Calcium enhanced the resistance against Phoma arachidicola by improving cell membrane stability and regulating reactive oxygen species metabolism in peanut.

BACKGROUND: Peanut (Arachis hypogaea), a vital oil and food crop globally, is susceptible to web blotch which is a significant foliar disease caused by Phoma arachidicola Marasas Pauer&Boerema leading to substantial yield losses in peanut production. Calcium treatment has been found to enhance plant resistance against pathogens. RESULTS: This study investigates the impact of exogenous calcium on peanut resistance to web blotch and explores its mechanisms. Greenhouse experiments revealed that exogenous calcium treatment effectively enhanced resistance to peanut web blotch. Specifically, amino acid calcium and sugar alcohol calcium solutions demonstrated the best induced resistance effects, achieving reduction rates of 61.54% and 60% in Baisha1016, and 53.94% and 50% in Luhua11, respectively. All exogenous calcium treatments reduced malondialdehyde (MDA) and relative electrical conductivity (REC) levels in peanut leaves, mitigating pathogen-induced cell membrane damage. Exogenous calcium supplementation led to elevated hydrogen peroxide (H2O2) content and superoxide anion (O2∙-) production in peanut leaves, facilitating the accumulation of reactive oxygen species (ROS) crucial for plant defense responses. Amino acid calcium and sugar alcohol calcium treatments significantly boosted activities of peroxidase (POD), superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) in peanut leaves. Activation of these antioxidant enzymes effectively scavenged excess ROS, maintaining ROS balance and mitigating cellular damage. CONCLUSIONS: In summary, exogenous calcium treatment triggered ROS production, which was subsequently eliminated by the activation of antioxidant enzymes, thereby reducing cell membrane damage and inducing defense responses against peanut web blotch.

The antimicrobial property of JY-1, a complex mixture of Traditional Chinese Medicine, is linked to it abilities to suppress biofilm formation and disrupt membrane permeability.

The substantial increase of infections, caused by novel, sudden, and drug-resistant pathogens, poses a significant threat to human health. While numerous studies have demonstrated the antibacterial and antiviral effects of Traditional Chinese Medicine, the potential of a complex mixture of traditional Chinese Medicine with a broad-spectrum antimicrobial property remains underexplored. This study aimed to develop a complex mixture of Traditional Chinese Medicine (TCM), JY-1, and investigate its antimicrobial properties, along with its potential mechanism of action against pathogenic microorganisms. Antimicrobial activity was assessed using a zone of inhibition assay and the drop plate method. Hyphal induction of Candida albicans was conducted using RPMI1640 medium containing 10% FBS, followed by microscopic visualization. Quantitative real-time PCR (RT-qPCR) was employed to quantify the transcript levels of hyphal-specific genes such as HWP1 and ALS3. The impact of JY-1 on biofilm formation was evaluated using both the XTT reduction assay and scanning electron microscopy (SEM). Furthermore, the cell membrane integrity was assessed by protein and nucleic acid leakage assays. Our results clearly showed that JY-1 significantly inhibits the vegetative growth of Candida spp. and Cryptococcus spp. In addition, this complex mixture is effectively against a wide range of pathogenic bacteria, including Staphylococcus aureus, Vancomycin-resistant enterococci, Escherichia coli, Klebsiella pneumoniae and Enterobacter cloacae. More interestingly, JY-1 plays a direct anti-viral role against the mammalian viral pathogen vesicular stomatitis virus (VSV). Further mechanistic studies indicate that JY-1 acts to reduce the expression of hyphal specific genes HWP1 and ALS3, resulting in the suppression of the hyphal formation of C. albicans. The antimicrobial property of JY-1 could be attributed to its ability to reduce biofilm formation and disrupt the cell membrane permeability, a process resulting in microbial cell death and the release of cellular contents. Taken together, our work identified a potent broad-spectrum antimicrobial agent, a complex mixture of TCM which might be developed as a potential antimicrobial drug.

Elevated Serum Levels of Zonulin Family Peptides in Anticitrullinated Protein Antibody-Positive At-Risk Individuals Without Arthritis.

OBJECTIVE: Recent advances imply that early events triggering rheumatoid arthritis (RA) occur at mucosal surfaces. We aimed to evaluate whether intestinal permeability is altered in patients at increased risk of RA, and/or predicts the development of clinical arthritis, by measuring serum zonulin family peptides (ZFP) levels, which are shown to reflect intestinal barrier integrity. METHODS: Two independent prospective observational cohorts were studied, including subjects with musculoskeletal symptoms and anticitrullinated protein antibodies (ACPA), but without clinical arthritis at baseline. In Sweden, 82 such at-risk patients were compared to 100 age-matched healthy blood donors. In the UK, 307 at-risk patients were compared to 100 ACPA-negative symptomatic controls. ZFP was measured in baseline sera by enzyme-linked immunoassays. RESULTS: In the Swedish at-risk cohort, ZFP levels were significantly increased in patients compared to controls (mean 41.4 vs 33.6 ng/mL, P < 0.001) and Cox regression analysis showed prognostic value of ZFP for arthritis development (hazard ratio [HZ] 1.04 per ng/mL ZFP increase, 95% CI 1.01-1.07, P = 0.02). Elevated ZFP levels among ACPA-positive at-risk patients compared to symptomatic ACPA-negative controls were confirmed in the UK at-risk cohort (mean 69.7 vs 36.0 ng/mL, P < 0.001), but baseline ZFP were not associated with arthritis development (HR 1.00 per ng/mL ZFP increase, 95% CI 1.00-1.01, P = 0.30). CONCLUSION: Serum ZFP levels are elevated in ACPA-positive at-risk patients when compared to both healthy blood donors and symptomatic ACPA-negative controls. Thus, gut barrier function may be of importance in RA-associated autoimmunity. A possible prognostic value of serum ZFP merits further investigation, preferably in larger prospective cohorts.

Understanding Stimulation of Conjugal Gene Transfer by Nonantibiotic Compounds: How Far Are We?

A myriad of nonantibiotic compounds is released into the environment, some of which may contribute to the dissemination of antimicrobial resistance by stimulating conjugation. Here, we analyzed a collection of studies to (i) identify patterns of transfer stimulation across groups and concentrations of chemicals, (ii) evaluate the strength of evidence for the proposed mechanisms behind conjugal stimulation, and (iii) examine the plausibility of alternative mechanisms. We show that stimulatory nonantibiotic compounds act at concentrations from 1/1000 to 1/10 of the minimal inhibitory concentration for the donor strain but that stimulation is always modest (less than 8-fold). The main proposed mechanisms for stimulation via the reactive oxygen species/SOS cascade and/or an increase in cell membrane permeability are not unequivocally supported by the literature. However, we identify the reactive oxygen species/SOS cascade as the most likely mechanism. This remains to be confirmed by firm molecular evidence. Such evidence and more standardized and high-throughput conjugation assays are needed to create technologies and solutions to limit the stimulation of conjugal gene transfer and contribute to mitigating global antibiotic resistance.

Structural optimization of a lysine demethylase 5 inhibitor for improvement of its cellular activity.

Lysine demethylase 5 (KDM5) subfamily proteins are important in epigenetic gene regulation. They are involved in the growth and drug resistance of cancer cells. Therefore, KDM5s are potential cancer therapeutic targets, and their inhibitors hold promise as anti-cancer drugs. Several KDM5 inhibitors, including KDM5-C49 (2a), have exhibited potent KDM5-inhibitory activities in in vitro enzyme assays. However, they do not show enough cellular activity despite being converted to their prodrugs. We hypothesized that their poor lipophilicity should prevent them from sufficiently penetrating the cell membrane, and introducing more lipophilic groups should improve cellular activities. In this study, we investigated 2a and KDM5-C70 (3a), a prodrug of 2a, and attempted to improve its cellular activity by replacing the N,N-dimethyl amino group of 3a with more lipophilic groups. N-Butyl, N-methyl amino compound 2e exhibited potent and selective KDM5-inhibitory activity equal to that of 2a. Furthermore, the cell membrane permeability of 3e, an ethyl ester prodrug of 2e, was six times higher than that of 3a in a parallel artificial membrane permeation assay. In addition, western blot analysis indicated that treating human lung cancer A549 cells with 3e increased histone methylation levels more strongly than that with 3a. Thus, we identified compound 3e as a more cell-active KDM5 inhibitor that has sufficient cell membrane permeability.

Deletion of ArmPT, a LamB-like protein, increases cell membrane permeability and antibiotic sensitivity in Vibrio alginolyticus.

Vibrio bacterial species are dominant pathogens in mariculture animals. However, the extensive use of antibiotics and other chemicals has increased drug resistance in Vibrio bacteria. Despite rigorous investigative studies, the mechanism of drug resistance in Vibrio remains a mystery. In this study, we found that a gene encoding LamB-like outer membrane protein, named ArmPT, was upregulated in Va under antibiotic stress by RT-qPCR. We speculated that ArmPT might play a role in Va's drug resistance. Subsequently, using ArmPT gene knockout and gene complementation experiments, we confirmed its role in resistance against a variety of antibiotics, particularly kanamycin (KA). Transcriptomic and proteomic analyses identified 188 and 83 differentially expressed genes in the mutant strain compared with the wild-type (WT) before and after KA stress, respectively. Bioinformatic analysis predicted that ArmPT might control cell membrane permeability by changing cadaverine biosynthesis, thereby influencing the cell entry of antibiotics in Va. The higher levels of intracellular reactive oxygen species and the infused content of KA showed that antibiotics are more likely to enter the Va mutant strain. These results uncover the drug resistance mechanism of Va that can also exist in other similar pathogenic bacteria.

Design, synthesis and antifungal activity of novel amide derivatives containing a pyrrolidine moiety as potential succinate dehydrogenase inhibitors.

To discover new succinate dehydrogenase inhibitors (SDHI) fungicides, a series of amide derivatives containing a pyrrolidine moiety were designed and synthesized, and their antifungal activities were evaluated against Monilinia fructicola (M. fructicola), Rhizoctonia solani (R. solani), Fusarium graminearum schw (F. graminearum), Fusarium oxysporum (F. oxysporum), and Phytophthora infestans (P. infestans). Some compounds showed excellent antifungal activities against the five fungi. Among them, compound 6 showed broad-spectrum inhibitory activities. The EC50 of compound 6 against M. fructicola, R. solani, F. graminearum, F. oxysporum, and P. infestans were 2.13, 14.42, 1.69, 27.79, and 27.12 mg/L, respectively. In addition, compound 6 can effectively inhibit the spore germination of M. fructicola and has moderate damage to the cell membrane. Compound 6 can effectively inhibit succinate dehydrogenase (SDH) of M. fructicola, and can significantly increase the expression levels of SDHC and SDHD. Compound 6 can be used as a lead structure for developing new SDH inhibitors.

Ascendancy of pyraclostrobin nanocapsule formulation against Rhizoctonia solani: From a perspective of fungus.

High-performance pesticide formulations are essential for sustainable agriculture. Among these, nano-pesticides exhibit great advantages in pest control because of their unique size effects. However, the direct effects of nano-formulation fungicides on fungal pathogens remain largely unexplored. In this study, three qualified formulations, suspension concentrate (SC), microcapsules (CS), and nanocapsules (NCS) of pyraclostrobin (PYR) were prepared and utilized to reveal their biocontrol activities against Rhizoctonia solani. Among these three formulations, NCS exhibited notable biocontrol efficacy against R. solani exemplified by an EC50 of 0.319 mg/L for mycelia, distortion of mycelia and abnormalities in cell ultrastructure. Moreover, NCS displayed excellent internalization within R. solani mycelia, contributing to severe damage to cell membrane permeability. Importantly, an equivalent quantity of NCS-PYR showed potent inhibitory effects on the target pathogen, as indicated by reduced adenosine triphosphate (ATP) content and mitochondrial Complex III activity. The NCS consistently exhibited superior in vivo protective and curative activities against R. solani compared to those of CS and SC in rice and faba bean. In summary, we uncovered the strength of rapid efficacy and biocontrol activity of NCS against R. solani and elucidated the advantages of NCS-PYR from the perspective of the target pathogen in agriculture.

Neurite Exchange Imaging (NEXI): A minimal model of diffusion in gray matter with inter-compartment water exchange.

Biophysical models of diffusion in white matter have been center-stage over the past two decades and are essentially based on what is now commonly referred to as the "Standard Model" (SM) of non-exchanging anisotropic compartments with Gaussian diffusion. In this work, we focus on diffusion MRI in gray matter, which requires rethinking basic microstructure modeling blocks. In particular, at least three contributions beyond the SM need to be considered for gray matter: water exchange across the cell membrane - between neurites and the extracellular space; non-Gaussian diffusion along neuronal and glial processes - resulting from structural disorder; and signal contribution from soma. For the first contribution, we propose Neurite Exchange Imaging (NEXI) as an extension of the SM of diffusion, which builds on the anisotropic Kärger model of two exchanging compartments. Using datasets acquired at multiple diffusion weightings (b) and diffusion times (t) in the rat brain in vivo, we investigate the suitability of NEXI to describe the diffusion signal in the gray matter, compared to the other two possible contributions. Our results for the diffusion time window 20-45 ms show minimal diffusivity time-dependence and more pronounced kurtosis decay with time, which is well fit by the exchange model. Moreover, we observe lower signal for longer diffusion times at high b. In light of these observations, we identify exchange as the mechanism that best explains these signal signatures in both low-b and high-b regime, and thereby propose NEXI as the minimal model for gray matter microstructure mapping. We finally highlight multi-b multi-t acquisition protocols as being best suited to estimate NEXI model parameters reliably. Using this approach, we estimate the inter-compartment water exchange time to be 15 - 60 ms in the rat cortex and hippocampus in vivo, which is of the same order or shorter than the diffusion time in typical diffusion MRI acquisitions. This suggests water exchange as an essential component for interpreting diffusion MRI measurements in gray matter.

A study of the antibacterial mechanism of pinocembrin against multidrug-resistant Aeromonas hydrophila.

Aeromonas hydrophila is a common pathogen in fish that has caused severe economic losses in aquaculture worldwide. With the emergence of bacterial resistance, it is necessary to develop new drugs to combat bacterial infection, particularly for multidrug-resistant bacteria. In this study, the antibacterial activity of pinocembrin was investigated by observing bacterial growth and microscopic structure, and its mechanism of action was identified by investigating its effect on protein and DNA. The antibacterial susceptibility test indicated that pinocembrin inhibits A. hydrophila growth. The minimal inhibitory concentration and minimum bactericidal concentration were 256 µg/mL and 512 µg/mL, respectively. Ultrastructurally, the bacteria treated with pinocembrin showed surface roughness and plasmolysis. When bacteria were treated with 512 µg/mL pinocembrin, lactate dehydrogenase activity and soluble protein content decreased significantly, and electrical conductivity and DNA exosmosis levels increased by 4.21 ± 0.64% and 15.98 ± 1.93 mg/L, respectively. Staining with 4', 6-Diamidino-2-phenylindole showed that the nucleic acid fluorescence intensity and density decreased after the treatment with pinocembrin. Pinocembrin may inhibit the growth of A. hydrophila by increasing cell membrane permeability and affecting protein and DNA metabolism. Thus, pinocembrin is a candidate drug for the treatment of A. hydrophila infection in aquaculture.

Antagonistic activity of selenium-enriched Bifidobacterium breve against Clostridioides difficile.

Probiotics have the potential to be used in the prevention of Clostridioides difficile infection (CDI). In this study, selenium (Se)-enriched Bifidobacterium breve YH68-Se was obtained under optimal culture conditions with single-factor and response surface optimization. The overall environmental resistance of YH68-Se was superior to that of the parental strain YH68, mainly reflected in the substantial improvement of antioxidant activity and gastrointestinal tolerance. YH68-Se dramatically inhibited C. difficile growth, spore, biofilm, toxin production, and virulence gene expression, rapidly disrupted C. difficile cell membrane permeability and integrity, and altered the membrane proton motive force (PMF), induced a large outflow of intracellular substances and eventually caused bacterial death. The main factor inducing this process originated from the lactic acid (LD) in YH68-Se. In addition, the LD production of YH68 increased with increasing selenite concentration and was accompanied by enhanced activities of thioredoxin reductase (TrxR), glutathione peroxidase (GSH-Px), and increased concentration of autoinducer-2 (AI-2), which may be the crucial factors contributing to the outstanding probiotic properties of YH68-Se and their potent antagonism of C. difficile. KEY POINTS: • Compared with the parental strain B. breve YH68, the environmental resistance of YH68-Se was improved. • YH68-Se was able to produce more lactic acid, which suppressed the important physiological activities of C. difficile and rapidly disrupted their cell membrane structures. • Sodium selenite in the suitable concentration range gradually increases the yield of lactic acid and phenylacetic acid, increased the concentration of autoinducer-2, and enhanced the activities of antioxidant enzymes TrxR and GSH-Px in YH68.

Enhancement of triterpene production via in situ extractive fermentation of Sanghuangporus vaninii YC-1.

There have been many studies on the activities and polysaccharide production of Sanghuangporus vaninii. However, few studies have looked at triterpene production from S. vaninii using liquid-state fermentation. A method for enhancing the production of triterpenes by in situ extractive fermentation (ISEF) was studied. Eight solvents were investigated as extractants for triterpene production in the ISEF system. The results showed that using vegetable oil as an extractant significantly increased the yield of total triterpenes and biomass of S. vaninii YC-1, reaching 18.98 ± 0.71 and 44.67 ± 2.21 g/L, respectively. In 5 L fermenter experiments, the added vegetable oil improved the dissolved oxygen condition of the fermentation broth and promoted the growth of S. vaninii YC-1. Furthermore, adding vegetable oil increased the expression of fatty acid synthesis-related genes such as FAD2 and SCD, thereby increasing the synthesis of unsaturated fatty acids in the cell membrane of S. vaninii YC-1. Therefore, the cell membrane permeability of S. vaninii YC-1 increased by 19%. Our results indicated that vegetable oil increased the permeability of S. vaninii YC-1 cell membranes to promote the production of total triterpenes. The use of vegetable oil as an extractant was thus effective in increasing the yield of triterpenes in the ISEF system.

Inactivation kinetics and cell envelope damages of foodborne pathogens Listeria monocytogenes and Salmonella Enteritidis treated with cold plasma.

In recent years, more attention has been paid to the application of cold plasma (CP) in eliminating foodborne pathogenic bacteria. This work investigated CP effects on inactivation kinetics and cell envelopes of Listeria monocytogenes (L. monocytogenes) and Salmonella Enteritidis (S. Enteritidis). Bacterial suspensions were treated with dielectric barrier discharge atmospheric CP at 75 kV for different treatment time. Three regression models were tested for estimating inactivation kinetics. Reactive species generated in plasma, the appearance and integrity of bacterial cells, the activity and secondary structure of enzymes in the cell envelope, and molecular docking, were measured for evaluating the envelope damages. Results indicated that Log-linear model was suitable for L. monocytogenes and the Weibull model was suitable for S. Enteritidis. S. Enteritidis was more sensitive to short-lived reactive species (such as OH radicals) in plasma than L. monocytogenes, and the cell envelope of S. Enteritidis was more severely damaged (the increased membrane permeability and leakage of intracellular substances) after plasma treatment. Interestingly, compared with S. Enteritidis, the decrease in the activity of enzymes existing in the cell envelope of L. monocytogenes did not contribute significantly to the death of bacteria. Molecular docking further suggested that the decrease in the enzyme activity might be due to the modification of the enzyme, by the interaction between reactive species in plasma (H2O2) and amino acid residues of the enzyme through the hydrogen bond.

The Cooperative Anti-Neoplastic Activity of Polyphenolic Phytochemicals on Human T-Cell Acute Lymphoblastic Leukemia Cell Line MOLT-4 In Vitro.

Acute lymphoblastic leukemia (ALL) is the most common hematological malignancy affecting pediatric patients. ALL treatment regimens with cytostatics manifest substantial toxicity and have reached the maximum of well-tolerated doses. One potential approach for improving treatment efficiency could be supplementation of the current regimen with naturally occurring phytochemicals with anti-cancer properties. Nutraceuticals such as quercetin, curcumin, resveratrol, and genistein have been studied in anti-cancer therapy, but their application is limited by their low bioavailability. However, their cooperative activity could potentially increase their efficiency at low, bioavailable doses. We studied their cooperative effect on the viability of a human ALL MOLT-4 cell line in vitro at the concentration considered to be in the bioavailable range in vivo. To analyze their potential side effect on the viability of non-tumor cells, we evaluated their toxicity on a normal human foreskin fibroblast cell line (BJ). In both cell lines, we also measured specific indicators of cell death, changes in cell membrane permeability (CMP), and mitochondrial membrane potential (MMP). Even at a low bioavailable concentration, genistein and curcumin decreased MOLT-4 viability, and their combination had a significant interactive effect. While resveratrol and quercetin did not affect MOLT-4 viability, together they enhanced the effect of the genistein/curcumin mix, significantly inhibiting MOLT-4 population growth in vitro. Moreover, the analyzed phytochemicals and their combinations did not affect the BJ cell line. In both cell lines, they induced a decrease in MMP and correlating CMP changes, but in non-tumor cells, both metabolic activity and cell membrane continuity were restored in time. (4) Conclusions: The results indicate that the interactive activity of analyzed phytochemicals can induce an anti-cancer effect on ALL cells without a significant effect on non-tumor cells. It implies that the application of the combinations of phytochemicals an anti-cancer treatment supplement could be worth further investigation regardless of their low bioavailability.

Herbicide promotes the conjugative transfer of multi-resistance genes by facilitating cellular contact and plasmid transfer.

The global dissemination of antibiotic resistance genes (ARGs), especially via plasmid-mediated horizontal transfer, is becoming a pervasive health threat. While our previous study found that herbicides can accelerate the horizontal gene transfer (HGT) of ARGs in soil bacteria, the underlying mechanisms by which herbicides promote the HGT of ARGs across and within bacterial genera are still unclear. Here, the underlying mechanism associated with herbicide-promoted HGT was analyzed by detecting intracellular reactive oxygen species (ROS) production, extracellular polymeric substance composition, cell membrane integrity and proton motive force combined with genome-wide RNA sequencing. Exposure to herbicides induced a series of the above bacterial responses to promote HGT except for the ROS response, including compact cell-to-cell contact by enhancing pilus-encoded gene expression and decreasing cell surface charge, increasing cell membrane permeability, and enhancing the proton motive force, providing additional power for DNA uptake. This study provides a mechanistic understanding of the risk of bacterial resistance spread promoted by herbicides, which elucidates a new perspective on nonantibiotic agrochemical acceleration of the HGT of ARGs.

LptB-LptF coupling mediates the closure of the substrate-binding cavity in the LptB2 FGC transporter through a rigid-body mechanism to extract LPS.

Lipopolysaccharides (LPS) are essential envelope components in many Gram-negative bacteria and provide intrinsic resistance to antibiotics. LPS molecules are synthesized in the inner membrane and then transported to the cell surface by the LPS transport (Lpt) machinery. In this system, the ATP-binding cassette (ABC) transporter LptB2 FGC extracts LPS from the inner membrane and places it onto a periplasmic protein bridge through a poorly understood mechanism. Here, we show that residue E86 of LptB is essential for coupling the function of this ATPase to that of its partners LptFG, specifically at the step where ATP binding drives the closure of the LptB dimer and the collapse of the LPS-binding cavity in LptFG that moves LPS to the Lpt periplasmic bridge. We also show that defects caused by changing residue E86 are suppressed by mutations altering either LPS structure or transmembrane helices in LptG. Furthermore, these suppressors also fix defects in the coupling helix of LptF, but not of LptG. Together, these results support a transport mechanism in which the ATP-driven movements of LptB and those of the substrate-binding cavity in LptFG are bi-directionally coordinated through the rigid-body coupling, with LptF's coupling helix being important in coordinating cavity collapse with LptB dimerization.

Selective Cadmium Fluorescence Probe Based on Bis-heterocyclic Molecule and its Imaging in Cells.

Fluorescence probes that selectively image cadmium are useful for detecting and tracking the amount of Cd2+ in cells and tissues. In this study, we designed and synthesized a novel Cd2+ fluorescence probe based on the pyridine-pyrimidine structure, 4-(methylsulfanyl)-6-(pyridin-2-yl)pyrimidin-2-amine (3), as a low-molecular-weight fluorescence probe for Cd2+. Compound 3 could successfully discriminate between Cd2+ and Zn2+ and exhibited a highly selective turn-on response toward Cd2+ over biologically related metal ions. The dissociation constant (Kd) and the limit of detection (LOD) of 5.4 × 10- 6 mol L- 1 and 4.4 × 10- 7 mol L- 1, respectively, were calculated using fluorescence titration experiments. Studies with closely related analogs showed that the bis-heterocyclic moiety of 3 acted as both a coordination site for Cd2+ and a fluorophore. Further, the methylsulfanyl group of compound 3 is essential for achieving selective and sensitive Cd2+ detection. Fluorescence microscopy studies using living cells revealed that the cell membrane permeability of compound 3 is sufficient to detect intracellular Cd2+. These results indicate that novel bis-heterocyclic molecule 3 has considerable potential as a fluorescence probe for Cd2+ in biological applications.

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.

Measuring diffusion exchange across the cell membrane with DEXSY (Diffusion Exchange Spectroscopy).

INTRODUCTION: To combine numerical simulations, in vitro and in vivo experiments to evaluate the feasibility of measuring diffusion exchange across the cell membrane with diffusion exchange spectroscopy (DEXSY). METHODS: DEXSY acquisitions were simulated over a range of permeabilities in nerve tissue and yeast substrates. In vitro measurements were performed in a yeast substrate and in vivo measurements in mouse tumor xenograft models, all at 9.4 T. RESULTS: Diffusion exchange was observed in simulations over a physiologically relevant range of cell permeability values. In vitro and in vivo measures also provided evidence of diffusion exchange, which was quantified with the Diffusion Exchange Index (DEI). CONCLUSIONS: Our findings provide preliminary evidence that DEXSY can be used to make in vivo measurements of diffusion exchange and cell membrane permeability.

Biological investigations of essential oils extracted from three Juniperus species and evaluation of their antimicrobial, antioxidant and cytotoxic activities.

AIMS: To conduct biological investigations and to evaluate the antimicrobial and antioxidant activities of the essential oils (EOs) extracted from Juniperus communis, J. scopulorum and J. horizontalis; to screen their mechanisms of action by conducting the cell membrane permeability assay (CMP); and to determine the possible cytotoxicity of the three EOs against human neuroblastoma cells (SH-SY5Y). METHODS AND RESULTS: The antifungal activity was tested against four phytopathogenic fungi (Monilinia fructicola, Aspergillus niger, Penicillium expansum and Botrytis cinerea). The antibacterial activity was evaluated against two Gram-positive (G+ve) (Bacillus megaterium and Clavibacter michiganensis) and three Gram-negative (G-ve) bacterial strains (Pseudomonas fluorescens, P. syringae pv. phaseolicola and Xanthomonas campestris). Results showed that the three tested EOs have antifungal activity against M. fructicola and P. expansum and effective antibacterial activity against P. syringae pv. phaseolicola and B. megaterium. Moreover, the three EOs were evaluated for their ability to inhibit the growth of SH-SY5Y cells with MTT assay. J. communis EO was the more effective with an IC50 of 53·7 µg ml-1 . The antioxidant capacity of the three EO did not differ as measured by the DPPH assay. CONCLUSIONS: The three tested juniper EOs showed promising antimicrobial and antioxidant activity and cytotoxic effects against human neuroblastoma cell line. SIGNIFICANCE AND IMPACT OF THE STUDY: The outfindings from this research showed promising antimicrobial effects of the three oils against the majority of the tested phytopathogens with a potential to utilize them as natural alternatives to synthetic drugs, the cause of global environmental problems, pathogen resistance and difficulty to control many post-harvest plant diseases.