The mechanistic insights of essential oil of Mentha piperita to control Botrytis cinerea and the prospection of lipid nanoparticles to its application.

Botrytis cinerea is the phytopathogenic fungus responsible for the gray mold disease that affects crops worldwide. Essential oils (EOs) have emerged as a sustainable tool to reduce the adverse impact of synthetic fungicides. Nevertheless, the scarce information about the physiological mechanism action and the limitations to applying EOs has restricted its use. This study focused on elucidating the physiological action mechanisms and prospection of lipid nanoparticles to apply EO of Mentha piperita. The results showed that the EO of M. piperita at 500, 700, and 900 µL L-1 inhibited the mycelial growth at 100 %. The inhibition of spore germination of B. cinerea reached 31.43 % at 900 µL L-1. The EO of M. piperita decreased the dry weight and increased pH, electrical conductivity, and cellular material absorbing OD260 nm of cultures of B. cinerea. The fluorescence technique revealed that EO reduced hyphae width, mitochondrial activity, and viability, and increased ROS production. The formulation of EO of M. piperita loaded- solid lipid nanoparticles (SLN) at 500, 700, and 900 µL L-1 had particle size ∼ 200 nm, polydispersity index < 0.2, and stability. Also, the thermogravimetric analysis indicated that the EO of M. piperita-loaded SLN has great thermal stability at 50 °C. EO of M. piperita-loaded SLN reduced the mycelial growth of B. cinerea by 70 %, while SLN formulation (without EO) reached 42 % inhibition. These results supported that EO of M. piperita-loaded SLN is a sustainable tool for reducing the disease produced by B. cinerea.

The efficient activity of plant essential oils for inhibiting Botrytis cinerea and Penicillium expansum: Mechanistic insights into antifungal activity.

Botrytis cinerea and Penicillium expansum produce deterioration in fruit quality, causing losses to the food industry. Thus, plant essential oils (EOs) have been proposed as a sustainable alternative for minimizing the application of synthetic fungicides due to their broad-spectrum antifungal properties. This study investigated the efficacy of five EOs in suppressing the growth of B. cinerea and P. expansum and their potential antifungal mechanisms. EOs of Mentha × piperita L., Origanum vulgare L., Thymus vulgaris L., Eucalyptus globules Labill., and Lavandula angustifolia Mill., were screened for both fungi. The results showed that the EO of T. vulgaris and O. vulgare were the most efficient in inhibiting the growth of B. cinerea and P. expansum. The concentration increase of all EO tested increased fungi growth inhibition. Exposure of fungi to EOs of T. vulgaris and O. vulgare increased the pH and the release of constituents absorbing 260 nm and soluble proteins, reflecting membrane permeability alterations. Fluorescence microscopic examination revealed that tested EOs produce structural alteration in cell wall component deposition, decreasing the hypha width. Moreover, propidium iodide and Calcein-AM stains evidenced the loss of membrane integrity and reduced cell viability of fungi treated with EOs. Fungi treated with EOs decreased the mitochondria activity and the respiratory process. Therefore, these EOs are effective antifungal agents against B. cinerea and P. expansum, which is attributed to changes in the cell wall structure, the breakdown of the cell membrane, and the alteration of the mitochondrial activity.

Association of Emotion Regulation and Dispositional Mindfulness in an Adolescent Sample: The Mediational Role of Time Perspective.

This study relates emotional regulation strategies with dispositional mindfulness and the mediating role of time perspective. It is based on the fact that one of the mechanisms of mindfulness consists in providing protective emotional regulation strategies. At the same time, a direct relationship between dispositional mindfulness and time perspective has been observed. To do this, a representative sample of 320 Chilean adolescents from the city of Talcahuano, whose age ranged between 14 and 17 years old, and who were attending high school, was evaluated. The Zimbardo Time Perspective Inventory, the Difficulties in Emotion Regulation Scale, and the Five Facet Mindfulness Questionnaire were applied. Regression analysis results verified the close relationship between emotional regulation and dispositional mindfulness (R2 = 0.54), as well as with the factors of time perspective (R2 = 0.41), explaining, between both of them, 60% of the variance of difficulties in emotional regulation. The possible mediational role of time perspective between dispositional mindfulness and emotional regulation is established.

Biological Crusts to Increase Soil Carbon Sequestration: New Challenges in a New Environment.

The major priority of research in the present day is to conserve the environment by reducing GHG emissions. A proposed solution by an expert panel from 195 countries meeting at COP 21 was to increase global SOC stocks by 0.4% year-1 to compensate for GHG emissions, the '4 per 1000' agreement. In this context, the application of biocrusts is a promising framework with which to increase SOC and other soil functions in the soil-plant continuum. Despite the importance of biocrusts, their application to agriculture is limited due to: (1) competition with native microbiota, (2) difficulties in applying them on a large scale, (3) a lack of studies based on carbon (C) balance and suitable for model parameterization, and (4) a lack of studies evaluating the contribution of biocrust weathering to increase C sequestration. Considering these four challenges, we propose three perspectives for biocrust application: (1) natural microbiome engineering by a host plant, using biocrusts; (2) quantifying the contribution of biocrusts to C sequestration in soils; and (3) enhanced biocrust weathering to improve C sequestration. Thus, we focus this opinion article on new challenges by using the specialized microbiome of biocrusts to be applied in a new environment to counteract the negative effects of climate change.

Manganese-Oxidizing Antarctic Bacteria (Mn-Oxb) Release Reactive Oxygen Species (ROS) as Secondary Mn(II) Oxidation Mechanisms to Avoid Toxicity.

Manganese (Mn) oxidation is performed through oxidative Mn-oxidizing bacteria (MnOxb) as the main bio-weathering mechanism for Mn(III/IV) deposits during soil formation. However, with an increase in temperature, the respiration rate also increases, producing Reactive Oxygen Species (ROS) as by-products, which are harmful to microbial cells. We hypothesize that bacterial ROS oxidize Mn(II) to Mn(III/IV) as a secondary non-enzymatic temperature-dependent mechanism for cell protection. Fourteen MnOxb were isolated from Antarctic soils under the global warming effect, and peroxidase (PO) activity, ROS, and Mn(III/IV) production were evaluated for 120 h of incubation at 4 °C, 15 °C, and 30 °C. ROS contributions to Mn oxidation were evaluated in Arthrobacter oxydans under antioxidant (Trolox) and ROS-stimulated (menadione) conditions. The Mn(III/IV) concentration increased with temperature and positively correlated with ROS production. ROS scavenging with Trolox depleted the Mn oxidation, and ROS-stimulant increased the Mn precipitation in A. oxydans. Increasing the Mn(II) concentration caused a reduction in the membrane potential and bacterial viability, which resulted in Mn precipitation on the bacteria surface. In conclusion, bacterial ROS production serves as a complementary non-enzymatic temperature-dependent mechanism for Mn(II) oxidation as a response in warming environments.

Soil Redox Controls CO2, CH4 and N2O Efflux from White-Rot Fungi in Temperate Forest Ecosystems.

Microaerophilic white-rot fungi (WRF) are impacted by oxygen depletion because of fluctuating redox occurrence in southern temperate forest soils of Chile (1500-5000 mm year-1). How these conditions influence WRF survival has been scarcely examined. We explored the contributions of WRF to greenhouse gas (GHG) emissions of N2O and CH4 and soil organic C oxidation (CO2) in five sterilized and inoculated forest soils derived from various parent materials and climates. The soil was incubated for 20 days following (i) oxic, (ii) anoxic, and (iii) fluctuating redox conditions. Fungi contributed to 45% of the total GHG under redox fluctuating conditions, including the contribution of bacteria, while the opposite (26%) was valid for oxic treatment. On average, the highest gas emission (62%) was N2O for WRF under redox treatment, followed by anoxic (22%) and oxic (16%) treatments, while CO2 and CH4 emissions followed oxic > redox > anoxic. These data suggest that indigenous microbial WRF communities are well adapted to fluctuating redox milieu with a significant release of GHG emissions in humid temperate forests of the southern cone.

Overtime expression of plasma membrane and mitochondrial function markers associated with cell death in human spermatozoa exposed to nonphysiological levels of reactive oxygen species.

In many cell types, the potential of reactive oxygen species to induce death processes has been largely demonstrated. Studies in spermatozoa have associated the imbalance of reactive oxygen species and phosphatidylserine externalisation as an apoptosis marker. However, the lack of consensus about time effect in the joint expression of these and other death markers has made it difficult to understand the set of mechanisms influenced beyond the concentration effect of reactive oxygen species to stimulate cell death. Here, the plasma membrane permeability and integrity, phosphatidylserine externalisation and mitochondrial membrane potential were jointly evaluated as death markers in human spermatozoa stimulated with H2 O2 . The results showed a profound and sustained effect of dissipation in the mitochondrial membrane potential and an increased phosphatidylserine externalisation in human spermatozoa exposed to 3 mmol-1 of H2 O2 at 30 min. This was followed by an increased membrane permeability after 45 min. The last observed event was the loss of cell membrane integrity at 60 min. In conclusion, mitochondria are rapidly affected in human spermatozoa exposed to reactive oxygen species, with the barely detectable mitochondrial membrane potential coexisting with the high phosphatidylserine externalisation in cells with normal membrane permeability.

Iron-reducing bacteria decompose lignin by electron transfer from soil organic matter.

Iron-reducing bacteria (IRB) are crucial for electron transfer in anaerobic soil microsites. The utilization of the energy gathered by this mechanism by decomposers of organic matter is a challenging and fascinating issue. We hypothesized that bacteria reducing Fe(III) (oxyhydr)oxides to soluble Fe(II) obtain electrons from reduced soil organic matter (SOMr) involving lignin oxidation. Iron-reducing bacteria were isolated from topsoils of various climates (humid temperate, cold temperate, subpolar), vegetation types (mostly grasslands and forests), and derived from various parent materials treatments assigned as Granitic, Volcanic-allophanic, Fluvio-glacial, Basaltic-Antarctic and Metamorphic. After the screening of IRB by phospholipid fatty acid (PLFA) analysis and PCR identification (full-length 16S rDNA), the IRB were inoculated to 20 samples (five soils and 4 replicates) and a broad range of parallel processes were traced. Geobacter metallireducens and Geobacter lovleyi were the main Geobacteraceae-strains present in all soils and strongly increased the activity of ligninolytic enzymes: lignin peroxidase and manganese peroxidase. Carbon dioxide (CO2) released from IRB-inoculated soils was 140% higher than that produced by Fenton reactions (induced by H2O2 and Fe(II) addition) but 40% lower than in non-sterile soils. CO2 release was closely correlated with the produced Fe (II) and H2O2 consumption. The highest CO2 was released from Basaltic-Antarctic soils with the highest Fe content and was closely correlated with lignin depolymerization (detection by fluorescence images). All IRB oxidized the lignin contained in the SOM within a wide pH range and in soils from all parent materials. We present a conceptual model showing electron shuttling from SOM containing lignin (as a C and energy source) to IRB to produce energy and promote Fe(III) (oxyhydr)oxides reduction was proposed and discussed.

Effect of MnTBAP on in vitro capacitation of frozen-thawed stallion sperm.

In vitro manipulation of spermatozoa leads to deleterious changes of structure and function that occur mainly due to oxidative stress, therefore, prevention or treatment is a strategy to improve the functions of processed sperm. In the present study, the aim was to evaluate the effects of MnTBAP supplementation, a compound with antioxidant activity, on in vitro capacitation conditions of thawed equine sperm. For this purpose, stallion spermatozoa (2 × 106 cells/mL) were incubated in the sperm-TLP base medium for 4 h in which there were three different conditions: non-capacitating, capacitating, and capacitating plus 150 mM MnTBAP. There were incubations for 4 h at 37.5 °C in a humidified air atmosphere. Sample analysis was performed immediately after thawing (0 h), and at the end of the incubation period (4 h), unless otherwise indicated. The following variables were evaluated for spermatozoa: plasma membrane integrity and fluidity, acrosome integrity, intracellular calcium concentrations, intracellular pH, tyrosine phosphorylation, ATP concentrations, motility and heterologous zona-binding assay, using flow cytometry, fluorescent microscopy and/or chemiluminescence, depending on the most appropriate procedure for the variable being evaluated. Results indicated that capacitation-like changes were synergistically induced by the cAMP agonists, phosphodiesterase inhibitor and bicarbonate. The presence of bovine serum albumin was harmful to the plasma membrane. The MnTBAP supplementation had a positive effect on viability-related markers (plasma membrane integrity, membrane fluidity, associated with greater intracellular pH) when there were capacitating conditions. In conclusion, the activity of MnTBAP contributes to improving the in vitro incubation conditions of frozen-thawed stallion sperm.

Aristoteline, an Indole-Alkaloid, Induces Relaxation by Activating Potassium Channels and Blocking Calcium Channels in Isolated Rat Aorta.

Alkaloids derived from plants have shown great medicinal benefits, and are often reported for their use in cardiovascular disease management. Aristotelia chilensis (Molina) Stuntz (Maqui) has shown important medicinal properties in traditional useage. In this study, we evaluated the effect of the indole-alkaloid aristoteline (ARI), isolated from leaves of Maqui, on vascular reactivity of isolated aortic rings from normotensive rats. ARI induced relaxation (100%) in a concentration-dependent manner in intact or denuded-endothelium aortic rings pre-contracted with phenylephrine (PE; 1 µM). However, a specific soluble guanylyl cyclase inhibitor (ODQ; 1 µM) significantly reduced the relaxation to ARI in aortic rings pre-contracted with PE. In the presence of ARI, the contraction induced by KCl or PE was significantly (p < 0.05) decreased. Interestingly, the potassium channel blockade with 10 µM BaCl2 (Kir), 10 µM glibenclamide (KATP), 1 mM tetraethylammonium (TEA; KCa1.1), or 1 mM 4-aminopyridine (4-AP; Kv) significantly (p < 0.05) reduced the ARI-induced relaxation. ARI significantly (p < 0.05) reduced the contractile response to agonist of CaV1.2 channels (Bay K8644; 10 nM), likely reducing the influx of extracellular calcium through plasma membrane. The mechanisms associated with this process suggest an activation of the potassium channels, a calcium-induced antagonism and endothelium independent vasodilation that possibly involves the nitric oxide-independent soluble guanylate cyclase pathway.

Antioxidant Effect of a Polyphenol-Rich Murtilla (Ugni molinae Turcz.) Extract and Its Effect on the Regulation of Metabolism in Refrigerated Boar Sperm.

The production of reactive oxygen species (ROS) in boar spermatozoa increases in refrigeration; this can have an impact on sperm quality and fertilization capacity. We evaluated the effect of polyphenol-rich aqueous extract of murtilla (Ugni molinae Turcz) on boar sperm stored at 17°C in order to reduce oxidative stress and improve sperm quality in the long term. Five experiments were performed: first, characterization of the polyphenol content from five genotypes of murtilla; second, determination of the genotype with the best antioxidant effect (MT-Ex); third, the antioxidant capacity on O2 - and lipid peroxidation; fourth, the influence of MT-Ex on motility, calcium movement, cAMP, and metabolic parameters; and fifth, analysis of long-term refrigeration. The average phenolic content was 344 ppm; gallic acid, catechin, quercetin, myricetin, and kaempferol were detected. All extracts evaluated presented a concentration-dependent antioxidant effect. MT-Ex reduces intracellular O2 -/peroxides but low lipid peroxidation. MT-Ex in nonstimulated ROS conditions reduces sperm motility, mitochondrial membrane potential, cAMP, and ATP, but the succinate dehydrogenase activity remained normal; also, we observed a reduction in calcium movement in in vitro sperm capacitation. The long-term analyses showed that MT-Ex improved sperm motility decay and reduced membrane damage and ROS at 168 h. Based on this study, we propose MT-Ex as a supplement in semen extenders.

Functional and transcriptomic characterization of carboplatin-resistant A2780 ovarian cancer cell line.

BACKGROUND: Ovarian cancer is a significant cancer-related cause of death in women worldwide. The most used chemotherapeutic regimen is based on carboplatin (CBDCA). However, CBDCA resistance is the main obstacle to a better prognosis. An in vitro drug-resistant cell model would help in the understanding of molecular mechanisms underlying this drug-resistance phenomenon. The aim of this study was to characterize cellular and molecular changes of induced CBDCA-resistant ovarian cancer cell line A2780. METHODS: The cell selection strategy used in this study was a dose-per-pulse method using a concentration of 100 µM for 2 h. Once 20 cycles of exposure to the drug were completed, the cell cultures showed a resistant phenotype. Then, the ovarian cancer cell line A2780 was grown with 100 µM of CBDCA (CBDCA-resistant cells) or without CBDCA (parental cells). After, a drug sensitivity assay, morphological analyses, cell death assays and a RNA-seq analysis were performed in CBDCA-resistant A2780 cells. RESULTS: Microscopy on both parental and CBDCA-resistant A2780 cells showed similar characteristics in morphology and F-actin distribution within cells. In cell-death assays, parental A2780 cells showed a significant increase in phosphatidylserine translocation and caspase-3/7 cleavage compared to CBDCA-resistant A2780 cells (P < 0.05 and P < 0.005, respectively). Cell viability in parental A2780 cells was significantly decreased compared to CBDCA-resistant A2780 cells (P < 0.0005). The RNA-seq analysis showed 156 differentially expressed genes (DEGs) associated mainly to molecular functions. CONCLUSION: CBDCA-resistant A2780 ovarian cancer cells is a reliable model of CBDCA resistance that shows several DEGs involved in molecular functions such as transmembrane activity, protein binding to cell surface receptor and catalytic activity. Also, we found that the Wnt/ß-catenin and integrin signaling pathway are the main metabolic pathway dysregulated in CBDCA-resistant A2780 cells.

Impact of post-thaw supplementation of semen extender with antioxidants on the quality and function variables of stallion spermatozoa.

During cryopreservation procedures, the spermatozoa are exposed to physical and chemical stressors that generate an increase in the intracellular concentration of reactive oxygen species (ROS). If ROS concentrations are too great, this can lead to a state of oxidative stress that are detrimental to sperm quality. The aim of this study was to ascertain the profile the ROS production and assess the effects of post-thaw supplementation of a semen extender with different antioxidant compounds on the quality and function variables of frozen-thawed stallion spermatozoa incubated in vitro. Frozen-thawed stallion spermatozoa (2 × 106 cells/mL) were incubated with three different antioxidants (MnTBAP, NAC and FeTPPS) for 4 h at 38 °C. An untreated sperm suspension and a fresh sample were included as controls. Plasma membrane integrity (SYBR-14/PI), intracellular ROS concentration (DHE and ROS-ID™ total ROS/Superoxide Detection Kit), lipid peroxidation (BODIPY), DNA damage (TUNEL) and mitochondrial membrane potential (ΔΨm; TMRE/SYTOX) were evaluated by flow cytometry and fluorescence microscopy. In addition, sperm motility was evaluated using the ISAS system. Evaluations were performed at 0 and 4 h of incubation. The results indicate that superoxide anion is the main ROS produced by frozen-thawed stallion spermatozoa and that the use of MnTBAP improved sperm motility and viability, decreased the lipid peroxidation and DNA damage. In conclusion, this study provides relevant data to improve in vitro incubations conditions and to establish futures therapies using MnTBAP after thawing with the aim being to overcome the deleterious effects of semen cryopreservation and consequently preserve the stallion sperm quality through avoiding oxidative stress.

Functional and transcriptomic characterization of carboplatin-resistant A2780 ovarian cancer cell line

BACKGROUND: Ovarian cancer is a significant cancer-related cause of death in women worldwide. The most used chemotherapeutic regimen is based on carboplatin (CBDCA). However, CBDCA resistance is the main obstacle to a better prognosis. An in vitro drug-resistant cell model would help in the understanding of molecular mechanisms underlying this drug-resistance phenomenon. The aim of this study was to characterize cellular and molecular changes of induced CBDCA-resistant ovarian cancer cell line A2780. METHODS: The cell selection strategy used in this study was a dose-per-pulse method using a concentration of 100 µM for 2 h. Once 20 cycles of exposure to the drug were completed, the cell cultures showed a resistant phenotype. Then, the ovarian cancer cell line A2780 was grown with 100 µM of CBDCA (CBDCA-resistant cells) or without CBDCA (parental cells). After, a drug sensitivity assay, morphological analyses, cell death assays and a RNA-seq analysis were performed in CBDCA-resistant A2780 cells. RESULTS: Microscopy on both parental and CBDCA-resistant A2780 cells showed similar characteristics in morphology and F-actin distribution within cells. In cell-death assays, parental A2780 cells showed a significant increase in phosphatidylserine translocation and caspase-3/7 cleavage compared to CBDCA-resistant A2780 cells (P < 0.05 and P < 0.005, respectively). Cell viability in parental A2780 cells was significantly decreased compared to CBDCA-resistant A2780 cells (P < 0.0005). The RNA-seq analysis showed 156 differentially expressed genes (DEGs) associated mainly to molecular functions. CONCLUSION: CBDCA-resistant A2780 ovarian cancer cells is a reliable model of CBDCA resistance that shows several DEGs involved in molecular functions such as transmembrane activity, protein binding to cell surface receptor and catalytic activity. Also, we found that the Wnt/3-catenin and integrin signaling pathway are the main metabolic pathway dysregulated in CBDCA-resistant A2780 cells.

Antioxidant and Vasodilator Activity of Ugni molinae Turcz. (Murtilla) and Its Modulatory Mechanism in Hypotensive Response.

Hypertension is a systemic condition with high morbidity and mortality rates worldwide, which poses an increased risk for cardiovascular diseases. In this study, we demonstrated the antioxidant and vasodilator activity of Ugni molinae Turcz. (Murtilla) fruit, a berry native to Chile and proposed models to explain its modulatory mechanism in hypotensive response. Murtilla fruits were cultivated in a germplasm bank and submitted to chemical and biological analyses. The phenolic compounds gallic acid, Catechin, Quercetin-3-ß-D-glucoside, Myricetin, Quercetin, and Kaempferol were identified. Murtilla extract did not generate toxic effects on human endothelial cells and had significant antioxidant activity against ROS production, lipid peroxidation, and superoxide anion production. Furthermore, it showed dose-dependent vasodilator activity in aortic rings in the presence of endothelium, whose hypotensive mechanism is partially mediated by nitric oxide synthase/guanylate cyclase and large-conductance calcium-dependent potassium channels. Murtilla fruits might potentially have beneficial effects on the management of cardiovascular diseases.

Oil bodies as a potential microencapsulation carrier for astaxanthin stabilisation and safe delivery.

Astaxanthin (AST) is a valued molecule because of its high antioxidant properties. However, AST is extremely sensitive to oxidation, causing the loss of its bioactive properties. The purposes of this study were to define conditions for microencapsulating AST in oil bodies (OB) from Brassica napus to enhance its oxidative stability, and to test the bioactivity of the microencapsulated AST (AST-M) in cells. Conditions for maximising microencapsulation efficiency (ME) were determined using the Response Surface Methodology, obtaining a high ME (>99%). OB loaded with AST showed a strong electrostatic repulsion in a wide range of pH and ionic strengths. It was found that AST-M exposed to air and light was more stable than free AST. In addition, the protective effect of AST against intracellular ROS production was positively influenced by microencapsulation in OB. These results suggest that OB offer a novel option for stabilising and delivering AST.