T-1-MRI Fluorescent Iron Oxide Nanoparticles by Microwave Assisted Synthesis

Iron oxide nanoparticles have long been studied as a T-2 contrast agent in MRI due to their superparamagnetic behavior. T-1-based positive contrast, being much more favorable for clinical application due to brighter and more accurate signaling is, however, still limited to gadolinium- or manganese-based imaging tools. Though being the only available commercial positive-contrast agents, they lack an efficient argument when it comes to biological toxicity and their circulatory half-life in blood. The need arises to design a biocompatible contrast agent with a scope for easy surface functionalization for long circulation in blood and/or targeted imaging. We hereby propose an extremely fast microwave synthesis for fluorescein-labeled extremely-small iron oxide nanoparticles (fdIONP), in a single step, as a viable tool for cell labeling and T-1-MRI. We demonstrate the capabilities of such an approach through high-quality magnetic resonance angiographic images of mice.This study was supported by a grant from Ministerio de Economia y Competitividad (MAT2013-47303-P) and European Union (Cardionext, ITN-FP7-608027).S

Microwave-driven synthesis of bisphosphonate nanoparticles allows in vivo visualisation of atherosclerotic plaque

A fast and reproducible microwave-driven process has allowed us to synthesise neridronate-functionalised nanoparticles. Contrary to tradition, the phosphate groups decorate the outside layer of the particles providing Ca2+ binding properties in vitro and selective accumulation in vivo in the atheroma plaque. In vivo and ex vivo detection by T2-weighted MRI is demonstrated and validated by histology. The accumulation in the plaque takes place in less than one hour following the intravenous injection, which is particularly suitable for clinical applications

Superparamagnetic Nanoparticles for Atherosclerosis Imaging

The production of magnetic nanoparticles of utmost quality for biomedical imaging requires several steps, from the synthesis of highly crystalline magnetic cores to the attachment of the different molecules on the surface. This last step probably plays the key role in the production of clinically useful nanomaterials. The attachment of the different biomolecules should be performed in a defined and controlled fashion, avoiding the random adsorption of the components that could lead to undesirable byproducts and ill-characterized surface composition. In this work, we review the process of creating new magnetic nanomaterials for imaging, particularly for the detection of atherosclerotic plaque, in vivo. Our focus will be in the different biofunctionalization techniques that we and several other groups have recently developed. Magnetic nanomaterial functionalization should be performed by chemoselective techniques. This approach will facilitate the application of these nanomaterials in the clinic, not as an exception, but as any other pharmacological compound.The authors thank the Spanish Ministry of Science (SAF2011-25445), the Comunidad de Madrid (S2010/BMD-2326, Inmunothercan-CM, NANOCOPD-CIBERES-CIBERBBN-SEPAR), and the EU 7th Framework Program (FP7-PEOPLE-ITN-264864 Pinet and FP7-PEOPLE-2013-ITN CardioNext).S

Synergistic effects of UVR and simulated stratification on commensalistic phytoplankton–bacteria relationship in two optically contrasting oligotrophic Mediterranean lakes

An indirect effect of global warming is a reduction in the depth of the upper mixed layer (UML) causing organisms to be exposed to higher levels of ultraviolet (UVR, 280–400 nm) and photosynthetically active radiation (PAR, 400–700 nm). This can affect primary and bacterial production as well as the commensalistic phytoplankton–bacteria relationship. The combined effects of UVR and reduction in the depth of the UML were assessed on variables related to the metabolism of phytoplankton and bacteria, during in situ experiments performed with natural pico- and nanoplankton communities from two oligotrophic lakes with contrasting UVR transparency (high-UVR versus low-UVR waters) of southern Spain. The negative UVR effects on epilimnetic primary production (PP) and on heterotrophic bacterial production (HBP), intensified under increased stratification, were higher in the low-UVR than in the high-UVR lake, and stronger on the phytoplanktonic than on the heterotrophic bacterial communities. Under UVR and increased stratification, the commensalistic phytoplankton–bacteria relationship was strengthened in the high-UVR lake where excretion of organic carbon (EOC) rates exceeded the bacterial carbon demand (BCD; i.e., BCD : EOC(%) ratio 100). The greater UVR damage to phytoplankton and bacteria and the weakening of their commensalistic interaction found in the low-UVR lake indicates that these ecosystems would be especially vulnerable to UVR and increased stratification as stressors related to global climate change. Thus, our findings may have important implications for the carbon cycle in oligotrophic lakes of the Mediterranean region.This study was supported by the Ministerio Español de Medio Ambiente, Rural y Marino (PN2009/067), Ciencia e Innovación (CGL2011-23681), Junta de Andalucía (Excelencia CVI-02598 and P09-RNM-5376), Consejo Nacional de Investigaciones Científicas y Técnicas – CONICET (PIP no. 112-201001-00228), and Fundación Playa Unión. G. Herrera and C. Durán were supported by a Formación de Profesorado Universitario grant from the Spanish government. The authors are indebted to the staff of Sierra Nevada National Park and Lagunas de Ruidera Natural Park for permission to work, to E. Jiménez-Coll for the bacterial production analysis, and to D. Nesbitt for writing assistance in English

Interactive effects of vertical mixing, nutrients and ultraviolet radiation: in situ photosynthetic responses of phytoplankton from high mountain lakes in Southern Europe

Global change, together with human activities, has resulted in increasing amounts of organic material (including nutrients) that water bodies receive. This input further attenuates the penetration of solar radiation, leading to the view that opaque lakes are more "protected" from solar ultraviolet radiation (UVR) than clear ones. Vertical mixing, however, complicates this view as cells are exposed to fluctuating radiation regimes, for which the effects have, in general, been neglected. Furthermore, the combined impacts of mixing, together with those of UVR and nutrient inputs are virtually unknown. In this study, we carried out complex in situ experiments in three high mountain lakes of Spain (Lake Enol in the National Park Picos de Europa, Asturias, and lakes Las Yeguas and La Caldera in the National Park Sierra Nevada, Granada), used as model ecosystems to evaluate the joint impact of these climate change variables. The main goal of this study was to address the question of how short-term pulses of nutrient inputs, together with vertical mixing and increased UVR fluxes modify the photosynthetic responses of phytoplankton. The experimentation consisted in all possible combinations of the following treatments: (a) solar radiation: UVR + PAR (280–700 nm) versus PAR (photosynthetically active radiation) alone (400–700 nm); (b) nutrient addition (phosphorus (P) and nitrogen (N)): ambient versus addition (P to reach to a final concentration of 30 μg P L−1, and N to reach N:P molar ratio of 31); and (c) mixing: mixed (one rotation from surface to 3 m depth (speed of 1 m 4 min−1, total of 10 cycles)) versus static. Our findings suggest that under ambient nutrient conditions there is a synergistic effect between vertical mixing and UVR, increasing phytoplankton photosynthetic inhibition and excretion of organic carbon (EOC) from opaque lakes as compared to algae that received constant mean irradiance within the epilimnion. The opposite occurs in clear lakes where antagonistic effects were determined, with mixing partially counteracting the negative effects of UVR. Nutrient input, mimicking atmospheric pulses from Saharan dust, reversed this effect and clear lakes became more inhibited during mixing, while opaque lakes benefited from the fluctuating irradiance regime. These climate change related scenarios of nutrient input and increased mixing, would not only affect photosynthesis and production in lakes, but might also further influence the microbial loop and trophic interactions via enhanced EOC under fluctuating UVR exposure.This work was supported by Ministerio Español de Medio Ambiente, Rural y Marino (PN2009/067) and Ciencia e Innovación (GLC2008-01127/BOS and CGL2011-23681), Junta de Andalucía (Excelencia CVI-02598), Agencia Nacional de Promoción Científica y Tecnológica (PICT 2007-1651) and Fundación Playa Unión; GH and CD were supported by the Spanish Government – Formación de Profesorado Universitario Grant

Phosphatidylcholine-coated iron oxide nanomicelles for in vivo prolonged circulation time with an antibiofouling protein corona

We report the synthesis of micellar phosphatidylcholine-coated superparamagnetic iron oxide nanoparticles as a new long circulation contrast agents for magnetic resonance imaging. Oleic acid-coated Fe3 O4 nanoparticles were first prepared through thermal degradation and then encapsulated into small clusters with a phosphatidylcholine coating to obtain hydrophilic nanomicelles. A thorough characterization confirmed the chemical nature of the coating and the excellent colloidal stability of these nanomicelles in aqueous media. Magnetization and relaxivity properties proved their suitability as magnetic resonance imaging (MRI) contrast agent and in vitro cell viability data showed low toxicity. Vascular lifetime and elimination kinetics in the liver were assessed by blood relaxometry and by in vivo MRI in rats and compared with "control" particles prepared with a polyethylene glycol derivative. These micellar particles had a lifetime in blood of more than 10 h, much longer than the control nanoparticles (≈2 h), which is remarkable considering that the coating molecule is a small biocompatible zwitterionic phospholipid. The protein corona was characterized after incubation with rat serum at different times by high-throughput proteomics, showing a higher proportion of bound apolipoproteins and other dysopsonins for the phosphatidylcholine particles. The antibiofouling properties of this corona and its resistance to the adsorption of proteins corroborate the observed enhanced stability and prolonged systemic circulation.This study is supported by a grant from FP7 Marie Curie, Pulmonary imaging network (PINET), by Fundacio La Maratode TV3 (70/C/2012) and by a grant from the Comunidad de Madrid (S2010/BMD-2326, Inmunothercan-CM) and by Spanish Economy Ministry (MAT2013-47303-P). We thank E. Urones (Centro Nacional de Microscopia de la Universidad Complutense de Madrid) for the transmission electronic microscopy imaging; P. Morales (Instituto de Ciencia de Materiales de la Universidad Autonoma de Madrid) for the thermogravimetric and magnetization analysis and B. Salinas (Fundacion Centro Nacional de Investigaciones Cardiovasculares and CIBER de Enfermedades Respiratorias) for the TEM picture of oleic acid coated Fe3 O4. The authors declare no competing financial interests.S

In vivo imaging of lung inflammation with neutrophil-specific Ga-68 nano-radiotracer

In vivo detection and quantification of inflammation is a major goal in molecular imaging. Furthermore, cell-specific detection of inflammation would be a tremendous advantage in the characterization of many diseases. Here, we show how this goal can be achieved through the synergistic combination of nanotechnology and nuclear imaging. One of the most remarkable features of this hybrid approach is the possibility to tailor the pharmacokinetics of the nanomaterial-incorporated biomolecule and radionuclide. A good example of this approach is the covalent binding of a large amount of a neutrophil-specific, hydrophobic peptide on the surface of Ga-68 core-doped nanoparticles. This new nano-radiotracer has been used for non-invasive in vivo detection of acute inflammation with very high in vivo labelling efficiency, i.e. a large percentage of labelled neutrophils. Furthermore, we demonstrate that the tracer is neutrophil-specific and yields images of neutrophil recruitment of unprecedented quality. Finally, the nano-radiotracer was successfully detected in chronic inflammation in atherosclerosis-prone ApoE(-/-) mice after several weeks on a high-fat diet

Comprehensive Quantification of the Modified Proteome Reveals Oxidative Heart Damage in Mitochondrial Heteroplasmy

Post-translational modifications hugely increase the functional diversity of proteomes. Recent algorithms based on ultratolerant database searching are forging a path to unbiased analysis of peptide modifications by shotgun mass spectrometry. However, these approaches identify only one-half of the modified forms potentially detectable and do not map the modified residue. Moreover, tools for the quantitative analysis of peptide modifications are currently lacking. Here, we present a suite of algorithms that allows comprehensive identification of detectable modifications, pinpoints the modified residues, and enables their quantitative analysis through an integrated statistical model. These developments were used to characterize the impact of mitochondrial heteroplasmy on the proteome and on the modified peptidome in several tissues from 12-week-old mice. Our results reveal that heteroplasmy mainly affects cardiac tissue, inducing oxidative damage to proteins of the oxidative phosphorylation system, and provide a molecular mechanism explaining the structural and functional alterations produced in heart mitochondria.We thank Simon Bartlett (CNIC) for English editing. This study was supported by competitive grants from the Spanish Ministry of Economy and Competitiveness (MINECO) (BIO2015-67580-P) through the Carlos III Institute of Health-Fondo de Investigacion Sanitaria (PRB2, IPT13/0001-ISCIII-SGEFI/FEDER; ProteoRed), by Fundacion La Marato TV3, and by FP7-PEOPLE-2013-ITN ``Next-Generation Training in Cardiovascular Research and Innovation-Cardionext.'' N.B. is a FP7-PEOPLE-2013-ITN-Cardionext Fellow. The CNIC is supported by the MINECO and the Pro-CNIC Foundation, and is a Severo Ochoa Center of Excellence (MINECO Award SEV-2015-0505).S