-Country Fights Tough Competitionhttps://digitalcommons.ursinus.edu/grizzlynews/1300/thumbnail.jp