The development of engineered nanomaterials is growing exponentially, despite concerns over their potential similarities to environmental nanoparticles that are associated with significant cardiorespiratory morbidity and mortality. The mechanisms through which inhalation of nanoparticles could trigger acute cardiovascular events are emerging, but a fundamental unanswered question remains: Do inhaled nanoparticles translocate from the lung in man and directly contribute to the pathogenesis of cardiovascular disease? In complementary clinical and experimental studies, we used gold nanoparticles to evaluate particle translocation, permitting detection by high-resolution inductively coupled mass spectrometry and Raman microscopy. Healthy volunteers were exposed to nanoparticles by acute inhalation, followed by repeated sampling of blood and urine. Gold was detected in the blood and urine within 15 min to 24 h after exposure, and was still present 3 months after exposure. Levels were greater following inhalation of 5 nm (primary diameter) particles compared to 30 nm particles. Studies in mice demonstrated the accumulation in the blood and liver following pulmonary exposure to a broader size range of gold nanoparticles (2–200 nm primary diameter), with translocation markedly greater for particles <10 nm diameter. Gold nanoparticles preferentially accumulated in inflammation-rich vascular lesions of fat-fed apolipoproteinE-deficient mice. Furthermore, following inhalation, gold particles could be detected in surgical specimens of carotid artery disease from patients at risk of stroke. Translocation of inhaled nanoparticles into the systemic circulation and accumulation at sites of vascular inflammation provides a direct mechanism that can explain the link between environmental nanoparticles and cardiovascular disease and has major implications for risk management in the use of engineered nanomaterials.

Nanoporous, thick (8 μm) films of titania (TiO2) were prepared and used for the immobilization of proteins. A detailed study has been made into the factors influencing protein adsorption on TiO2. Among these, we investigated pH, ionic strength of solution, protein surface charge, protein size, and immobilization time. Protein immobilization is found to be remarkably stable, attributed to secondary binding processes occurring after the initial immobilization. We also investigated the electrochemical properties of these films using cyclic voltammetry and spectroelectrochemistry and found that not only was direct reduction of the FeIII−heme to FeII−heme of both cytochrome-c and hemoglobin possible but that all the protein in the film is electroactive. We further demonstrate the use of a hemoglobin/TiO2 film as an aerobic sensor for nitric oxide. Optical sensing is demonstrated, with a limit of detection of 1 μM.

The field of regenerative medicine spans a wide area of the biomedical landscape—from single cell culture in laboratories to human whole-organ transplantation. To ensure that research is transferrable from bench to bedside, it is critical that we are able to assess regenerative processes in cells, tissues, organs and patients at a biochemical level. Regeneration relies on a large number of biological factors, which can be perturbed using conventional bioanalytical techniques. A versatile, non-invasive, non-destructive technique for biochemical analysis would be invaluable for the study of regeneration; and Raman spectroscopy is a potential solution. Raman spectroscopy is an analytical method by which chemical data are obtained through the inelastic scattering of light. Since its discovery in the 1920s, physicists and chemists have used Raman scattering to investigate the chemical composition of a vast range of both liquid and solid materials. However, only in the last two decades has this form of spectroscopy been employed in biomedical research. Particularly relevant to regenerative medicine are recent studies illustrating its ability to characterise and discriminate between healthy and disease states in cells, tissue biopsies and in patients. This review will briefly outline the principles behind Raman spectroscopy and its variants, describe key examples of its applications to biomedicine, and consider areas of regenerative medicine that would benefit from this non-invasive bioanalytical tool.

Marine photosynthetic picoeukaryotes (PPEs), representing organisms < 3 µm in size, are major contributors to global carbon cycling. However, the key members of the PPE community and hence the major routes of carbon fixation, particularly in the open ocean environment, are poorly described. Here, we have accessed PPE community structure using the plastid encoded 16S rRNA gene. Plastid 16S rRNA genes were sequenced from 65 algal cultures, about half being PPEs, representing 14 algal classes. These included sequences from 5 classes where previously no such sequences from cultured representatives had been available (Bolidophyceae, Dictyochophyceae, Eustigmatophyceae, Pelagophyceae and Pinguiophyceae). Sequences were also obtained for 6 of the 7 (according to 18S rRNA gene sequence) prasinophyte clades. Phylogenetic analysis revealed plastids from the same class as clustering together. Using all the obtained sequences, as well as plastid sequences currently in public databases, a non-degenerate marine algal plastid-biased PCR primer, PLA491F, was developed to minimize amplification of picocyanobacteria, which often dominate numerically environmental samples. Clone libraries subsequently constructed from the pico-sized fraction from 2 open ocean sites in the Arabian Sea, revealed an abundance of 16S rRNA gene clones phylogenetically related to chrysophytes, whilst prymnesiophyte, clade II prasinophyte (Ostreococcus-like) and pelagophyte clones were also well represented. The finding of a wealth of novel clones related to the Chrysophyceae highlights the utility of a PCR biased towards marine algal plastids as a valuable complement to 18S rDNA based studies of PPE diversity. KEY WORDS: Photosynthetic picoeukaryotes · Plastid 16S rRNA · Arabian Sea · PCR Resale or republication not permitted without written consent of the publisherAquat Microb Ecol 43: [79][80][81][82][83][84][85][86][87][88][89][90][91][92][93] 2006 guiophyceae (Kawachi et al. 2002). Yet despite the evident ecological significance of PPEs, relatively little is known of their diversity in the marine environment, particularly in the open ocean. This has been attributed mainly to difficulties in identification by light microscopy. Only recently, with the advent of molecular techniques, has picoeukaryote diversity begun to be revealed (for a recent review see Moreira & LopezGarcia 2002). Thus, phylogenetic studies based on 18S rRNA gene sequence analysis are now beginning to show the extent of taxonomic diversity within this group of organisms (Diez et al. 2001, Lopez-Garcia et al. 2001, Moon-van der Staay et al. 2001, Stoeck & Epstein 2003. Such studies, using 'universal' 18S rRNA gene primers (which target both [photo]autotrophs and heterotrophs) and clone library construction from environmental samples have demonstrated the presence of PPEs affiliated with many different algal classes including the Bacillariophyceae, Bolidophyceae, Chrysophyceae, Cryptophyceae, Dictyochophyceae, Dinophyceae, Eustigmatophyceae, Glaucocystophyceae, Pelagoph...

The application of gold nanoshells (NS) as a surface-enhanced Raman (SER) platform for intracellular sensing in NIH-3T3 fibroblast cells was studied by using a near-infrared Raman system. To show the feasibility of using these 151 +/- 5 nm sized solution-stable nanoparticles inside living cells, we investigated the uptake, cellular response, and the health of the cell population. We show that NS are taken up voluntarily and can be found in the cytosol by transmission electron microscopy (TEM), which also provides detailed information about location and immediate surrounding of the NS. The internalization into cells has been found to be independent of active cellular mechanisms, such as endocytosis, and can be suggested to be of passive nature. Uptake of NS into cells can be controlled, and cells show no increase in necrosis or apoptosis as a result; we show that NS-based intracytosolic SER spectra can be measured on biological samples using short acquisition times and low laser powers. We demonstrate its application using 4-mercaptobenzoic acid (4-MBA)-functionalized nanoshells as a pH sensor.