Fractional Release Research Articles

Characterization of RyR2 Mutations Located at the Caffeine and FKBP Binding Sites in hiPSC-CMs

Signaling of Ca2+ release from RyR2 is a critical step in cardiac EC-coupling. To gain insight into regulatory mechanisms of Ca2+ release channel (RyR2), we introduced CPVT1-associated W4645R mutation (caffeine-binding site) and N771D mutation (FKBP binding site) corresponding with skeletal disease RyR1 mutation in RYR2 gene of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) by CRISPR/Cas9 gene-editing. Simultaneous Ca2+ imaging and electrophysiological measurement in patch clamped hiPSC-CMs showed: (1) there was no significant differences in activation threshold, reversal potential or voltage-dependent activation and inactivation of Ca2+ current in the two mutants as compared to wild type (WT) myocytes. (2) The W4645R mutant was unresponsive to rapid caffeine (5-20mM) applications, but showed 60% higher ICa-gated Ca2+ release as compared to WT myocytes. (3) N771D mutant had similar fractional release, higher CICR gain, but unexpectedly lower diastolic Ca2+ leak compared to WT cells; and (4) β-adrenergic stimulation (500nM isoproterenol) normalized the significantly lower amplitude and frequency of spontaneous cytosolic Ca2+ oscillations in W4645R, but had no differential effect on N771D mutant compared to WT cells. Our data suggests that W4645R mutation abolishes caffeine-triggered release, but increases ICa-gated Ca2+ release possibly related to enhanced Ca2+ sensitivity of RyR2 as shown in [3H]ryanodine binding assay with recombinant W4645R-RyR2. N771D-RyR2 mutation surprisingly did not show any aberrancy in Ca2+ signaling in sharp contrast to the finding with the truncated RyR protein, showing impaired FKBP binding. Supported by NIH grant HL147054.

Leaching behavior of prototypical corium samples: A step to understand the interactions between the fuel debris and water at the Fukushima Daiichi reactors

On 11 March 2011, the Great East Japan Earthquake occurred and caused a severe nuclear accident at the Fukushima Daiichi Nuclear Power Station (1F). A policy for corium debris retrieval will be decided in the near future. To this end, debris management after removal from reactors must be optimized. From the viewpoint of cost and waiting for definitive solutions, interim or long-term underwater storage is one of the considered options after debris removal. Leaching data on the long term behavior of debris in water are therefore needed before debris removal. In addition, it is important to understand potential differences in leaching behaviors between debris and Spent Fuel (SF) to get comparative elements.For these reasons, simulated in-vessel and ex-vessel corium debris, fabricated in the Colima experimental facility set up in the PLINIUS platform at CEA Cadarache, were selected and leaching experiments were carried out under oxidizing conditions. In parallel, geochemical modeling was performed to better understand the experimental concentrations, pH evolutions and secondary phase’s formation.Finally, the Fractional Release Rates of the (U, Zr)O2 matrix for the two kinds of samples (in-vessel and ex-vessel) were found to be close to or one order of magnitude lower than that of SF under oxidizing conditions (from 10−6 to 10−7 per day) but the release processes are different. Indeed, the Fraction of Inventory in Aqueous Phase of Mo, Ba, Sr, Rh, La, Nd, Ce and Y tended to be higher than that of uranium and zirconium in this study whatever the element considered. The release of simulating PFs, including low volatile element like trivalent rare earths, does not appear to be fully controlled by that of the (U, Zr)O2 matrix from a macroscopic and average point of view. The FPs could be partially located outside the (U, Zr)O2 solid solution as a function of the corium history (quenching rate and thermal history, chemical interactions…). The location of inventories and their chemical forms are not very well known moreover uncertainties related to heterogeneity are important and may also impact the calculation of the global released fractions over time. For the ex-vessel sample, releases into the solution of the main concrete elements (Si, Ca, Mg) seem well correlated and dissociated from the behavior of the (U, Zr)O2 phase. Further studies are needed especially on real or prototypical corium approaching the conditions of the Fukushima accident. The study of simpler and model systems could also provide information on the reactivity of the (U, Zr)O2 matrix.

Design and characterisation of PHBV-magnesium oleate directional nanofibers for neurosupport

The focus of significance in neuronal repair strategies is the design of scaffold systems capable of promoting neuronal regeneration and directional guidance via provision of a biomimetic environment resemblance of native neural tissue. The purpose of this study was to synthesize triple-cue electrospun aligned nanofibrous films (physical cue) of poly(3-hyroxybutyric acid-co-3-hydroxyvaleric acid) (PHBV) blended with magnesium-oleate (MgOl) (chemical cue) and N-acetyl-L-cysteine (NAC) (therapeutic cue) with potential incorporation into hollow nerve guidance conduits for an enhanced regenerative strategy. A Box–Behnken experimental design of 15 formulations, were analysed for crystallinity, textural properties and in vitro water-uptake, erosion, NAC-release and PC12 cell viability. Nucleating effects of MgOl provided tuning of PHBV electrospinning-induced crystallinity and mechanical properties. Tensile strengths and deformation moduli of ±12 MPa and ±7 MP, respectively, were attainable, thereby matching native nerve mechanics. Crystallinity changes ascribed differing release kinetics to NAC over 30 d: diffusion-based (42%–58% crystallinity with 33%–47% fractional release) and polymer-relaxational (59%–65% crystallinity with 60%–82% fractional release). The synergistic activity of MgOl and NAC increased PC12 proliferation by 32.6% compared to the control. MgOl produced dual actions as non-toxic plasticiser and PC12 cell proliferation-promoter via nucleation and neurotrophic-like effects, respectively. Controlled release of NAC imparted neuro-protectant effects on PC12 cells and promoted neurite extension, thus, making electrospun PHBV-MgOl nanofibrous films a versatile and promising approach for axonal guidance in peripheral nerve repair strategies.

P1622Cancer drugs induce functional and structural impairment in adult cardiomyocytes

Abstract Introduction The addition of anti-human epidermal growth factor receptor 2 (HER2; ErbB2) monoclonal antibody Trastuzumab (TRZ) to Doxorubicin (DOXO) chemotherapy is associated with a synergistic increase in cardiac toxicity. While previous studies have addressed the toxicity of both agents on isolated cardiomyocytes (CMs), little is known regarding this process in vivo, especially with respect to electrophysiological changes. Purpose To investigate electrical and structural changes in LV and RV CMs using an in vivo rat model of DOXO/TRZ cardiotoxicity. Methods Rats received 6 IP injections of either DOXO or TRZ over a 2-week period, or 6 doses of DOXO followed by 6 doses of TRZ (COMBO), or saline as a control. In-vivo echocardiography was performed. Electrical activity and Ca2+ handling were assessed in LV and RV CMs from rat hearts. Single cell patch-clamp and field stimulation experiments were performed. Spontaneous sarcoplasmic reticulum Ca2+ release events (Ca2+ sparks) were recorded at x100 magnification in line-scan mode (sampling rate 0.7 kHz) from 2 μM Fluo4-AM loaded CMs. To assess T-tubular disarray, CMs were incubated with di-3-ANEPPDHQ and periodic component was quantified by Fast Fourier Transform (FFT) analysis of confocal microscopy images. Results DOXO, and to a greater extent COMBO treatment was associated with significant increases in both LV end-systolic and end-diastolic volumes, and decreases in LVEF and fractional shortening. By contrast, TRZ alone merely increased LV end-systolic volume. Electrophysiological studies showed increases in action potential duration (APD), beat-to-beat variability of repolarization (BVR), delayed after depolarizations (DADs), and Ca2+-sparks in both DOXO and COMBO groups. Stimulated intracellular Ca2+ transients (1,2 and 4 Hz) showed significant changes with respect to time to peak, tau decay, amplitude, and fractional release in the DOXO group. These changes were associated with a significant downregulation of sarco/endoplasmic reticulum Ca2+ ATPase pump (SERCA) expression. From a structural viewpoint, these changes were associated with T-tubular disarray in the DOXO and COMBO groups. Conclusions DOXO, and to a greater extent COMBO treatment (but not TRZ alone) cause LV dysfunction in vivo. Moreover, both DOXO and COMBO treatments, but not TRZ alone, induce electrophysiological abnormalities and both structural and functional changes in the sarcoplasmic reticulum. These findings provide novel insights into the cellular mechanisms of CM dysfunction and arrhythmias associated with combined DOXO/TRZ therapy. Acknowledgement/Funding Swiss League against Cancer

Mathematical Modeling of Drug Release in a Phase-Transient Temperature-Responsive Drug Delivery System in Spherical Coordinates

A sphere-shaped drug delivery system responsive to temperature, as a unique external stimulus, was introduced and its performance mathematically studied at the pseudo-steady state. The system is composed of three individual sections, including the drug core, phase-transient intermediate shell, and protective polymeric shell. An ON-OFF release of drug could be achieved by increasing or decreasing the environmental temperature around the melting point of the intermediate shell and the smartness of system is due to the solid-liquid phase transition of this shell. The ON-OFF response of the system was mathematically modeled by solving the governing heat and mass transfer equations at the pseudo-steady state. The results showed the lag time of the system in the ON state, the cumulative released drug in the ON state and the fractional undesired release of drug in the OFF state are strongly under the influences of different kinds of factors, including the geometrical characteristics of the system (e.g., the radius of the drug core and the thicknesses of the intermediate and polymeric shells), the physical properties of the system (e.g., the thermal conductivities and diffusion coefficients of the intermediate and polymeric shells), and the environmental and operation conditions.

Leaching of spent nuclear fuels in aerated conditions: Influences of sample preparation on radionuclide release patterns

The release of radionuclides from spent nuclear fuel upon contact with water is a central issue for the assessment of the safety of geological disposal concepts. Several studies have been conducted aiming at understanding matrix dissolution as well as the rapid/instant release of radionuclides separated from the UO2 matrix; there are however questions remaining regarding how higher burn-up affects the amount of fission products segregated to the fuel-cladding gap and grain boundaries.In this study we have performed aerated fuel corrosion and leaching experiments using spent nuclear fuels with a range of burnups, from ∼52 to ∼70 MWd/KgU. The samples have been prepared in different manners in order to investigate the release of the segregated fission products from the gap and the grain boundaries. Thus, the fuel samples are prepared either as segments with the cladding attached, as fragments without cladding, or as milled powder. The fuel samples are leached in aqueous solution in aerated conditions and the radionuclide release is monitored during several contact periods for up to five years total leaching time.The results from the leaching of fuel segments reveal congruent release of e.g. europium and neodymium whereas the release of those elements was lower than the U-238 release for the fragment samples. Potential explanations for this are discussed in the paper.The results show that the release rates of elements segregated from the fuel matrix were in general found to be lower from segments samples as compared to fragment samples, which can be attributed to the closed fuel-cladding gap inhibiting the exposure of the gap inventory to water for the high burnup fuels used in this study.Leaching of fuel powder (78 MWd/kgU local BU) by simultaneous grinding and leaching showed a fractional release of Cs-137 and I-129 of 1.5% and 1.8% respectively.

EVALUACIÓN DEL MECANISMO DE ABSORCIÓN DE ACEITE Y PÉRDIDA DE AGUA DURANTE EL FREIDO DE TORTILLA CHIPS

Simple equation Mt/M∞ = Ktn is evaluated to describe the Fickian mechanism of oil absorption and water release during the frying of tortilla chips in nixtamalized samples with lime and without lime where Mt/M∞ is the fractional solute absorption or release, and n is the diffusional exponent characteristic of the Fickian mechanism that changes with food physical and chemical properties. For the Fickian mechanism, the exponent (n) is ≤0.50, and non-Fickian is 0.5<n<1 for slabs. Moisture releases showed a Fickian mechanism form tortilla chips with lime (n = 0.5) due to water interaction with matrix components and non-Fickian for tortilla chips without lime (n = 0.6). Oil absorption showed a Fickian mechanism for nixtamalization with lime n = 0.4 and for nixtamalization without lime, one of n = 0.3. The larger water diffusion coefficient in the tortilla chip compared to that of oil diffusion indicates that water release and oil absorption during frying is not a simultaneous event. The behavior of the water and oil diffusion is an interesting topic that, in this study, was investigated using a practical equation with potential application in food processing.

The effects of macrophages on cardiomyocyte calcium-handling function using in vitro culture models.

Following myocardial infarction (MI), myocardial inflammation plays a crucial role in the pathogenesis of MI injury and macrophages are among the key cells activated during the initial phases of the host response regulating the healing process. While macrophages have emerged as attractive effectors in tissue injury and repair, the contribution of macrophages on cardiac cell function and survival is not fully understood due to complexity of the in vivo inflammatory microenvironment. Understanding the key cells involved and how they communicate with one another is of paramount importance for the development of effective clinical treatments. Here, novel in vitro myocardial inflammation models were developed to examine how both direct and indirect interactions with polarized macrophage subsets present in the post‐MI microenvironment affect cardiomyocyte function. The indirect model using conditioned medium from polarized macrophage subsets allowed examination of the effects of macrophage‐derived factors on stem cell‐derived cardiomyocyte function for up to 3 days. The results from the indirect model demonstrated that pro‐inflammatory macrophage‐derived factors led to a significant downregulation of cardiac troponin T (cTnT) and sarcoplasmic/endoplasmic reticulum calcium ATPase (Serca2) gene expression. It also demonstrated that inhibition of macrophage‐secreted matricellular protein, osteopontin (OPN), led to a significant decrease in cardiomyocyte store‐operated calcium entry (SOCE). In the direct model, stem cell‐derived cardiomyocytes were co‐cultured with polarized macrophage subsets for up to 3 days. It was demonstrated that anti‐inflammatory macrophages significantly increased cardiomyocyte Ca2+ fractional release while macrophages independent of their subtypes led to significant downregulation of SOCE response in cardiomyocytes. This study describes simplified and controlled in vitro myocardial inflammation models, which allow examination of potential beneficial and deleterious effects of macrophages on cardiomyocytes and vise versa. This can lead to our improved understanding of the inflammatory microenvironment post‐MI, otherwise difficult to directly investigate in vivo or by using currently available in vitro models.

Magnetic Graphene Oxide Nanocarrier for Targeted Delivery of Cisplatin: A Perspective for Glioblastoma Treatment.

Selective vectorization of Cisplatin (CisPt) to Glioblastoma U87 cells was exploited by the fabrication of a hybrid nanocarrier composed of magnetic γ-Fe2O3 nanoparticles and nanographene oxide (NGO). The magnetic component, obtained by annealing magnetite Fe3O4 and characterized by XRD measurements, was combined with NGO sheets prepared via a modified Hummer’s method. The morphological and thermogravimetric analysis proved the effective binding of γ-Fe2O3 nanoparticles onto NGO layers. The magnetization measured under magnetic fields up to 7 Tesla at room temperature revealed superparamagnetic-like behavior with a maximum value of MS = 15 emu/g and coercivity HC ≈ 0 Oe within experimental error. The nanohybrid was found to possess high affinity towards CisPt, and a rather slow fractional release profile of 80% after 250 h. Negligible toxicity was observed for empty nanoparticles, while the retainment of CisPt anticancer activity upon loading into the carrier was observed, together with the possibility to spatially control the drug delivery at a target site.

An Assessment of Initial Leaching Characteristics of Alkali-Borosilicate Glasses for Nuclear Waste Immobilization.

Initial leaching characteristics of simulated nuclear waste immobilized in three alkali- borosilicate glasses (ABS-waste) were studied. The effects of matrix composition on the containment performance and degradation resistance measures were evaluated. Normalized release rates are in conformance with data reported in the literature. High Li and Mg loadings lead to the highest initial de-polymerization of sample ABS-waste (17) and contributed to its thermodynamic instability. Ca stabilizes non-bridging oxygen (NBO) and reduces the thermodynamic instability of the modified matrix. An exponential temporal change in the alteration thickness was noted for samples ABS-waste (17) and Modified Alkali-Borosilicate (MABS)-waste (20), whereas a linear temporal change was noted for sample ABS-waste (25). Leaching processes that contribute to the fractional release of all studied elements within the initial stage of glass corrosion were quantified and the main controlling leach process for each element was identified. As the waste loading increases, the contribution of the dissolution process to the overall fractional release of structural elements decreases by 43.44, 5.05, 38.07, and 52.99% for Si, B, Na, and Li respectively, and the presence of modifiers reduces this contribution for all the studied metalloids. The dissolution process plays an important role in controlling the release of Li and Cs, and this role is reduced by increasing the waste loading.

Iron-deficiency anemia reduces cardiac contraction by downregulating RyR2 channels and suppressing SERCA pump activity.

Iron deficiency is present in ~50% of heart failure (HF) patients. Large multicenter trials have shown that treatment of iron deficiency with i.v. iron benefits HF patients, but the underlying mechanisms are not known. To investigate the actions of iron deficiency on the heart, mice were fed an iron-depleted diet, and some received i.v. ferric carboxymaltose (FCM), an iron supplementation used clinically. Iron-deficient animals became anemic and had reduced ventricular ejection fraction measured by magnetic resonance imaging. Ca2+ signaling, a pathway linked to the contractile deficit in failing hearts, was also significantly affected. Ventricular myocytes isolated from iron-deficient animals produced smaller Ca2+ transients from an elevated diastolic baseline but had unchanged sarcoplasmic reticulum (SR) Ca2+ load, trigger L-type Ca2+ current, or cytoplasmic Ca2+ buffering. Reduced fractional release from the SR was due to downregulated RyR2 channels, detected at protein and message levels. The constancy of diastolic SR Ca2+ load is explained by reduced RyR2 permeability in combination with right-shifted SERCA activity due to dephosphorylation of its regulator phospholamban. Supplementing iron levels with FCM restored normal Ca2+ signaling and ejection fraction. Thus, 2 Ca2+-handling proteins previously implicated in HF become functionally impaired in iron-deficiency anemia, but their activity is rescued by i.v. iron supplementation.

Parametric study of iodine-129 releases from nuclear fuel to fuel-clad gap & primary coolant in PWRs

A numerical methodology is developed to compute volatile fission fragment release from fuel to gap and then to the primary coolant under changing reactor conditions. Based on diffusion of fission fragments and mass balances in gap and in coolant, a FORTRAN-77 based program DIFFP (Diffusion of Fission Products) has been developed to solve numerically the time & space dependent equations using source release rate respectively. For power shutdown effect of spiking have been modeled for given release rate. The predicted gas fractional release as a function of time using DIFFP program was in good agreement with the reported results. Then parametric studies for empirical diffusion constant (D′), coolant purification rate (βP), and gas escape rate coefficient (ν) respectively were done to find the sensitivity of parameters. For operation time >150 days, the sensitivity increases from 4.5% to 20.4% with an increase D′ from 4.1 × 10−10 s−1 to 6.9 × 10−10 s−1 respectively and sensitivity is a nonlinear function of D′: sensitivity=(-62.20±0.78)+(16.83±0.37)D′Do′+(-0.703±0.0398)D′Do′2where, 0.69<D′/D0′<1.17 and D′ is the reference value (5.9 × 10−10 s−1) for the defective fuel rods. Sensitivity for gas escape rate coefficient (ν) was also determined by increasing it from 5% to 50% from its reference value (1.4 × 10−4 s−1) and results show that the coolants activity becomes insensitive to changes in the gas escape rate coefficient and sensitivity value becomes less than 1% when effective full power days are larger than 150. If coolant purification rate constant (βP) is parametrically changed from 5% to 30%, then the absolute sensitivity value increases from 4.8% to 23% respectively and for all cases, the absolute sensitivity remains independent of time (in terms of effective full power days) for a given value of βP and rises linearly with an increase in the value.

Contraction and intracellular calcium transport in epicardial and endocardial ventricular myocytes from streptozotocin-induced diabetic rat

Introduction: Diabetes mellitus (DM) is a global health problem. According to the International Diabetes Federation, 424.9 million people suffered from DM in 2017 and this number is expected to rise to 628.6 million by 2045. Although diabetes can affect every organ in the body, cardiovascular disease is a major cause of death and disability in people with diabetes. Diabetic patients frequently suffer from systolic and diastolic dysfunction. Within the ventricles, the electromechanical properties of cardiac myocytes vary transmurally. Aims and Objectives: The aim of this study was to investigate contraction and Ca2+ transport in epicardial (EPI) and endocardial (ENDO) myocytes from the left ventricle in the streptozotocin (STZ) – induced diabetic rat heart. Materials and Methods: Experiments were performed 5-6 months after STZ treatment. Ventricular myocytes were isolated by enzymic and mechanical dispersal techniques from EPI and ENDO regions of the left ventricle. Contraction and free intracellular Ca2+ concentration [Ca2+]i were measured by video edge detection and fluorescence photometry techniques, respectively. Results: Myocyte length and calculated surface area were smaller in EPI-STZ compared to EPI-CON. Time to peak (TPK) shortening was prolonged in EPI-STZ compared to EPI-CON and in ENDO-STZ compared to ENDO-CON myocytes. Time to half (THALF) relaxation of shortening was prolonged in EPI-STZ compared to EPI-CON. TPK Ca2+ transient was prolonged in EPI-STZ compared to EPI-CON, ENDO-STZ compared to ENDO-CON, ENDO-STZ compared to EPI-STZ and in ENDO-CON compared to EPI-CON myocytes. THALF decay of the Ca2+ transient was prolonged in ENDO-STZ compared to ENDO-CON. Fractional release of Ca2+ was increased in ENDO-STZ compared to ENDO-CON and in ENDO-STZ compared to EPI-STZ. Recovery of the Ca2+ transient was prolonged in ENDO-STZ compared to ENDO-CON. Conclusion: In conclusion the kinetics of contraction and Ca2+ transient and fractional release of Ca2+ from the sarcoplasmic reticulum are altered to different extents in EPI and ENDO myocytes from STZ-induced diabetic rat.

3,4-Methylenedioxymethamphetamine, mephedrone, and β-phenylethylamine release dopamine from the cytoplasm by means of transporters and keep the concentration high and constant by blocking reuptake

The addiction-related behavioural effects of drugs of abuse are mediated by the mesocorticolimbic monoamine systems. We investigated the effects of 3,4-methylenedioxymethamphetamine (MDMA), mephedrone, β-phenylethylamine (β-PEA) methylphenidate (MPH) on dopamine release from mouse perfused nucleus accumbens and prefrontal cortex slices. The fractional release of [3H]-dopamine was measured at rest and in response to field stimulation. The distributions of [3H]-dopamine and its metabolites were determined using high-pressure liquid chromatography. The effect of drugs on [3H]-dopamine uptake was measured in synaptosomal P2 preparations from the frontal cortex and striatum. Similar to MDMA, mephedrone β-PEA increased the resting release of [3H]-dopamine from the nucleus accumbens and prefrontal cortex in a [Ca2+]o-independent manner, and the stimulation-evoked release was also augmented. In contrast, MPH failed to affect the resting release but potentiated the release in response to axonal activity. Similar to dopamine transporter antagonist GBR 12909, mephedrone, MDMA and MPH biphasically inhibited the [3H]-dopamine uptake. The administration of GBR 12909 and nisoxetine, or lowering the bath temperature prevented MDMA, mephedrone and β-PEA from enhancing the resting, cytoplasmic release of [3H]-dopamine, indicating the role of transporters in the release process. We conclude that amphetamine-like drugs of abuse and the trace amine β-PEA excessively increase the [Ca2+]o-independent, non-vesicular release of dopamine from the cytoplasm into the extrasynaptic space and inhibit the high-affinity transporters, thereby maintaining a high ambient, non-synaptic concentration of dopamine that may tonically control the activity of neurons equipped with dopamine receptors and is likely involved in the reinforcing effects and abusive potential of amphetamines.

Phase evolution and chemical stability of the Nd2O3-ZrO2-SiO2 system synthesized by a novel hydrothermal-assisted sol-gel process

A hydrothermal-assisted sol-gel process, a novel method of combining both advantages of sol-gel and hydrothermal processes, was developed to synthesize dense Nd-doped ZrSiO4 ceramics with high Nd (surrogate for trivalent actinides) loading at low sintering temperature (1723 K) and short time (6 h). The evolutions of phase and microstructure of the obtained ceramics with the varying composition, sintering temperature and pH value were investigated. The chemical durability of the ceramics was also evaluated. The results show that the properties of ceramics strongly depend on sintering temperature and pH value. The acidic condition is beneficial to phase formation and densification of ZrSiO4 ceramics. It was found that the maximum immobilization capacity of Nd in the single-phase ZrSiO4 ceramics is highly dependent upon the synthesis method and sintering temperature. With decreasing sintering temperature, the maximum immobilization capacity increases. And the immobilization capacity of Nd in ZrSiO4 ceramics was found to be enhanced by the hydrothermal-assisted sol-gel method. Furthermore, the normalized release rate of Nd in Nd-doped ZrSiO4 ceramics decreases with increasing time and remains almost unchanged (∼10−4 g m-2 d-1) after 28 days, and the fractional release of Nd in all samples is about ∼10−5 wt%, exhibiting high chemical stability.

The Role Of P62 On Chronic Oxidative Stress‐Induced Myocytes Ca handling

IntroductionRecent studies have shown that the activity of autophagy impairs, and the accumulation of p62 protein, autophagy substrate, in the heart observes in heart failure patients and mouse models of cardiomyopathy. Accumulations of p62 protein are known to activate the intrinsic cellular stress. Abnormal Ca release is partly responsible for oxidative stress‐induced contractile dysfunction of myocytes in ischemic heart disease. Myocytes Ca release is controlled by a function of sarcoplasmic reticulum (SR) Ca store. However, the role of p62 protein on myocytes Ca regulation in the ischemic heart is not clear.PurposeTo investigate the role of p62 on SR Ca regulation in myocytes isolated from mice with the Isoproterenol‐induced myocardial injury.MethodsTo test our hypothesis, both wild‐type (WT) and p62 knockout (KO) mice were used at 10–12 weeks of age. Myocytes isolated from WT and p62 KO mice (n=3 per group) injected with 200 mg/kg Isoproterenol (ISO; beta agonist, twice per day) for 0 (Base), 5 days, and 10 days. SR Ca handling and contractility measured in field‐stimulated single myocytes loaded with fluorescent Ca indicators (Fura‐2AM; cytosolic Ca loading). Amplitudes of caffeine (10 mM, 5 sec)‐induced Ca transients used as estimates of SR Ca store. Sets of heart from WT and p62 mice were processed for western blot (n=3 per group).ResultsThe p62‐deficient myocytes had no difference of myocytes contractility, and SR Ca handling at the baseline compared to WT myocytes (n=35–45 cells per group). After acute ISO exposure (1μM for 1 min), WT myocytes increased systolic Ca release whereas p62‐deficient myocytes did not. An exposure of a submaximal oxidative stress (1μM; H2O2) in p62‐deficient myocytes, however, increased myocytes contractility and the fraction of Ca released from the SR during each beat (fractional Ca release; systolic Ca release over SR Ca store) after acute ISO stimulation (p&lt;0.05). In WT mice injected with ISO for 5 days and 10 days, myocytes had a biphasic effect on myocytes contractility and fractional Ca release (n=15–45 cells per group, p&lt;0.05). Interestingly, in p62‐deficient myocytes further increased myocytes contractility and fractional Ca release (Figure) on chronic ISO injections from 5 days to 10 days. WT mice hearts with chronic exposure to ISO for 10 days increased p62 protein expressions and suppressed the conversion of LC3 I to LC3 II.ConclusionTaken together, our results suggest that accumulations of p62 protein may directly cause SR Ca defects via beta‐adrenergic signaling in myocytes under chronic myocardial injury.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Evaluation of stratospheric age of air from CF&amp;lt;sub&amp;gt;4&amp;lt;/sub&amp;gt;, C&amp;lt;sub&amp;gt;2&amp;lt;/sub&amp;gt;F&amp;lt;sub&amp;gt;6&amp;lt;/sub&amp;gt;, C&amp;lt;sub&amp;gt;3&amp;lt;/sub&amp;gt;F&amp;lt;sub&amp;gt;8&amp;lt;/sub&amp;gt;, CHF&amp;lt;sub&amp;gt;3&amp;lt;/sub&amp;gt;, HFC-125, HFC-227ea and SF&amp;lt;sub&amp;gt;6&amp;lt;/sub&amp;gt;; implications for the calculations of halocarbon lifetimes, fractional release factors and ozone depletion potentials

Abstract. In a changing climate, potential stratospheric circulation changes require long-term monitoring. Stratospheric trace gas measurements are often used as a proxy for stratospheric circulation changes via the mean age of air values derived from them. In this study, we investigated five potential age of air tracers – the perfluorocarbons CF4, C2F6 and C3F8 and the hydrofluorocarbons CHF3 (HFC-23) and HFC-125 – and compare them to the traditional tracer SF6 and a (relatively) shorter-lived species, HFC-227ea. A detailed uncertainty analysis was performed on mean ages derived from these new tracers to allow us to confidently compare their efficacy as age tracers to the existing tracer, SF6. Our results showed that uncertainties associated with the mean age derived from these new age tracers are similar to those derived from SF6, suggesting that these alternative compounds are suitable in this respect for use as age tracers. Independent verification of the suitability of these age tracers is provided by a comparison between samples analysed at the University of East Anglia and the Scripps Institution of Oceanography. All five tracers give younger mean ages than SF6, a discrepancy that increases with increasing mean age. Our findings qualitatively support recent work that suggests that the stratospheric lifetime of SF6 is significantly less than the previous estimate of 3200 years. The impact of these younger mean ages on three policy-relevant parameters – stratospheric lifetimes, fractional release factors (FRFs) and ozone depletion potentials – is investigated in combination with a recently improved methodology to calculate FRFs. Updates to previous estimations for these parameters are provided.

Comparison of structure, morphology, and leach characteristics of multi-phase ceramics produced via melt processing and hot isostatic pressing

Melt processing of multi-phase ceramic waste forms offers potential advantages over traditional solid-state synthesis methods given both the prevalence of melters currently in use and the ability to reduce the possibility of airborne radionuclide contamination. In this work, multi-phase ceramics with a targeted hollandite composition of Ba1.0Cs0.3Cr1.0Al0.3Fe1.0Ti5.7O16 were fabricated by melt processing at 1675 °C and hot isostatic pressing (HIP) at 1250 and 1300 °C. X-ray diffraction analysis (XRD) confirmed hollandite as the major phase in all specimens. Zirconolite/pyrochlore peaks and weaker perovskite reflections were observed after melt processing, while HIP samples displayed prominent perovskite peaks and low-intensity zirconolite reflections. Melt processing produced specimens with large (>50 μm) well-defined hollandite grains, while HIP yielded samples with a more fine-grained morphology. Elemental analysis showed “islands” rich in Cs and Ti across the surface of the 1300 °C HIP sample, suggesting partial melting and partitioning of Cs into multiple phases. Photoemission data revealed multiple Cs 3d spin-orbit pairs for the HIP samples, with the lower binding energy doublets likely corresponding to Cs located in more leachable phases. Among all specimens examined, the melt-processed sample exhibited the lowest fractional release rates for Rb and Cs. However, the retention of Sr and Mo was greater in the HIP specimens.

Mechanotransduction via No Signaling Auto-Regulates Cardiomyocyte Contractility

Background: The heart must adjust its contractile force to compensate for the blood pressure changes in daily life (due to changes in posture, physical activity, emotional state, etc.) in order to maintain adequate cardiac output. Here we investigate the mechano-chemo-transduction (MCT) mechanisms that transduce mechanical load to regulate cardiac Ca2+ signaling and contractility. Methods: We used a Cell-in-Gel system to embed freshly isolated rabbit ventricular myocytes in a 3-D viscoelastic hydrogel. The cell in-gel was continuously perfused with Tyrode's solution and electrically paced at 0.5 Hz. Self-control experiments were conducted by (1) studying the myocytes contracting in-gel under mechanical load, (2) then dissolving the gel to release the cell into Tyrode's solution and (3) studying the cell contracting in solution load-free. Results: The cardiomyocytes contracting in-gel exhibited a larger cytosolic Ca2+ transient than load-free cells (Fura-2 ratio 2.71±0.07 vs 2.08±0.04), revealing the MCT effect on regulating intracellular Ca2+. The SR Ca2+ content was higher and the fractional release of Ca2+ from SR was also larger when the cell was in-gel than load-free (measured with Fluo-5N), which should contribute to increasing the cytosolic Ca2+ transient. Furthermore, the MCT effects on the SR Ca2+ and cytosolic Ca2+ were abolished by using 1 mM L-NAME to inhibit nitric oxide synthases (NOS), suggesting a critical role of NOS-NO signaling in mediating MCT. The cell in-gel contraction was reduced from 13.5±0.4% in untreated cell to 10.7±0.6% after L-NAME treatment, showing the MCT effect on enhancing contraction. Conclusion: Mechano-chemo-transduction in cardiomyocyte is mediated by NOS-NO signaling to regulate the SR Ca2+ release to increase the cytosolic Ca2+ transient, which in turn enhances the contractility in compensatory response to increased load. This MCT mechanism contributes to the Anrep effect and is fundamental to cardiac regulation in health and disease.

A refined method for calculating equivalent effective stratospheric chlorine

Abstract. Chlorine and bromine atoms lead to catalytic depletion of ozone in the stratosphere. Therefore the use and production of ozone-depleting substances (ODSs) containing chlorine and bromine is regulated by the Montreal Protocol to protect the ozone layer. Equivalent effective stratospheric chlorine (EESC) has been adopted as an appropriate metric to describe the combined effects of chlorine and bromine released from halocarbons on stratospheric ozone. Here we revisit the concept of calculating EESC. We derive a refined formulation of EESC based on an advanced concept of ODS propagation into the stratosphere and reactive halogen release. A new transit time distribution is introduced in which the age spectrum for an inert tracer is weighted with the release function for inorganic halogen from the source gases. This distribution is termed the release time distribution. We show that a much better agreement with inorganic halogen loading from the chemistry transport model TOMCAT is achieved compared with using the current formulation. The refined formulation shows EESC levels in the year 1980 for the mid-latitude lower stratosphere, which are significantly lower than previously calculated. The year 1980 is commonly used as a benchmark to which EESC must return in order to reach significant progress towards halogen and ozone recovery. Assuming that – under otherwise unchanged conditions – the EESC value must return to the same level in order for ozone to fully recover, we show that it will take more than 10 years longer than estimated in this region of the stratosphere with the current method for calculation of EESC. We also present a range of sensitivity studies to investigate the effect of changes and uncertainties in the fractional release factors and in the assumptions on the shape of the release time distributions. We further discuss the value of EESC as a proxy for future evolution of inorganic halogen loading under changing atmospheric dynamics using simulations from the EMAC model. We show that while the expected changes in stratospheric transport lead to significant differences between EESC and modelled inorganic halogen loading at constant mean age, EESC is a reasonable proxy for modelled inorganic halogen on a constant pressure level.