O-acetylation of sialic acids in N-glycans of Atlantic salmon (Salmo salar) serum is altered by handling stress

O-acetylation is one of the major modifications of sialic acids that significantly alters biological properties of the parent molecule. These O-acetylated forms are components of the cellular membrane and can affect physiological and pathological responses. Understanding the role of N-glycans in physiology is of increasing relevance to cellular biologists in various disciplines who study glycoproteomics yet lack information regarding the function of the attached glycans. It is well known that stress may decrease immune function in fish; however, there are only few suitable biomarkers available to monitor the physiological responses under the stress conditions. This study is the first report on the effect of stress on the profile of O-acetylation of sialic acids in fish serum. In order to preserve the relevant structural characteristics as much as possible, native N-glycans were directly analyzed using CE-MS. We have characterized the N-glycans in serum of salmon (Salmo salar) exposed to long-term handling stress (15 s out of the water, daily for 4 wk) and compared with the results obtained from sera of control fish. The results indicated that major N-glycans in salmon serum contained mono-acetylated sialic acids (83%), and that the O-acetylation pattern of sialic acids could be altered by long-term stress.

Zinc and docosahexaenoic acid metabolism in brain cells

This thesis examines the direct interaction of Docosahexaenoic acid (DHA, an omega-3 fatty acid) against zinc-induced mitochondrial dysfunction and involvement of bioenergetic regulation as a zinc toxicity target, which may be the initiator of oxidative stress, caspase cascade, alteration in epigenetic patterns and therefore gene expression in human neuronal cells.

Eicosapentaenoic acid and oxypurinol in the treatment of muscle wasting in a mouse model of cancer cachexia

Cancer cachexia is a wasting condition, driven by systemic inflammation and oxidative stress. This study investigated eicosapentaenoic acid (EPA) in combination with oxypurinol as a treatment in a mouse model of cancer cachexia. Mice with cancer cachexia were randomized into 4 treatment groups (EPA (0.4 g/kg/day), oxypurinol (1 mmol/L ad-lib), combination, or control), and euthanized after 29 days. Analysis of oxidative damage to DNA, mRNA analysis of pro-oxidant, antioxidant and proteolytic pathway components, along with enzyme activity of pro- and antioxidants were completed on gastrocnemius muscle. The control group displayed earlier onset of tumor compared to EPA and oxypurinol groups (P<0.001). The EPA group maintained body weight for an extended duration (20 days) compared to the oxypurinol (5 days) and combination (8 days) groups (P<0.05). EPA (18.2±3.2 pg/ml) and combination (18.4±3.7 pg/ml) groups had significantly higher 8-OH-dG levels than the control group (12.9±1.4 pg/ml, P≤0.05) indicating increased oxidative damage to DNA. mRNA levels of GPx1, MURF1 and MAFbx were higher following EPA treatment compared to control (P≤0.05). Whereas oxypurinol was associated with higher GPx1, MnSOD, CAT, XDH, MURF1, MAFbx and UbB mRNA compared to control (P≤0.05). Activity of total SOD was higher in the oxypurinol group (32.2±1.5 U/ml) compared to control (27.0±1.3 U/ml, P<0.01), GPx activity was lower in the EPA group (8.76±2.0 U/ml) compared to control (14.0±1.9 U/ml, P<0.05), and catalase activity was lower in the combination group (14.4±2.8 U/ml) compared to control (20.9±2.0 U/ml, P<0.01). There was no change in XO activity. The increased rate of weight decline in mice treated with oxypurinol indicates that XO may play a protective role during the progression of cancer cachexia, and its inhibition is detrimental to outcomes. In combination with EPA, there was little significant improvement from control, indicating oxypurinol is unlikely to be a viable treatment compound in cancer cachexia.

In depression, bacterial translocation may drive inflammatory responses, oxidative and nitrosative stress (O&NS) and autoimmune responses directed against O&NS- damaged neoepitopes

Objective: Depression is accompanied by activation of immuno-inflammatory and oxidative and nitrosative stress (IO&NS) pathways, and increased IgM/IgA responses to lipopolysaccharide (LPS) of gram-negative commensal bacteria. The latter suggests that bacterial translocation has caused IgM/IgA responses directed against LPS. Bacterial translocation may drive IO&NS responses.

Method: To examine the associations between IgM/IgA responses to LPS and IO&NS measurements, including plasma/serum interleukin-1 (IL-1), tumor necrosis factor (TNF)α, neopterin, lysozyme, oxidized LDL (oxLDL) antibodies, peroxides, and IgM (auto)immune responses against malondialdehyde (MDA), azelaic acid, phophatidyl inositol (Pi), NO-tryptophan and NO-tyrosine in depressed patients and controls.

Results: We found significant positive associations between IgM/IgA responses to LPS and oxLDL antibodies, IgM responses against MDA, azelaic acid, Pi, NO-tryptophan, and NO-tyrosine. The IgA responses to LPS were correlated with lysozyme. There were no significant positive correlations between the IgM/IgA responses to LPS and IL-1 and neopterin.

Conclusion: The findings show that in depression there is an association between increased bacterial translocation and lysozyme production, an antibacterial compound, O&NS processes, and autoimmune responses directed against O&NS generated neoantigenic determinants. It is suggested that bacterial translocation may drive IO&NS pathways in depression and thus play a role in its pathophysiology.

Green electrospun pantothenic acid/silk fibroin composite nanofibers: fabrication, characterization and biological activity

Silk fibroin (SF) from Bombyx mori has many established excellent properties and has found various applications in the biomedical field. However, some abilities or capacities of SF still need improving to meet the need for using practically. Indeed, diverse SF-based composite biomaterials have been developed. Here we report the feasibility of fabricating pantothenic acid (vitamin B5, VB5)-reinforcing SF nanofibrous matrices for biomedical applications through green electrospinning. Results demonstrated the successful loading of D-pantothenic acid hemicalcium salt (VB5-hs) into resulting composite nanofibers. The introduction of VB5-hs did not alter the smooth ribbon-like morphology and the silk I structure of SF, but significantly decreased the mean width of SF fibers. SF conformation transformed into β-sheet from random coil when composite nanofibrous matrices were exposed to 75% (v/v) ethanol vapor. Furthermore, nanofibers still remained good morphology after being soaked in water environment for five days. Interestingly, as-prepared composite nanofibrous matrices supported a higher level of cell viability, especially in a long culture period and significantly assisted skin cells to survive under oxidative stress compared with pure SF nanofibrous matrices. These findings provide a basis for further extending the application of SF in the biomedical field, especially in the personal skin-care field.

Suberoylanilide hydroxamic acid radiosensitizes tumor hypoxic cells in vitro through the oxidation of nitroxyl to nitric oxide

The pharmacological effects of hydroxamic acids are partially attributed to their ability to serve as HNO and/or NO donors under oxidative stress. Previously, it was concluded that oxidation of the histone deacetylase inhibitor suberoylanilide hydroxamic acid (SAHA) by the metmyoglobin/H2O2 reaction system releases NO, which was based on spin trapping of NO and accumulation of nitrite. Reinvestigation of this system demonstrates the accumulation of N2O, which is a marker of HNO formation, at similar rates under normoxia and anoxia. In addition, the yields of nitrite that accumulated in the absence and the presence of O2 did not differ, implying that the source of nitrite is other than autoxidation of NO. In this system metmyoglobin is instantaneously and continuously converted into compound II, leading to one-electron oxidation of SAHA to its respective transient nitroxide radical. Studies using pulse radiolysis show that one-electron oxidation of SAHA (pKa=9.56 ± 0.04) yields the respective nitroxide radical (pKa=9.1 ± 0.2), which under all experimental conditions decomposes bimolecularly to yield HNO. The proposed mechanism suggests that compound I oxidizes SAHA to the respective nitroxide radical, which decomposes bimolecularly in competition with its oxidation by compound II to form HNO. Compound II also oxidizes HNO to NO and NO to nitrite. Given that NO, but not HNO, is an efficient hypoxic cell radiosensitizer, we hypothesized that under an oxidizing environment SAHA might act as a NO donor and radiosensitize hypoxic cells. Preincubation of A549 and HT29 cells with 2.5 μM SAHA for 24h resulted in a sensitizer enhancement ratio at 0.01 survival levels (SER0.01) of 1.33 and 1.59, respectively. Preincubation of A549 cells with oxidized SAHA had hardly any effect and, with 2mM valproic acid, which lacks the hydroxamate group, resulted in SER0.01=1.17. Preincubation of HT29 cells with SAHA and Tempol, which readily oxidizes HNO to NO, enhanced the radiosensitizing effect of SAHA. Pretreatment with SAHA blocked A549 cells at the G1 stage of the cell cycle and upregulated γ-H2AX after irradiation. Overall, we conclude that SAHA enhances tumor radioresponse by multiple mechanisms that might also involve its ability to serve as a NO donor under oxidizing environments.

Nucleic acid-based aptamers: applications, development and clinical trials

Short single-stranded oligonucleotides called aptamers, often termed as chemical antibodies, have been developed as powerful alternatives to traditional antibodies with respect to their obvious advantages like high specificity and affinity, longer shelf-life, easier manufacturing protocol, freedom to introduce chemical modifications for further improvement, etc. Reiterative selection process of aptamers over 10-15 cycles starting from a large initial pool of random nucleotide sequences renders them with high binding affinity, thereby making them extremely specific for their targets. Aptamer-based detection systems are well investigated and likely to displace primitive detection systems. Aptamer chimeras (combination of aptamers with another aptamer or biomacromolecule or chemical moiety) have the potential activity of both the parent molecules, and thus hold the capability to perform diverse functions at the same time. Owing to their extremely high specificity and lack of immunogenicity or pathogenicity, a number of other aptamers have recently entered clinical trials and have garnered favorable attention from pharmaceutical companies. Promising results from the clinical trials provide new hope to change the conventional style of therapy. Aptamers have attained high therapeutic relevance in a short time as compared to synthetic drugs and/or other modes of therapy. This review follows the various trends in aptamer technology including production, selection, modifications and success in clinical fields. It focusses largely on the various applications of aptamers which mainly depend upon their selection procedures. The review also sheds light on various modifications and chimerizations that have been implemented in order to improve the stability and functioning of the aptamers, including introduction of locked nucleic acids (LNAs). The application of various aptamers in detection systems has been discussed elaborately in order to stress on their role as efficient diagnostic agents. The key aspect of this review is focused on success of aptamers on the basis of their performance in clinical trials for various diseases.

Alzheimer's Abeta fused to green fluorescent protein induces growth stress and a heat shock response

The 42 amino acid Alzheimer's Aβ peptide is involved in the progression of Alzheimer's disease. Here we describe the effects of intracellular Aβ, produced through its attachment to either end of a green fluorescent protein, in yeast. Cells producing Aβ exhibited a lower growth yield and a heat shock response, showing that Aβ fusions promote stress in cells and supporting the notion that intracellular Aβ is a toxic molecule. These studies have relevance in understanding the role of Aβ in the death of neuronal cells, and indicate that yeast may be a new tractable model system for the screening for inhibitors of the stress caused by Aβ.

The deleterious effects of oxidative and nitrosative stress on palmitoylation, membrane lipid rafts and lipid-based cellular signalling : new drug targets in neuroimmune disorders

Oxidative and nitrosative stress (O&NS) is causatively implicated in the pathogenesis of Alzheimer’s and Parkinson’s disease, multiple sclerosis, chronic fatigue syndrome, schizophrenia and depression. Many of the consequences stemming from O&NS, including damage to proteins, lipids and DNA, are well known, whereas the effects of O&NS on lipoprotein-based cellular signalling involving palmitoylation and plasma membrane lipid rafts are less well documented. The aim of this narrative review is to discuss the mechanisms involved in lipid-based signalling, including palmitoylation, membrane/lipid raft (MLR) and n-3 polyunsaturated fatty acid (PUFA) functions, the effects of O&NS processes on these processes and their role in the abovementioned diseases. S-palmitoylation is a post-translational modification, which regulates protein trafficking and association with the plasma membrane, protein subcellular location and functions. Palmitoylation and MRLs play a key role in neuronal functions, including glutamatergic neurotransmission, and immune-inflammatory responses. Palmitoylation, MLRs and n-3 PUFAs are vulnerable to the corruptive effects of O&NS. Chronic O&NS inhibits palmitoylation and causes profound changes in lipid membrane composition, e.g. n-3 PUFA depletion, increased membrane permeability and reduced fluidity, which together lead to disorders in intracellular signal transduction, receptor dysfunction and increased neurotoxicity. Disruption of lipid-based signalling is a source of the neuroimmune disorders involved in the pathophysiology of the abovementioned diseases. n-3 PUFA supplementation is a rational therapeutic approach targeting disruptions in lipid-based signalling.

Stress and reproduction: central mechanisms and sex differences in non-rodent species

Despite extensive research, the mechanisms by which stress affects reproduction are unknown. Activation of stress systems could potentially influence reproduction at any level of the hypothalamo-pituitary gonadal axis. Nonetheless, the predominant impact is on the secretion of gonadotrophin releasing hormone (GnRH) from the brain and the secretion of the gonadotrophins, luteinizing hormone (LH) and follicle stimulating hormone (FSH), from the gonadotrophs of the anterior pituitary gland. When stress is prolonged, it is likely that secretion of the gonadotrophins will be suppressed but the effects of acute stress or repeated acute stress are not clear. Different stressors activate different pathways for varying durations, and the actions of stress vary with sex and are influenced by the predominance of particular sex steroids in the circulation. The mechanisms by which stress influences reproduction are likely to involve complex interactions between a number of central and peripheral pathways and may be different in males and females. To understand these mechanisms, it is important to determine the stress pathways that are activated by particular stressors and to establish how these pathways affect the secretion and actions of GnRH. Furthermore, there is a need to know how stress influences the feedback actions of gonadal steroids and inhibin.

Nanostructured liquid crystalline particle assisted delivery of 2,4-dichlorophenoxyacetic acid to weeds, crops and model plants

Routine agricultural practices are heavily dependent on the use of surfactants, many of which are toxic to humans and detrimental to the environment. In proof of concept work we have previously shown the potential of nanostructured liquid crystalline particles (NLCP) to safely interact with plant leaf cuticular surfaces with minimal impact on epicuticular waxes. Here we demonstrate the use of NLCP to effectively deliver the auxin herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) to plant leaves in laboratory and field studies. In the laboratory, the physiological stress responses of lupin, Lupinus angustifolius (L.) (Fabaceae) towards NLCP spray applications were shown to be much reduced in comparison with application of two common surfactants. Phytotoxicity assays of 2,4-D loaded NLCP were used to validate the herbicidal effects on Arabidopsis thaliana (L.) Heynth. (Brassicaceae) and established a similarity with that of surfactant assisted 2,4-D delivery when tested at a concentration of 0.1%. Field trials were conducted to test the efficacy of NLCP-assisted delivery of 2,4-D in comparison with commercial surfactants for the control of the invasive weed wild radish, Raphanus raphanistrum (L.) (Brassicaceae), in wheat, Triticum aestivum (L.) (Poaceae) crop fields. Compared against Estercide 800, a commercially available 2,4-D formulation, NLCP assisted delivery of 2,4-D was effective at low concentrations of 0.03% and 0.06%. The crop yield remained similar for all the tested concentrations and formulations of 2,4-D loaded NLCP and Estercide 800. This is the first report to directly show that, as an alternative to conventional methods, NLCP can be used under both laboratory and field conditions to successfully delivery an agrochemical.

Arachidonic acid and eicosapentaenoic acid metabolism in juvenile Atlantic Salmon as affected by water temperature

Salmons raised in aquaculture farms around the world are increasingly subjected to sub-optimal environmental conditions, such as high water temperatures during summer seasons. Aerobic scope increases and lipid metabolism changes are known plasticity responses of fish for a better acclimation to high water temperature. The present study aimed at investigating the effect of high water temperature on the regulation of fatty acid metabolism in juvenile Atlantic salmon fed different dietary ARA/EPA ratios (arachidonic acid, 20:4n-6/ eicosapentaenoic acid, 20:5n-3), with particular focus on apparent in vivo enzyme activities and gene expression of lipid metabolism pathways. Three experimental diets were formulated to be identical, except for the ratio EPA/ARA, and fed to triplicate groups of Atlantic salmon (Salmo salar) kept either at 10°C or 20°C. Results showed that fatty acid metabolic utilisation, and likely also their dietary requirements for optimal performance, can be affected by changes in their relative levels and by environmental temperature in Atlantic salmon. Thus, the increase in temperature, independently from dietary treatment, had a significant effect on the β-oxidation of a fatty acid including EPA, as observed by the apparent in vivo enzyme activity and mRNA expression of pparα -transcription factor in lipid metabolism, including β-oxidation genes- and cpt1 -key enzyme responsible for the movement of LC-PUFA from the cytosol into the mitochondria for β-oxidation-, were both increased at the higher water temperature. An interesting interaction was observed in the transcription and in vivo enzyme activity of Δ5fad-time-limiting enzyme in the biosynthesis pathway of EPA and ARA. Such, at lower temperature, the highest mRNA expression and enzyme activity was recorded in fish with limited supply of dietary EPA, whereas at higher temperature these were recorded in fish with limited ARA supply. In consideration that fish at higher water temperature recorded a significantly increased feed intake, these results clearly suggested that at high, sub-optimal water temperature, fish metabolism attempted to increment its overall ARA status -the most bioactive LC-PUFA participating in the inflammatory response- by modulating the metabolic fate of dietary ARA (expressed as % of net intake), reducing its β-oxidation and favouring synthesis and deposition. This correlates also with results from other recent studies showing that both immune- and stress- responses in fish are up regulated in fish held at high temperatures. This is a novel and fundamental information that warrants industry and scientific attention, in consideration of the imminent increase in water temperatures, continuous expansion of aquaculture operations, resources utilisation in aquafeed and much needed seasonal/adaptive nutritional strategies.

Hydroxyoctadecadienoic acids: Oxidised derivatives of linoleic acid and their role in inflammation associated with metabolic syndrome and cancer

Linoleic acid (LA) is a major constituent of low-density lipoproteins. An essential fatty acid, LA is a polyunsaturated fatty acid, which is oxidised by endogenous enzymes and reactive oxygen species in the circulation. Increased levels of low-density lipoproteins coupled with oxidative stress and lack of antioxidants drive the oxidative processes. This results in synthesis of a range of oxidised derivatives, which play a vital role in regulation of inflammatory processes. The derivatives of LA include, hydroxyoctadecadienoic acids, oxo-​octadecadienoic acids, epoxy octadecadecenoic acid and epoxy-keto-octadecenoic acids. In this review, we examine the role of LA derivatives and their actions on regulation of inflammation relevant to metabolic processes associated with atherogenesis and cancer. The processes affected by LA derivatives include, alteration of airway smooth muscles and vascular wall, affecting sensitivity to pain, and regulating endogenous steroid hormones associated with metabolic syndrome. LA derivatives alter cell adhesion molecules, this initial step, is pivotal in regulating inflammatory processes involving transcription factor peroxisome proliferator-activated receptor pathways, thus, leading to alteration of metabolic processes. The derivatives are known to elicit pleiotropic effects that are either beneficial or detrimental in nature hence making it difficult to determine the exact role of these derivatives in the progress of an assumed target disorder. The key may lie in understanding the role of these derivatives at various stages of development of a disorder. Novel pharmacological approaches in altering the synthesis or introduction of synthesised LA derivatives could possibly help drive processes that could regulate inflammation in a beneficial manner. Chemical Compounds: Linoleic acid (PubChem CID: 5280450), 9- hydroxyoctadecadienoic acid (PubChem CID: 5312830), 13- hydroxyoctadecadienoic acid (PubChem CID: 6443013), 9-oxo-​octadecadienoic acid (PubChem CID: 3083831), 13-oxo-​octadecadienoic acid (PubChem CID: 4163990), 9,10-epoxy-12-octadecenoate (PubChem CID: 5283018), 12,13-epoxy-9-keto-10- trans -octadecenoic acid (PubChem CID: 53394018), Pioglitazone (PubChem CID: 4829).

Effect of bulk microstructure of commercially pure titanium on surface characteristics and fatigue properties after surface modification by sand blasting and acid-etching

Surface modification techniques are widely used to enhance the biological response to the implant materials. These techniques generally create a roughened surface, effectively increasing the surface area thus promoting cell adhesion. However, a negative side effect is a higher susceptibility of a roughened surface to failure due to the presence of multiple stress concentrators. The purpose of the study reported here was to examine the effects of surface modification by sand blasting and acid-etching (SLA) on the microstructure and fatigue performance of coarse-grained and ultrafine-grained (UFG) commercially pure titanium. Finer grain sizes, produced by equal channel angular pressing, resulted in lower values of surface roughness in SLA-processed material. This effect was associated with greater resistance of the UFG structure to plastic deformation. The fatigue properties of UFG Ti were found to be superior to those of coarse-grained Ti and conventional Ti-6Al-4V, both before and after SLA-treatment.