Kandelia candel Thioredoxin f Confers Osmotic Stress Tolerance in Transgenic Tobacco.

Water deficit caused by osmotic stress and drought limits crop yield and tree growth worldwide. Screening and identifying candidate genes from stress-resistant species are a genetic engineering strategy to increase drought resistance. In this study, an increased concentration of mannitol resulted in elevated expression of thioredoxin f (KcTrxf) in the nonsecretor mangrove species Kandelia candel. By means of amino acid sequence and phylogenetic analysis, the mangrove Trx was classified as an f-type thioredoxin. Subcellular localization showed that KcTrxf localizes to chloroplasts. Enzymatic activity characterization revealed that KcTrxf recombinant protein possesses the disulfide reductase function. KcTrxf overexpression contributes to osmotic and drought tolerance in tobacco in terms of fresh weight, root length, malondialdehyde (MDA) content, and hydrogen peroxide (H2O2) production. KcTrxf was shown to reduce the stomatal aperture by enhancing K+ efflux in guard cells, which increased the water-retaining capacity in leaves under drought conditions. Notably, the abscisic acid (ABA) sensitivity was increased in KcTrxf-transgenic tobacco, which benefits plants exposed to drought by reducing water loss by promoting stomatal closure. KcTrxf-transgenic plants limited drought-induced H2O2 in leaves, which could reduce lipid peroxidation and retain the membrane integrity. Additionally, glutathione (GSH) contributing to reactive oxygen species (ROS) scavenging and transgenic plants are more efficient at regenerating GSH from oxidized glutathione (GSSG) under conditions of drought stress. Notably, KcTrxf-transgenic plants had increased glucose and fructose contents under drought stress conditions, presumably resulting from KcTrxf-promoted starch degradation under water stress. We conclude that KcTrxf contributes to drought tolerance by increasing the water status, by enhancing osmotic adjustment, and by maintaining ROS homeostasis in transgene plants.

Individual and combined effects of fluoranthene, phenanthrene, mannitol and sulfuric acid on marigold (Calendula officinalis).

A study was conducted to characterize marigold stress response to polycyclic aromatic hydrocarbons (PAHs) (oxidative stress inducers) with and without sulfuric acid (S.Acid; pH 3) (acid-stress inducer), and to evaluate reactive oxygen species (ROS) scavenging activity of mannitol (Mann). Marigold (Calendula officinalis) seedlings were grown in a greenhouse and fumigated with fluoranthene (FLU), phenanthrene (PHE), Mann, and S.Acid individually and in various combinations for 40 days. Various physiological and biochemical parameters among others were analyzed using standard methods. The results revealed that fumigation of FLU induced oxidative stress to the plants via ROS generation leading to negative effects on photosynthesis at near saturating irradiance (Amax), stomatal conductance (Gs), internal carbon dioxide concentration (Ci), leaf water relations and chlorophyll pigments. Significant per cent inhibition of Amax (54%), Gs (86%) and Ci (32%), as well as per cent reductions in chlorophyll a (Chl.a) (33%), Chl.b (34%), and total chlorophyll (Tot. Chl) (48%) contents were recorded in FLU fumigated treatment in comparison to control. Combination of Mann with FLU scavenged the generated ROS and substantially lowered the oxidative stress on the plants hence all the measured parameters were not significantly different from control. PHE fumigation had varied effects on marigold plants and was not as deleterious as FLU. Combined fumigation of S.Acid with both the PAHs had significant negative effect on leaf water relations, and positive effect on fresh and turgid weight of the plants but had no effect on the other measured parameters. The lowest proline contents and highest catalase and ascorbate peroxidase activities in FLU fumigated plants further confirmed that oxidative stress was imposed via the generation of ROS. From the results, it is evident that Mann could be an efficient scavenger of ROS-generated by FLU in the marigold plants. We recommend Mann to be widely used for the protection of higher plants from FLU-generated stress in the urban areas.

High glucose-induced hyperosmolarity contributes to COX-2 expression and angiogenesis: implications for diabetic retinopathy.

BACKGROUND: We tested the hypothesis that glucose-induced hyperosmolarity, occurring in diabetic hyperglycemia, promotes retinal angiogenesis, and that interference with osmolarity signaling ameliorates excessive angiogenesis and retinopathy in vitro and in vivo. METHODS AND RESULTS: We incubated human aortic (HAECs) and dermal microvascular endothelial cells (HMVECs) with glucose or mannitol for 24 h and tested them for protein levels and in vitro angiogenesis. We used the Ins2 Akita mice as a model of type 1 diabetes to test the in vivo relevance of in vitro observations. Compared to incubations with normal (5 mmol/L) glucose concentrations, cells exposed to both high glucose and high mannitol (at 30.5 or 50.5 mmol/L) increased expression of the water channel aquaporin-1 (AQP1) and cyclooxygenase (COX)-2. This was preceded by increased activity of the osmolarity-sensitive transcription factor Tonicity enhancer binding protein (TonEBP), and enhanced endothelial migration and tubulization in Matrigel, reverted by treatment with AQP1 and TonEBP siRNA. Retinas of Ins2 Akita mice showed increased levels of AQP1 and COX-2, as well as angiogenesis, all reverted by AQP1 siRNA intravitreal injections. CONCLUSIONS: Glucose-related hyperosmolarity seems to be able to promote angiogenesis and retinopathy through activation of TonEBP and possibly increasing expression of AQP1 and COX-2. Osmolarity signaling may be a target for therapy.

Non-Nutritive Polyol Sweeteners Differ in Insecticidal Activity When Ingested by Adult Drosophila melanogaster (Diptera: Drosophilidae).

Previous work showed the non-nutritive polyol sweetener Erythritol was toxic when ingested by Drosophila melanogaster (Meigen, 1930). This study assessed whether insect toxicity is a general property of polyols. Among tested compounds, toxicity was highest for erythritol. Adult fruit flies (D. melanogaster) fed erythritol had reduced longevity relative to controls. Other polyols did not reduce longevity; the only exception was a weaker but significant reduction of female (but not male) longevity when flies were fed D-mannitol. We conclude at least some non-nutritive polyols are not toxic to adult D. melanogaster when ingested for 17 days. The longer time course (relative to erythritol) and female specificity of D-mannitol mortality suggests different mechanisms for D-mannitol and erythritol toxicity to D. melanogaster.

Mannitol-induced drought stress on calli of Trigonellafoenum-graecum L. Var. RMt-303.

Different explants of fenugreek, T. foenum-graecum L. (Var. RMt-303), were compared for their callus induction and subsequent shoot regeneration capabilities on Murashige and Skoog media supplemented with different phytohormones in varying concentration. The highest percentage of callus induction frequency was observed in 1 ppm benzylaminopurine (BAP). Maximum shoots were induced on media supplemented with 0.5 ppm BAP using leaf and stem tissues as explants. However, root tissues showed only callusing with no subsequent shooting. Cotyledonary node responded better than hypocotyls in terms of shoot induction on media supplemented with thidiazuron (0.1 ppm). The callus was subjected to drought stress as simulated by reduced water potential of growth media due to addition of mannitol. Calli could withstand -2 MPa water potential till 30 days indicating that the drought stress tolerance mechanisms are functional in this variety. Chlorophyll a and b and total chlorophyll, proline and total phenolic contents, total peroxidase and catalase activities increased under stress conditions suggesting the tolerance of callus to drought stress. However, ascorbate peroxidase, guaiacol peroxidase activities were found to decrease slightly. Malondialdehyde and H2O2 contents were found to decrease while only a slight disturbance was found in membrane stability index. These results underline the mechanisms that are crucial for drought stress tolerance in fenugreek.

Determination of the genotoxic effects of sweeteners, mannitol and lactitol.

The changes in dietary habit around the world have led to an increased use of additives in the food. The safety of food additives has been a main focus of research for many years due to the ongoing debate on their potential effects on health. In this study, the in vitro genotoxic effects of mannitol and lactitol, polyols used as sweetener food additives, were evaluated using chromosomal aberrations (CAs) and micronucleus (MN) assays in human peripheral lymphocytes. Additionally, the effects of these sweeteners on the mitotic index (MI) and nuclear division index (NDI) were investigated. Concentrations of 500, 1000, 2000, 4000, and 8000 µg/mL for mannitol and 250, 500, 1000, 2000, and 4000 µg/mL for lactitol were used. The results indicated that both polyols did not affect CA and MN frequency, and did not cause a significant change in NDI at all treatment concentratoins. However, mannitol (except at concentrations of 500 and 1000 µg/mL) and lactitol (except at 250 µg/mL) significantly decreased the MI compared to the control at almost all concentrations and treatment times. In conclusion, it was observed that mannitol and lactitol did not have a significant genotoxic effect at the concentrations used in human lymphocytes in vitro.

Cryopreservation of scalps of Malaysian bananas using a pre-growth method.

The effect of preculture with different sugars and mannitol on cryopreservation of scalps of the banana (Musa) cvs. Pisang Mas, Pisang Nangka, Pisang Berangan and Pisang Awak was investigated. Scalps (0.3 square cm) were precultured on semi-solid MS-based medium, containing 0.4 or 0.5 M sucrose, glucose, fructose, trehalose or mannitol, for 14 days under a 16 h light and 8 h dark photoperiod prior to rapid cooling and storage in liquid nitrogen. Explants were rewarmed rapidly in a water bath at 40 degree C for 1 min, followed by recovery on two layers of sterile filter paper overlaying 25 ml aliquots of semi-solid MS-based medium with 5 mg per liter benzylaminopurine, 0.2 mg per liter indole acetic acid and 10 mg per liter ascorbic acid (PM8 medium) for 2 days in the dark. Subsequently, scalps were transferred onto 25 ml aliquots of semi-solid PM8 medium and incubated in the dark for 1 week prior to incubation in the light. Shoot regeneration from 5 - 48 percent of cryopreserved scalps of all the banana cvs., was observed only following preculture with 0.4 or 0.5 M glucose or fructose, and with 0.4 M trehalose for the cvs. Pisang Berangan and Pisang Awak. Preculture with 0.4 M glucose resulted in maximum shoot regeneration of cryopreserved scalps of 10 percent, 13 percent, 42 percent and 48 percent for the cvs. Pisang Mas, Pisang Nangka, Pisang Berangan and Pisang Awak, respectively. Concentrations of 0.5 M trehalose, or 0.4 and 0.5 M sucrose or mannitol were extremely toxic to scalps of all the cvs. investigated.

Enrichment of the erythrocyte miR-451a in brain extracellular vesicles following impairment of the blood-brain barrier.

Extracellular RNAs (exRNAs) are present in all biofluids and incorporate many types of RNAs including miRNA. To enhance their stability outside of the cell, exRNAs are bound within ribonucleoprotein complexes or packaged into extracellular vesicles (EVs). The blood-brain barrier (BBB) is a dynamic interface between the systemic circulation and the CNS and is responsible for maintaining a stable extracellular environment for CNS cells. The intent of this study was to determine if EVs and their contents are transferred from the peripheral circulation to the CNS under conditions of an impaired BBB. The BBB of mice was disrupted by unilateral intracarotid artery infusion with hyperosmolar mannitol solution. To validate barrier opening, the uptake clearance of [13C12]-sucrose in the left forebrain (i.e. the ipsilateral, mannitol injected hemisphere) was quantified and revealed a 14-fold increase in the mannitol perfused hemisphere compared to sham treated mice. EVs were isolated from the extracellular spaces of the left forebrain following gentle tissue lysis and differential ultracentrifugation. EVs were confirmed using nanotracking analysis, electron microscopy and western blotting. qRT-PCR showed that the erythrocyte-enriched miR-451a in brain tissue EVs increased with mannitol treatment by 24-fold. Small RNA sequencing performed on the EVs isolated from the sham and mannitol treated mice showed that miR-9-5p was the most abundant miRNA contained within the brain EVs. qRT-PCR analysis of plasma EVs did not produce a statistically significant difference in the expression of the CNS-enriched miR-9-5p or miR-9-3p, suggesting that transfer of CNS EVs to the peripheral circulation did not occur under the conditions of our experiment. We demonstrate that EVs containing miR-451a, a highly abundant miRNA present within erythrocytes and erythrocyte EVs, are enhanced in the CNS upon BBB disruption.

A Systematic Safety Evaluation of Nanoporous Mannitol Material as a Dry-Powder Inhalation Carrier System.

For carrier-based dry-powder inhaler (DPI) formulations, the adhesion between carrier particles and active pharmaceutical ingredients (API) particles have a significant influence on the aerosolization performance of the API-carrier complexes and the desired detachment of the API for efficient pulmonary delivery. In our previous study, nanoporous mannitol material was successfully fabricated as carriers by a one-step nonorganic solvent spray drying method with the thermal degradation of ammonium carbonate. These carriers were shown to achieve excellent aerosolization performance. In addition, no residue of ammonium carbonate was detected on the powder surface. However, the safety of nanoporous mannitol carriers (Nano-PMCs) during pulmonary administration/delivery was still unknown because the lung is vulnerable to the inhaled particles. To address this question, the present study was conducted to construct a systematic safety evaluation for DPIs carriers to investigate the safety of Nano-PMCs in the whole inhalation, which would make up for the lack of detailed and standardized method in this field. In vitro safety evaluation was carried out using respiratory and pulmonary cytotoxicity tests, hemolysis assay, and ciliotoxicity test. In vivo safety evaluation was studied by measuring inflammatory indicators in the bronchoalveolar lavage fluid, assessing the pulmonary function and observing pulmonary pathological changes. Nano-PMCs showed satisfactory biocompatibility on respiratory tracts and lungs in vitro and in vivo. It was suggested that Nano-PMCs were safe for intrapulmonary delivery and potential as DPI carriers.

An in vitro self-organized three-dimensional model of the blood-brain barrier microvasculature.

The blood-brain barrier (BBB) protects the human brain from external aggression. Despite its great importance, very few in vitro models of the BBB reproducing its complex organization are available yet. Here we fabricated such a three-dimensional (3D) self-organized in vitro model of BBB microvasculature by means of a combination of collagen microfibers (CMF) and fibrin gel. The interconnected fibers supported human brain microvascular endothelial cell migration and the formation of a capillary-like network with a lumen diameter close to in vivo values. Fibrin, a protein involved in blood vessel repair, favored the further 3D conformation of the brain microvascular endothelial cells, astrocytes and pericytes, ensured gel cohesion and avoided shrinkage. The maturation of the BBB microvasculature network was stimulated by both the CMF and the fibrin in the hydrogel. The expression of essential tight-junction proteins, carriers and transporters was validated in regards to bidimensional simple coculture. The volume of gel drops was easily tunable to fit in 96-well plates. The cytotoxicity of D-Mannitol and its impacts on the microvascular network were evaluated, as an example of the pertinence of this 3D BBB capillary model for screening applications.

Genome-wide identification of Arabidopsis long noncoding RNAs in response to the blue light.

Long non-coding RNAs (lncRNAs) have been shown in animals to play roles in a wide range of biological processes. In plant, light modulates the growth and development as a key external signal. However, little is known about the role of plant lncRNA in response to light. In this study, we sequenced the messenger RNAs (mRNAs), lncRNAs and microRNAs (miRNAs) in Arabidopsis seedlings under blue light for 2 h and 8 h. Compared to dark, we identified 4197 mRNAs, 375 miRNAs and 481 lncRNAs, or 5207 mRNAs, 286 miRNAs and 545 lncRNAs of differential expressions under blue light treatments for 2 h or 8 h respectively. Subsequently, a total of 407 competing endogenous RNA (ceRNA) pairs (lncRNA-mRNA-miRNA) were constructed. We identified a blue light-induced lncRNA which plays roles in blue light-directed plant photomorphogenesis and response to mannitol stress by serving as a ceRNA to sequester miR167 in a type of target mimicry. These results revealed previously unknown roles of the lncRNA in blue light signaling and mannitol stress, and provided useful resources of lncRNAs associated with miRNAs in response to blue light.

The effect of hyperosmosis on paracellular permeability in Caco-2 cell monolayers.

The intestinal epithelium is a significant barrier to oral absorption of hydrophilic compounds, and their passage through the intercellular space is restricted by the tight junctions. In this study we found that hyperosmosis is a significant factor altering paracellular transport in Caco-2 cell monolayers. Osmotic regulators, such as sodium chloride, mannitol, and raffinose, decreased transepithelial electrical resistance and enhanced lucifer yellow permeability. The effect of these osmotic regulators on Caco-2 cell monolayers was not likely to be caused by gross cytotoxicity. Although certain amino acids and oligosaccharides have been reported to have specific tight junction-modulating activity, we found that the increased paracellular permeability of Caco-2 monolayers induced by these compounds was at least partly due to the increased osmotic pressure of the test solutions. These findings provide a new potential precaution in the evaluation of paracellular permeability-modulating substances using the Caco-2 cell monolayer system.

Mannitol ingestion causes concentration-dependent, sex-biased mortality in adults of the fruit fly (Drosophila melanogaster).

Mannitol, a sugar alcohol used in commercial food products, has been previously shown to induce sex-biased mortality in female Drosophila melanogaster when ingested at a single concentration (1 M). We hypothesized that sex differences in energy needs, related to reproductive costs, contributed to the increased mortality we observed in females compared to males. To test this, we compared the longevity of actively mating and non-mating flies fed increasing concentrations of mannitol. We also asked whether mannitol-induced mortality was concentration-dependent for both males and females, and if mannitol's sex-biased effects were consistent across concentrations. Females and males both showed concentration-dependent increases in mortality, but female mortality was consistently higher at concentrations of 0.75 M and above. Additionally, fly longevity decreased further for both sexes when housed in mixed sex vials as compared to single sex vials. This suggests that the increased energetic demands of mating and reproduction for both sexes increased the ingestion of mannitol. Finally, larvae raised on mannitol produced expected adult sex ratios, suggesting that sex-biased mortality due to the ingestion of mannitol occurs only in adults. We conclude that sex and reproductive status differences in mannitol ingestion drive sex-biased differences in adult fly mortality.

Acute kidney injury, seizures, and hypertonic hyponatremia secondary to mannitol intoxication in a dog.

OBJECTIVE: To describe a case of mannitol overdose associated with acute kidney injury (AKI), hypertonic hyponatremia, and neurologic abnormalities in a dog. CASE SUMMARY: A 10-year-old intact male Shiba Inu dog was referred to the emergency service of a veterinary teaching hospital for inappetence and acute onset of seizures. The dog had received 2 IV boluses of 3 g/kg of mannitol in less than 24 hours for a glaucoma crisis. Twelve hours after the second injection, the dog became inappetant and developed 2 generalized seizures. Seizure activity was treated with diazepam (0.5 mg/kg IV). Serum biochemistry profile showed severe hyponatremia and hypochloremia, mild hypokalemia, marked increased creatinine (381 µmol/L [44-133 µmol/L]) and moderately increased BUN (13.8 mmol/L [1.6-10.9 mmol/L]). Urinalysis revealed a urine specific gravity of 1.018, glucosuria, proteinuria, pigmenturia and the presence of vacuolized tubular epithelial cells. A presumptive diagnosis of mannitol intoxication was made based on the high dose of mannitol, severe hyponatremia, neurological abnormalities suggestive of intracranial disease, AKI, and urine cytology. Initial calculated plasma osmolality was 263.4 mOsm/kg and measured plasma osmolality was 332 mOsm/kg with an osmolal gap of 68.6 mOsm/kg, confirming the presence of an unmeasured solute attributed to mannitol. Treatment consisted of fluid therapy and supportive care. On day 3, osmolal gap had resolved and serum creatinine concentration returned to normal within 12 days. NEW OR UNIQUE INFORMATION PROVIDED: Mannitol intoxication has been reported in human medicine. This case report is, to our knowledge, the first to describe AKI, hypertonic hyponatremia, and neurological abnormalities secondary to mannitol overdose in a dog.

Cyclosporine A as a Cardioprotective Agent During Donor Heart Retrieval, Storage, or Transportation: Benefits and Limitations.

BACKGROUND: Storage of donor hearts in cardioplegic solutions supplemented with conditioning agents activating endogenous mitochondrial protective signaling enhanced their postreperfusion recovery. The present study investigates the role of timing and duration of cardiac exposure to cyclosporine A (CsA), another putative mitochondrial protectant, on cardiac functional recovery and potential mechanisms of CsA action in an isolated working rat heart model of donor heart retrieval and storage. METHODS: After measurement of baseline function, hearts were arrested and stored for 6 hours at 4°C in either Celsior alone or Celsior + CsA (0.2 µM), then reperfused for 45 minutes in Krebs solution, when functional recovery was assessed. Two additional groups of Celsior-alone stored hearts were exposed to 0.2 µM CsA for the initial 15 minutes (nonworking period) or the full 45-minute period of reperfusion. Coronary effluent was collected pre- and poststorage for assessment of lactate dehydrogenase release. Tissue samples were collected at the end of each study for immunoblotting and histological studies. RESULTS: CsA supplementation during cold storage or the first 15-minute reperfusion significantly improved functional recovery and significantly increased phospho-AMPKαThr172 and phospho-ULK-1Ser757. Hearts exposed to CsA for 45 minutes at reperfusion recovered poorly with no phospho-AMP-activated protein kinase α activation, decreased phospho-eNOSSer633, and decreased mitochondrial cytochrome c content with increased lactate dehydrogenase release. CONCLUSIONS: Inclusion of CsA during cold storage is cardioprotective. Effects of CsA addition to the perfusate during reperfusion were time dependent, with benefits at 15 minutes but not 45 minutes of reperfusion. The toxic effect with the presence of CsA for the full 45-minute reperfusion is associated with impaired mitochondrial integrity and decreased eNOS phosphorylation.

Examining Mannitol Use in Kidney Cancer Surgery: A Cautionary Tale of Extrapolated Surgical Data.

Mannitol for renal protection during partial nephrectomy is common but unsupported by evidence from clinical trials. The impact of mannitol on renal physiology includes both beneficial and harmful effects. Current understanding of renal ischemia reperfusion injury suggests potentially targetable processes that have a lower potential risk.

HIF1A and NFAT5 coordinate Na+-boosted antibacterial defense via enhanced autophagy and autolysosomal targeting.

Infection and inflammation are able to induce diet-independent Na+-accumulation without commensurate water retention in afflicted tissues, which favors the pro-inflammatory activation of mouse macrophages and augments their antibacterial and antiparasitic activity. While Na+-boosted host defense against the protozoan parasite Leishmania major is mediated by increased expression of the leishmanicidal NOS2 (nitric oxide synthase 2, inducible), the molecular mechanisms underpinning this enhanced antibacterial defense of mouse macrophages with high Na+ (HS) exposure are unknown. Here, we provide evidence that HS-increased antibacterial activity against E. coli was neither dependent on NOS2 nor on the phagocyte oxidase. In contrast, HS-augmented antibacterial defense hinged on HIF1A (hypoxia inducible factor 1, alpha subunit)-dependent increased autophagy, and NFAT5 (nuclear factor of activated T cells 5)-dependent targeting of intracellular E. coli to acidic autolysosomal compartments. Overall, these findings suggest that the autolysosomal compartment is a novel target of Na+-modulated cell autonomous innate immunity. Abbreviations: ACT: actins; AKT: AKT serine/threonine kinase 1; ATG2A: autophagy related 2A; ATG4C: autophagy related 4C, cysteine peptidase; ATG7: autophagy related 7; ATG12: autophagy related 12; BECN1: beclin 1; BMDM: bone marrow-derived macrophages; BNIP3: BCL2/adenovirus E1B interacting protein 3; CFU: colony forming units; CM-H2DCFDA: 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate, acetyl ester; CTSB: cathepsin B; CYBB: cytochrome b-245 beta chain; DAPI: 4,6-diamidino-2-phenylindole; DMOG: dimethyloxallyl glycine; DPI: diphenyleneiodonium chloride; E. coli: Escherichia coli; FDR: false discovery rate; GFP: green fluorescent protein; GSEA: gene set enrichment analysis; GO: gene ontology; HIF1A: hypoxia inducible factor 1, alpha subunit; HUGO: human genome organization; HS: high salt (+ 40 mM of NaCl to standard cell culture conditions); HSP90: heat shock 90 kDa proteins; LDH: lactate dehydrogenase; LPS: lipopolysaccharide; Lyz2/LysM: lysozyme 2; NFAT5/TonEBP: nuclear factor of activated T cells 5; MΦ: macrophages; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFI: mean fluorescence intensity; MIC: minimum inhibitory concentration; MOI: multiplicity of infection; MTOR: mechanistic target of rapamycin kinase; NaCl: sodium chloride; NES: normalized enrichment score; n.s.: not significant; NO: nitric oxide; NOS2/iNOS: nitric oxide synthase 2, inducible; NS: normal salt; PCR: polymerase chain reaction; PGK1: phosphoglycerate kinase 1; PHOX: phagocyte oxidase; RFP: red fluorescent protein; RNA: ribonucleic acid; ROS: reactive oxygen species; sCFP3A: super cyan fluorescent protein 3A; SBFI: sodium-binding benzofuran isophthalate; SLC2A1/GLUT1: solute carrier family 2 (facilitated glucose transporter), member 1; SQSTM1/p62: sequestosome 1; ULK1: unc-51 like kinase 1; v-ATPase: vacuolar-type H+-ATPase; WT: wild type.

Inherent toxicity of organ preservation solutions to cultured hepatocytes.

Organ preservation solutions have been designed to protect grafts against the injury inflicted by cold ischemia. However, toxicity of University of Wisconsin (UW) solution during rewarming has been reported. Therefore, we here assessed the toxicity of UW, histidine-tryptophan-ketoglutarate (HTK), Euro-Collins, histidine-lactobionate (HL), sodium-lactobionate-sucrose and Celsior solutions in cultured hepatocytes under hypothermic (4 degrees C), intermediate (21 degrees C) and physiological (37 degrees C) conditions. Marked toxicity of UW, HTK, HL and Euro-Collins solutions was observed at both 37 and 21 degrees C. With the exception of UW solution, these solutions also increased cell injury during cold incubation (LDH release after 18 h at 4 degrees C: HTK 76+/-2%, Euro-Collins 78+/-17%, HL 81+/-15%; control: Krebs-Henseleit buffer 20+/-6%). Testing of individual components using modified Krebs-Henseleit buffers suggested that histidine and phosphate are responsible for (part of) this toxicity. These potential toxicities should be taken into account in the development of future preservation solutions.

Improved corrosion resistance of commercially pure magnesium after its modification by plasma electrolytic oxidation with organic additives.

The optimal mechanical properties render magnesium widely used in industrial and biomedical applications. However, magnesium is highly reactive and unstable in aqueous solutions, which can be modulated to increase stability of reactive metals that include the use of alloys or by altering the surface with coatings. Plasma electrolytic oxidation is an efficient and tuneable method to apply a surface coating. By varying the plasma electrolytic oxidation parameters voltage, current density, time and (additives in the) electrolytic solution, the morphology, composition and surface energy of surface coatings are set. In the present study, we evaluated the influence on surface coatings of two solute additives, i.e. hexamethylenetetramine and mannitol, to base solutes silicate and potassium hydroxide. Results from in vitro studies in NaCl demonstrated an improvement in the corrosion resistance. In addition, coatings were obtained by a two-step anodization procedure, firstly anodizing in an electrolyte solution containing sodium fluoride and secondly in an electrolyte solution with hexamethylenetetramine and mannitol, respectively. Results showed that the first layer acts as a protective layer which improves the corrosion resistance in comparison with the samples with a single anodizing step. In conclusion, these coatings are promising candidates to be used in biomedical applications in particular because the components are non-toxic for the body and the rate of degradation of the surface coating is lower than that of pure magnesium.

Signs in vector-electrocardiography (VECG) predicting the fibrillatory propensity of iodixanol and mannitol solutions after injection into the left coronary artery of pigs.

RATIONALE AND OBJECTIVES: To find signs in vector-electrocardiography (VECG) predicting the ventricular fibrillatory propensity (VF-PROP) of iodixanol and mannitol solutions after injection into the left coronary artery (LCA) of pigs. MATERIALS AND METHODS: Five plasma-isotonic solutions perfused LCA: Iod 320 + Na/Ca (iodixanol 320 mg I/mL, 19 mM NaCl, 0.3 mM CaCl(2)), Iod 320 + Mann (iodixanol 320 mg I/mL, 50 mM mannitol), Mann + Na/Ca (240 mM mannitol, 19 mM NaCl, 0.3 mM CaCl(2)), Mann (275 mM mannitol), and Ringer (representing "physiologic electrolytes"). The first two solutions have at 37 degrees C viscosity 13 mPas and the others <1 mPas. In eight pigs, 20 mL of each solution was injected twice for 10 seconds, and in 15 pigs, each solution was injected for 11-40 seconds (0.5 mL/second) through a wedged catheter in the LCA. If ventricular fibrillation (VF) occurred, injection was stopped and heart was defibrillated. If VF did not occur, perfusion period was 40 seconds. A higher frequency of VF and a shorter period from start of injection until start of VF gave a solution a higher ranking of VF-PROP. RESULTS: The 10-second injections caused no VF. Ringer and Iod 320 + Na/Ca caused no VF after 40-second injections, whereas the other solutions caused VF. Ranking the solutions from lowest to highest VF- PROP gave: Ringer = Iod 320 + Na/Ca < Iod 320 + Mann < Mann + Na/Ca < Mann. Prolongation of QRS time and QTc time were the only VECG signs that showed significant differences (P < .05) between all solutions and correctly ranked the VF-PROP of all solutions in both animal groups. CONCLUSION: The results fit with the concept that a more physiologic electrolyte composition and a higher viscosity of a test solution will, after start of injection of that solution into LCA, delay changes in the electrolyte composition in myocardial interstitial fluid and also delay start of VF. If a plasma isotonic contrast medium (CM) with lower viscosity than that of iodixanol at 320 mgI/mL were created, we conclude that such a CM should have electrolyte composition closer to that of Ringer than present composition (19 mM NaCl and 0.3 mM CaC1(2)) to counteract the effects of faster diffusion of nonphysiologic electrolyte composition from the low-viscosity CM to myocardial interstitial fluid.