Polyunsaturated fatty acid oxidation in Alzheimer’s disease.

Alzheimer’s disease is a neurodegenerative disorder which has been characterised with genetic (apolipoproteins), protein (ß-amyloid and tau) and lipid oxidation/metabolism alterations in its pathogenesis. In conjunction with the Dementia Research Group, Bristol University, investigation into genetic, protein and lipid oxidation in Alzheimer’s disease was conducted. A large sample cohort using the double-blind criteria, along with various clinical and chemical data sets were used to improve the statistical analysis and therefore the strength of this particular study. Bristol University completed genetic and protein analysis with lipid oxidation assays performed at Aston University. Lipid oxidation is a complex process that creates various biomarkers, from transient intermediates, to short carbon chain products and cyclic ring structures. Quantification of these products was performed on lipid extracts of donated clinical diseased and non-diseased frontal and temporal brain regions, from the Brain Bank within Frenchay Hospital. The initial unoxidised fatty acids, first transient oxidation intermediates the conjugated dienes and lipid hydroperoxides, the endpoint aldehyde biomarkers and finally the cyclic isoprostanes and neuroprostanes were determined to investigate lipid oxidation in Alzheimer’s. Antioxidant levels were also investigated to observe the effect of oxidation on the defence pathways. Assays utilised in this analysis included; fatty acid composition by GC-FID, conjugated diene levels by HPLC-UV and UV-spec, lipid hydroperoxide levels by FOX, aldehyde content by TBARs, antioxidant status by TEAC and finally isoprostane and neuroprostane quantification using a newly developed EI-MS method. This method involved the SIM of specific ions from F-ring isoprostane and neuroprostane fragmentation, which enabled EI-MS to be used for their quantification. Analyses demonstrated that there was no significant difference between control and Alzheimer samples across all the oxidation biomarkers for both brain regions. Antioxidants were the only marker that showed a clear variance; with Alzheimer samples having higher levels than the age matched controls. This unique finding is supported with the observed lower levels of lipid oxidation biomarkers in Alzheimer brain region samples. The increased antioxidant levels indicate protection against oxidation which may be a host response to counteract the oxidative pathways, but this requires further investigation. In terms of lipid oxidation, no definitive markers or target site for therapeutic intervention have been revealed. This study concludes that dietary supplementation of omega-3 fatty acids or antioxidants would most likely be ineffective against Alzheimer disease, although it may support improvement in other areas of general health.

Biomarkers

Introduction – Why do we need ‘biomarkers? Biomarkers of protein oxidation Introduction Major issues/questions Protein carbonyl biomarkers Biochemistry Methods of measurement Storage, stability and limitations in use Protein thiol biomarkers Biochemistry Methods of measurement Storage, stability and limitations on use Aliphatic amino acid biomarkers Biochemistry Methods of measurement Storage, stability and limitations on use Oxidised Tryptophan Biomarkers Biochemistry Method of measurement Storage, stability and limitations on use Oxidised tyrosine biomarkers Biochemistry Methods of measurement Storage, stability and limitations on use Formation of neoepitopes on oxidised proteins Validation of assays for protein oxidation biomarkers Relationship of protein oxidation to disease Modulation of protein oxidation biomarkers by antioxidants Future perspectives Introduction to lipid peroxidation biomarkers Introduction: biochemistry of lipid peroxidation Malondialdehyde Methods of measurement Storage, stability and limitations on use Conjugated dienes Method of measurement Storage, stability and limitations of use LDL lag phase Method of measurement Storage, stability and limitations of use Hydrocarbon gases Biochemistry Method of measurement Storage, stability and limitations on use Lipofuscin Biochemistry Method of measurement Storage, stability and limitation on use Lipid peroxides Biochemistry Method of measurement Storage, stability and limitations on use Isoprostanes Biochemistry Method of measurement Storage, stability and limitations on use Possible new biomarkers of lipid oxidation Relationship of lipid peroxidation to disease Modulation of lipid peroxidation biomarkers by antioxidants Functional consequences of lipid peroxidation Contribution of dietary intake to lipid peroxidation products Biomarkers of DNA oxidation Introduction Confounding factors Units and terminology Nuclear and mitochondrial DNA damage Lymphocytes as surrogate tissues Measurement of DNA damage with the comet assay Practical details Storage, stability, and limitations of the assay Measurement of DNA base oxidation by HPLC Practical details Storage, stability and limitations of the method Measurement of DNA base oxidation by GC–MS Biochemistry of 8-oxoguanine, adenine and fapy derivatives Methods of measurement Storage, stability and limitations of the method Analysis of guanine oxidation products in urine Method of measurement Limitations and criticisms Immunochemical methods Methods of measurement Storage, stability, and limitations of the assay 32P post-labelling Method of measurement Limitations and criticisms Validation of assays for DNA oxidation Oxo-dGuo in lymphocyte DNA Urinary measurements DNA–aldehyde adducts Biochemistry Method of measurement Products of reactive nitrogen species Endpoints arising from oxidative DNA damage Mutations Chromosome aberrations Micronuclei Site-specific DNA damage Relationship of DNA oxidation to disease Modulation of DNA oxidation biomarkers by antioxidants Direct and indirect effects of oxidative stress: measures of total oxidant/antioxidant levels Visualisation of cellular oxidants Biochemistry: histochemical detection of ROS Method of measurement Limitations, storage and stability Measurement of hydrogen peroxide Biochemistry Methods of measurement Storage, stability and limitations on use Measurement of the ratio of antioxidant/oxidised antioxidant Biochemistry Method of measurement Storage, stability and limitations on use Total antioxidant capacity Biochemistry Terminology Methods of measurement Storage, stability and limitations on use Validation of assays for direct oxidant and antioxidant biomarkers Relationship of oxidant/antioxidant measurement to disease Modulation of oxidant/antioxidant biomarkers by dietary antioxidants Induction of genes in response to oxidative stress Background Measurement of antioxidant responsive genes and proteins Effects of antioxidant intake on the activity of antioxidant enzymes

Characterization of FITC-conjugated lectin binding to Candida albicans

Avidity of yeast and hyphal forms of Candida albicans for FITC-conjugated lectins was determined by flow cytometry and digital microscopy. Yeast phase cells bound Con A, a lectin with marked affinity for mannose, irrespective of growth phase, yet demonstrated little avidity for WGA and SBA. Yeast phase cell avidity for mannose-specific lectins was characterized through determination of FITC-conjugated Con A, LcH, PSA and GNA binding and subsequent calculation of Bmax, EC50 and Hn values. Such an approach, through comparison among FITC-conjugated lectins of differing specific activities, furnishes further insight into exposed outer cell wall mannose moieties. The rank order of lectin affinity as defined by EC50 values was GNA > Con A > LcH > PSA. Values for Hn suggest that lectins predominantly bind to a single receptor class, the relative abundance of which as defined by Bmax values was PSA > GNA > Con A > LcH. Hyphal surfaces in common with yeast phase cells demonstrated marked avidity for FITC-Con A, however, fluorescence of Candida morphological forms differed significantly, indicative of varying outer cell wall mannose exposure.

In vivo effects of tailored laminin-332 α3 conjugated scaffolds enhances wound healing:a histomorphometric analysis

Surface modification techniques have been used to develop biomimetic scaffolds by incorporating cell adhesion peptides. In our previous work, we have shown the tethering of laminin-332 α3 chain to type I collagen scaffold using microbial transglutaminase (mTGase), promotes cell adhesion, migration, and proliferation. In this study, we evaluated the wound healing properties of tailored laminin-332 α3 chain (peptide A: PPFLMLLKGSTR) tethered to a type I collagen scaffold using mTGase by incorporating transglutaminase substrate peptide sequences containing either glutamine (peptide B: PPFLMLLKGSTREAQQIVM) or lysine (peptide C: PPFLMLLKGSTRKKKKG) in rat full-thickness wound model at two different time points (7 and 21 days). Histological evaluations were assessed for wound closure, epithelialization, angiogenesis, inflammatory, fibroblastic cellular infiltrations, and quantified using stereological methods (p < 0.05). Peptide A and B tethered to collagen scaffold using mTGase stimulated neovascularization, decreased the inflammatory cell infiltration and prominently enhanced the fibroblast proliferation which significantly accelerated the wound healing process. We conclude that surface modification by incorporating motif of laminin-332 α3 chain (peptide A: PPFLMLLK GSTR) domain and transglutaminase substrate to the laminin-332 α3 chain (peptide B: PPFLMLLKGSTREAQQIVM) using mTGase may be a potential candidate for tissue engineering applications and skin regeneration. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 101A:2788-2795, 2013. Copyright © 2013 Wiley Periodicals, Inc., a Wiley Company.

Phase-conjugated pilots for fibre nonlinearity compensation in CO-OFDM transmission

We experimentally demonstrate a novel fibre nonlinearity compensation technique for CO-OFDM based on phase-conjugated pilots (PCPs), showing that, by varying the PCP overhead a performance improvement up to 4 dB can be achieved allowing highly flexible adaptation to link characteristics.

Phase-conjugated subcarrier coding for fibre nonlinearity mitigation in CO-OFDM transmission

We demonstrate a novel subcarrier coding scheme combined with pre-EDC for fibre nonlinearity mitigation in CO-OFDM, showing that a performance improvement of 1.5 dB can be achieved in a 150 Gb/s BPSK PDM CO-OFDM transmission.

Demonstration of phase-conjugated subcarrier coding for fiber nonlinearity compensation in CO-OFDM transmission

In this paper, we demonstrate through computer simulation and experiment a novel subcarrier coding scheme combined with pre-electrical dispersion compensation (pre-EDC) for fiber nonlinearity mitigation in coherent optical orthogonal frequency division multiplexing (CO-OFDM) systems. As the frequency spacing in CO-OFDM systems is usually small (tens of MHz), neighbouring subcarriers tend to experience correlated nonlinear distortions after propagation over a fiber link. As a consequence, nonlinearity mitigation can be achieved by encoding and processing neighbouring OFDM subcarriers simultaneously. Herein, we propose to adopt the concept of dual phase conjugated twin wave for CO-OFDM transmission. Simulation and experimental results show that this simple technique combined with 50% pre-EDC can effectively offer up to 1.5 and 0.8 dB performance gains in CO-OFDM systems with BPSK and QPSK modulation formats, respectively.

Phase-conjugated pilots for fibre nonlinearity compensation in CO-OFDM transmission

In this paper, we demonstrate a novel fiber nonlinearity compensation technique for coherent optical orthogonal frequency-division multiplexing (CO-OFDM) systems based on the transmission of phase-conjugated pilots (PCPs). In this scheme, a portion of OFDM subcarriers (up to 50%) is transmitted with its phase conjugates, which is used at the receiver to estimate the nonlinear distortions in the respective subcarriers and other subcarriers, which are not accompanied by PCPs. Simulation and experimental results show that by varying the PCP overhead, a performance improvement up to 4 dB can be achieved. In addition, the proposed technique can be effectively applied in both single polarization and polarization-division multiplexed systems, in both single channel and wavelength-division multiplexing systems, thus, offering highest flexibility in implementations.