Curcumin prevents alcohol-induced liver disease in rats by inhibiting the expression of NF-kappa B-dependent genes

Induction of NF-kappaB-mediated gene expression has been implicated in the pathogenesis of alcoholic liver disease (ALD). Curcumin, a phenolic antioxidant, inhibits the activation of NF-kappaB. We determined whether treatment with curcumin would prevent experimental ALD and elucidated the underlying mechanism. Four groups of rats (6 rats/group) were treated by intragastric infusion for 4 wk. One group received fish oil plus ethanol (FE); a second group received fish oil plus dextrose (FD). The third and fourth groups received FE or FD supplemented with 75 mg. kg(-1). day(-1) of curcumin. Liver samples were analyzed for histopathology, lipid peroxidation, NF-kappaB binding, TNF-alpha, IL-12, monocyte chemotactic protein-1, macrophage inflammatory protein-2, cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and nitrotyrosine. Rats fed FE developed fatty liver, necrosis, and inflammation, which was accompanied by activation of NF-kappaB and the induction of cytokines, chemokines, COX-2, iNOS, and nitrotyrosine formation. Treatment with curcumin prevented both the pathological and biochemical changes induced by alcohol. Because endotoxin and the Kupffer cell are implicated in the pathogenesis of ALD, we investigated whether curcumin suppressed the stimulatory effects of endotoxin in isolated Kupffer cells. Curcumin blocked endotoxin-mediated activation of NF-kappaB and suppressed the expression of cytokines, chemokines, COX-2, and iNOS in Kupffer cells. Thus curcumin prevents experimental ALD, in part by suppressing induction of NF-kappaB-dependent genes.     

Hospital Patient‐Care and Outside‐the‐Hospital Energy Profiles for Hemodialysis Services

Studies investigated the patient‐care (in‐hospital) and outside‐the‐hospital energy consumptions for delivering the hemodialysis (HD) service. A life cycle inventory methodology was used for this patient‐based analysis for two hospitals located in Wichita, Kansas. It was found that, for both hospitals, the actual HD machines consumed approximately 3.5 kilowatt‐hours (kWh) of electrical energy per HD, only 8% to 16% of the total energy used for delivering the HD service (in hospital). This increases to 9.6 to 28.9 kWh of hospital billable energy for the whole system of HD machine, auxiliaries, and dialysis water treatment. Converting these hospital direct electrical energy values to natural resource energy (nre) then adding the cradle‐to‐gate natural resource energy for the manufacturing and supply chain of all the HD consumables, the total is 78 to 149 kWh nre/HD. The nre measures all the direct fuel burned to generate energy and is thus directly related to emissions to the air, water, and land and is a direct secondary impact on public health from HD. The ratio of outside‐the‐hospital energy to direct hospital HD electrical energy consumption is 4:1 to 7:1, so a broader base exists for improvement than just the hospital.     

Isolation of a new ssDNA aptamer against staphylococcal enterotoxin B based on CNBr‐activated sepharose‐4B affinity chromatography

Staphylococcus aureus are potent human pathogens possessing arsenal of virulence factors. Staphylococcal food poisoning (SFP) and respiratory infections mediated by staphylococcal enterotoxin B (SEB) are common clinical manifestations. Many diagnostic techniques are based on serological detection and quantification of SEB in different food and clinical samples. Aptamers are known as new therapeutic and detection tools which are available in different ssDNA, dsDNA and protein structures. In this study, we used a new set of ssDNA aptamers against SEB. The methods used included preparation of a dsDNA library using standard SEB protein as the target analyte, affinity chromatography matrix in microfuge tubes, SELEX procedures to isolate specific ssDNA‐aptamer as an affinity ligand, aptamer purification using ethanol precipitation method, affinity binding assay using ELISA, aptamer cloning and specificity test. Among 12 readable sequences, three of them were selected as the most appropriate aptamer because of their affinity and specificity to SEB. This study presents a new set of ssDNA aptamer with favorable selectivity to SEB through 12 rounds of SELEX. Selected aptamers were used to detect SEB in infected serum samples. Results showed that SEB c1 aptamer (2 µg SEB/100 nM aptamer) had favorable specificity to SEB (k_d = 2.3 × 10^?11). In conclusion, aptamers can be considered as useful tools for detecting and evaluating SEB. The results showed that affinity chromatography was an affordable assay with acceptable accuracy to isolate sensitive and selective novel aptamers. Copyright © 2016 John Wiley & Sons, Ltd.     

Dopamine release and molecular modeling study of some coumarin derivatives

4-aryl-2-amino-6-(4-hydroxy-2-oxo-2H-chromen-3-yl)-pyridin-3-carbonitrile (1), 4-aryl-2-oxo-6-(4-hydroxy-2-oxo-2H-chromen-3-yl)-pyridin-3-carbonitriles (2a-2c), 3-(6-aryl-1,2,5,6- tetrahydro-2-thioxopyrimidin-4-yl)-4-hydroxy-2H-chromen-2-one (3a, 3b) and pyrazol-3-yl-4-hydroxycoumarin derivatives (4a-4c, 5, 6a, 6b, 7a, 7b, and 8a-8c) were prepared in order to measure their % change dopamine release in comparison to amphetamine as reference, using PC-12 cells in different concentrations. In addition, the molecular modeling study of the compounds into 3BHH receptor was also demonstrated. The calculated inhibition constant (k_i) implemented in the AutoDock program revealed identical correlation with the experimental results to that obtained binding free energy (ΔG_b) as both parameters revealed reasonable correlation coefficients (R²) being 0.51 involving 10 compounds; (1, 2b, 2c, 3a, 3b, 4a, 4b, 6a, and 8c).     

Multilamellar liposomes of triamcinolone acetonide: preparation,stability, and characterization

The aim of this study was to assess and characterize the stability of multilamellar liposomes as a delivery vehicle for triamcinolone acetonide. A standardized preparation method for a liposomal delivery vehicle was developed, after varying composition and storage conditions, and assessing encapsulation efficiency and loss of active principle. The assessment of temperature as a factor in formula stability during storage showed that stability improved under refrigeration (4–6°C) (less early diffusion of active principle through the liposomal wall), in comparison with samples stored at room temperature. To improve stability, cholesterol was added to some formulae, which although resulting in a decrease in average encapsulation efficiency, mitigated subsequent losses of retained active principle (formulae 4, 5, and 6), in comparison with those without cholesterol (formulae 1, 2, and 3). This was evident both under refrigerated and room-temperature conditions. Finally, after testing the effects of adding an antioxidant and/or preservative to the formulae, a liposomal design was achieved with acceptable stability, vesicle dimensions, and encapsulation efficiency.     

Structural basis for catalytic differences between alpha class human glutathione transferases hGSTA1-1 and hGSTA2-2 for glutathione conjugation of environmental carcinogen benzo[a]pyrene-7,8-diol-9,10-epoxide

The ultimate diol epoxide carcinogens derived from polycyclic aromatic hydrocarbons, such as benzo[a]pyrene (BP), are metabolized primarily by glutathione (GSH) conjugation reaction catalyzed by GSH transferases (GSTs). In human liver and probably lung, the alpha class GSTs are likely to be responsible for the majority of this reaction because of their high abundance. The catalytic efficiency for GSH conjugation of the carcinogenic (+)-anti-benzo[a]pyrene-7,8-diol-9,10-epoxide [(+)-anti-BPDE] is more than 5-fold higher for hGSTA1-1 than for hGSTA2-2. Here, we demonstrate that mutation of isoleucine-11 of hGSTA2-2, a residue located in the hydrophobic substrate-binding site (H-site) of the enzyme, to alanine (which is present in the same position in hGSTA1-1) results in about a 7-fold increase in catalytic efficiency for (+)-anti-BPDE-GSH conjugation. Thus, a single amino acid substitution is sufficient to convert hGSTA2-2 to a protein that matches hGSTA1-1 in its catalytic efficiency. The increased catalytic efficiency of hGSTA2/I11A is accompanied by greater enantioselectivity for the carcinogenic (+)-anti-BPDE over (-)-anti-BPDE. Further remodeling of the H-site of hGSTA2-2 to resemble that of hGSTA1-1 (S9F, I11A, F110V, and S215A mutations, SIFS mutant) results in an enzyme whose catalytic efficiency is approximately 13.5-fold higher than that of the wild-type hGSTA2-2, and about 2.5-fold higher than that of the wild-type hGSTA1-1. The increased activity upon mutations can be rationalized by the interactions of the amino acid side chains with the substrate and the orientation of the substrate in the active site, as visualized by molecular modeling. Interestingly, the catalytic efficiency of hGSTA2-2 toward (-)-anti-BPDE was increased to a level close to that of hGSTA1-1 upon F110V, not I11A, mutation. Similar to (+)-anti-BPDE, however, the SIFS mutant was the most efficient enzyme for GSH conjugation of (-)-anti-BPDE.     

In vitro response of apical bud explants from mature trees of jackfruit (Artocarpus heterophyllus)

A tissue culture technique for rapid vegetative propagation of mature jackfruit trees using apical bud cultures has been developed. Shoot-tip cultures were established on MS medium with 5–10 mm explants dissected from terminal buds of new growth from trunk. After initial culture of bud explants, one- to two-node pieces were taken from the microshoots formed and used to proliferate further axillary shoots for multiplying and maintaining shoot cultures. Benzyladenine and kinetin (4.5–9.0 µM), either separately or together, supported shoot proliferation; higher concentrations of the cytokinins inhibited bud breaking and favoured callus formation at the explant bases. Bud explants taken from emerging trunk sprouts invariably produced clumps of multiple shoots, whereas buds obtained from actively growing top branches generally elongated to form a solitary shoot. November to January was the best season for initiation of cultures from field-grown trees. Shoots proliferated at the initial subcultures had mature morphology and were difficult-to-root. Shoots assumed to be juvenile-like developed at the later passages and could be rooted with 60–80% success using 1/2-MS salts and 10 µM of indolebutyric acid or naphthaleneacetic acid. Regenerated plantlets were transferred to the soil and about 50% survived.     

Two-spotted spider mite reared on resistant eggplant affects consumption rate and life table parameters of its predator,Typhlodromus bagdasarjani (Acari: Phytoseiidae)

The blue tick, Rhipicephalus microplus, threatens cattle production in most tropical and subtropical areas of the world. Delayed skin hypersensitivity reactions are thought to cause Nguni cattle to be more resistant to R. microplus than Bonsmara cattle yet the cellular mechanisms responsible for these differences have not been classified. Tick counts and inflammatory cell infiltrates in skin biopsies from feeding sites of adult R. microplus ticks were determined in 9-month-old Nguni and Bonsmara heifers to determine the cellular mechanisms responsible for tick immunity. Nguni heifers (1.7 ± 0.03) had lower (P < 0.05) tick counts than the Bonsmaras (2.0 ± 0.03). Parasitized sites in Nguni heifers had higher counts of basophils, mast and mononuclear cells than those in the Bonsmara heifers. Conversely, parasitized sites in Nguni heifers had lower neutrophil and eosinophil counts than those in the Bonsmara heifers. Tick count was negatively correlated with basophil and mast cell counts and positively correlated with eosinophil counts in both breeds. In the Bonsmara breed, tick count was positively correlated with mononuclear cell counts. Cellular responses to adult R. microplus infestations were different and correlated with differences in tick resistance in Nguni and Bonsmara cattle breeds. It is essential to further characterise the molecular composition of the inflammatory infiltrate elicited by adult R. microplus infestation to fully comprehend immunity to ticks in cattle.     

Probing tRNA interaction with biogenic polyamines

Biogenic polyamines are found to modulate protein synthesis at different levels. This effect may be explained by the ability of polyamines to bind and influence the secondary structure of tRNA, mRNA, and rRNA. We report the interaction between tRNA and the three biogenic polyamines putrescine, spermidine, spermine, and cobalt(III)hexamine at physiological conditions, using FTIR spectroscopy, capillary electrophoresis, and molecular modeling. The results indicated that tRNA was stabilized at low biogenic polyamine concentration, as a consequence of polyamine interaction with the backbone phosphate group. The main tRNA reactive sites for biogenic polyamine at low concentration were guanine-N7/O6, uracil-O2/O4, adenine-N3, and 2′OH of the ribose. At high polyamine concentration, the interaction involves guanine-N7/O6, adenine-N7, uracil-O2 reactive sites, and the backbone phosphate group. The participation of the polycation primary amino group, in the interaction and the presence of the hydrophobic contact, are also shown. The binding affinity of biogenic polyamine to tRNA molecule was in the order of spermine > spermidine > putrescine with K_Spm = 8.7 × 10⁵ M⁻¹, K_Spd = 6.1 × 10⁵ M⁻¹, and K_Put = 1.0 × 10⁵ M⁻¹, which correlates with their positively charged amino group content. Hill analysis showed positive cooperativity for the biogenic polyamines and negative cooperativity for cobalt-hexamine. Cobalt(III)hexamine contains high- and low-affinity sites in tRNA with K₁ = 3.2 × 10⁵ M⁻¹ and K₂ = 1.7 × 10⁵ M⁻¹, that have been attributed to the interactions with guanine-N7 sites and the backbone PO₂ group, respectively. This mechanism of tRNA binding could explain the condensation phenomenon observed at high Co(III) content, as previously shown in the Co(III)–DNA complexes.     

The effect of nutrients on carbon and nitrogen fixation by the UCYN-A–haptophyte symbiosis

Symbiotic relationships between phytoplankton and N₂-fixing microorganisms play a crucial role in marine ecosystems. The abundant and widespread unicellular cyanobacteria group A (UCYN-A) has recently been found to live symbiotically with a haptophyte. Here, we investigated the effect of nitrogen (N), phosphorus (P), iron (Fe) and Saharan dust additions on nitrogen (N₂) fixation and primary production by the UCYN-A–haptophyte association in the subtropical eastern North Atlantic Ocean using nifH expression analysis and stable isotope incubations combined with single-cell measurements. N₂ fixation by UCYN-A was stimulated by the addition of Fe and Saharan dust, although this was not reflected in the nifH expression. CO₂ fixation by the haptophyte was stimulated by the addition of ammonium nitrate as well as Fe and Saharan dust. Intriguingly, the single-cell analysis using nanometer scale secondary ion mass spectrometry indicates that the increased CO₂ fixation by the haptophyte in treatments without added fixed N is likely an indirect result of the positive effect of Fe and/or P on UCYN-A N₂ fixation and the transfer of N₂-derived N to the haptophyte. Our results reveal a direct linkage between the marine carbon and nitrogen cycles that is fuelled by the atmospheric deposition of dust. The comparison of single-cell rates suggests a tight coupling of nitrogen and carbon transfer that stays balanced even under changing nutrient regimes. However, it appears that the transfer of carbon from the haptophyte to UCYN-A requires a transfer of nitrogen from UCYN-A. This tight coupling indicates an obligate symbiosis of this globally important diazotrophic association.