Analysis of radicals and radical reaction products in cell signalling and biomolecular damage:the long hard road to gold-standard measures

The field of free radical biology and medicine continues to move at a tremendous pace, with a constant flow of ground-breaking discoveries. The following collection of papers in this issue of Biochemical Society Transactions highlights several key areas of topical interest, including the crucial role of validated measurements of radicals and reactive oxygen species in underpinning nearly all research in the field, the important advances being made as a result of the overlap of free radical research with the reinvigorated field of lipidomics (driven in part by innovations in MS-based analysis), the acceleration of new insights into the role of oxidative protein modifications (particularly to cysteine residues) in modulating cell signalling, and the effects of free radicals on the functions of mitochondria, extracellular matrix and the immune system. In the present article, we provide a brief overview of these research areas, but, throughout this discussion, it must be remembered that it is the availability of reliable analytical methodologies that will be a key factor in facilitating continuing developments in this exciting research area.

Matrix-assisted diffusion-ordered spectroscopy:mixture resolution by NMR using SDSmicelles

Diffusion-ordered spectroscopy (DOSY) is a powerful technique for mixture analysis, but in its basic form it cannot separate the component spectra for species with very similar diffusion coefficients. It has been recently demonstrated that the component spectra of a mixture of isomers with nearly identical diffusion coefficients (the three dihydroxybenzenes) can be resolved using matrix-assisted DOSY (MAD), in which diffusion is perturbed by the addition of a co-solute such as a surfactant [R. Evans, S. Haiber, M. Nilsson, G. A. Morris, Anal. Chem. 2009, 81, 4548-4550]. However, little is known about the conditions required for such a separation, for example, the concentrations and concentration ratios of surfactant and solutes. The aim of this study was to explore the concentration range over whichmatrix-assisted DOSY using the surfactant SDS can achieve diffusion resolution of a simple model set of isomers, the monomethoxyphenols. The results show that the separation is remarkably robust with respect to both the concentrations and the concentration ratios of surfactant and solutes, supporting the idea that MAD may become a valuable tool formixture analysis. © 2010 John Wiley & Sons, Ltd.

Copper(II) complexes with unsymmetrical pentadentate ed3a-type diamino-tricarboxylate ligands. Crystal structures, configurational analysis and DFT study of complexes

The O–O–N–N–O-type pentadentate ligands H3ed3a, H3pd3a and H3pd3p (H3ed3a stands ethylenediamine-N,N,N′-triacetic acid; H3pd3a stands 1,3-propanediamine-N,N,N′-triacetic acid and H3pd3p stands 1,3-propanediamine-N,N,N′-tri-3-propionic acid) and the corresponding novel octahedral or square-planar/trigonal-bipyramidal copper(II) complexes have been prepared and characterized. H3ed3a, H3pd3a and H3pd3p ligands coordinate to copper(II) ion via five donor atoms (three deprotonated carboxylate atoms and two amine nitrogens) affording octahedral in case of ed3a3− and intermediate square-pyramidal/trigonal-bipyramidal structure in case of pd3a3− and pd3p3−. A six coordinate, octahedral geometry has been established crystallographically for the [Mg(H2O)6][Cu(ed3a)(H2O)]2 · 2H2O complex and five coordinate square-pyramidal for the [Mg(H2O)5Cu(pd3a)][Cu(pd3a)] · 2H2O. Structural data correlating similar chelate Cu(II) complexes have been used for the better understanding the pathway: octahedral → square-pyramidal ↔ trigonal- bipyramid geometry. An extensive configuration analysis is discussed in relation to information obtained for similar complexes. The infra-red and electronic absorption spectra of the complexes are discussed in comparison with related complexes of known geometries. Molecular mechanics and density functional theory (DFT) programs have been used to model the most stable geometric isomer yielding, at the same time, significant structural data. The results from density functional studies have been compared with X-ray data.

Adipose atrophy in cancer cachexia:morphologic and molecular analysis of adipose tissue in tumour-bearing mice

Extensive loss of adipose tissue is a hallmark of cancer cachexia but the cellular and molecular basis remains unclear. This study has examined morphologic and molecular characteristics of white adipose tissue in mice bearing a cachexia-inducing tumour, MAC16. Adipose tissue from tumour-bearing mice contained shrunken adipocytes that were heterogeneous in size. Increased fibrosis was evident by strong collagen-fibril staining in the tissue matrix. Ultrastructure of 'slimmed' adipocytes revealed severe delipidation and modifications in cell membrane conformation. There were major reductions in mRNA levels of adipogenic transcription factors including CCAAT/enhancer binding protein alpha (C/EBPα), CCAAT/enhancer binding protein beta, peroxisome proliferator-activated receptor gamma, and sterol regulatory element binding protein-1c (SREBP-1c) in adipose tissue, which was accompanied by reduced protein content of C/EBPα and SREBP-1. mRNA levels of SREBP-1c targets, fatty acid synthase, acetyl CoA carboxylase, stearoyl CoA desaturase 1 and glycerol-3-phosphate acyl transferase, also fell as did glucose transporter-4 and leptin. In contrast, mRNA levels of peroxisome proliferators-activated receptor gamma coactivator-1alpha and uncoupling protein-2 were increased in white fat of tumour-bearing mice. These results suggest that the tumour-induced impairment in the formation and lipid storing capacity of adipose tissue occurs in mice with cancer cachexia. © 2006 Cancer Research UK.

Mechanical properties and the evolution of matrix molecules in PTFE upon irradiation with MeV alpha particles

The morphology, chemical composition, and mechanical properties in the surface region of α-irradiated polytetrafluoroethylene (PTFE) have been examined and compared to unirradiated specimens. Samples were irradiated with 5.5 MeV 4He2+ ions from a tandem accelerator to doses between 1 × 106 and 5 × 1010 Rad. Static time-of-flight secondary ion mass spectrometry (ToF-SIMS), using a 20 keV C60+ source, was employed to probe chemical changes as a function of a dose. Chemical images and high resolution spectra were collected and analyzed to reveal the effects of a particle radiation on the chemical structure. Residual gas analysis (RGA) was utilized to monitor the evolution of volatile species during vacuum irradiation of the samples. Scanning electron microscopy (SEM) was used to observe the morphological variation of samples with increasing a particle dose, and nanoindentation was engaged to determine the hardness and elastic modulus as a function of a dose. The data show that PTFE nominally retains its innate chemical structure and morphology at a doses <109 Rad. At α doses ≥109 Rad the polymer matrix experiences increased chemical degradation and morphological roughening which are accompanied by increased hardness and declining elasticity. At  α doses >1010 Rad the polymer matrix suffers severe chemical degradation and material loss. Chemical degradation is observed in ToF-SIMS by detection of ions that are indicative of fragmentation, unsaturation, and functionalization of molecules in the PTFE matrix. The mass spectra also expose the subtle trends of crosslinking within the α-irradiated polymer matrix. ToF-SIMS images support the assertion that chemical degradation is the result of a particle irradiation and show morphological roughening of the sample with increased a dose. High resolution SEM images more clearly illustrate the morphological roughening and the mass loss that accompanies high doses of a particles. RGA confirms the supposition that the outcome of chemical degradation in the PTFE matrix with continuing irradiation is evolution of volatile species resulting in morphological roughening and mass loss. Finally, we reveal and discuss relationships between chemical structure and mechanical properties such as hardness and elastic modulus.

Comparison of the ageing behaviour of PM 2124 Al alloy and Al-SiCp metal-matrix composite

The ageing response of 2124 Al-SiC particulate metal-matrix composite (MMC) and unreinforced alloy has been examined using hardness measurements and Arrhenius analysis. The formation of phases during precipitation has been studied using differential scanning calorimetry (DSC). The MMC exhibits accelerated ageing compared to unreinforced alloy, due to enhanced S′ formation. The activation energy for diffusion is lower in the MMC than in the unreinforced alloy. DSC scans show Guinier-Preston B (GPB) zone nucleation to occur at a lower temperature in the MMC, whilst the total volume of GPB zones formed is smaller than in the unreinforced alloy. A model has been proposed to explain the GPB zone formation behaviour, in which ease of GPB zone nucleation varies within the MMC, as a function of ageing time and of position within the matrix. S′ formation is enhanced in the MMC because of improved diffusion and a large increase in density of heterogeneous nucleation sites compared to the unreinforced alloy. © 1994 Chapman & Hall.