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Background and Purpose: Reperfusion therapy is the standard of care for ischaemic stroke; however, there is a need to identify new therapeutic targets able to ameliorate cerebral damage. Neutrophil β1 adrenoceptors (β1AR) have been linked to neutrophil migration during exacerbated inflammation. Given the central role of neutrophils in cerebral damage during stroke, we hypothesize that β1AR blockade will improve stroke outcomes. Experimental Approach: Rats were subjected to middle cerebral artery occlusion–reperfusion to evaluate the effect on stroke of the selective β1AR blocker metoprolol (12.5 mg·kg−1) when injected i.v. 10 min before reperfusion. Key Results: Magnetic resonance imaging and histopathology analysis showed that pre-reperfusion i.v. metoprolol reduced infarct size. This effect was accompanied by reduced cytotoxic oedema at 24 h and vasogenic oedema at 7 days. Metoprolol-treated rats showed reduced brain neutrophil infiltration and those which infiltrated displayed a high proportion of anti-inflammatory phenotype (N2, YM1+). Additional inflammatory models demonstrated that metoprolol specifically blocked neutrophil migration via β1AR and excluded a significant effect on the glia compartment. Consistently, metoprolol did not protect the brain in neutrophil-depleted rats upon stroke. In patients suffering an ischaemic stroke, β1AR blockade by metoprolol reduced circulating neutrophil–platelet co-aggregates. Conclusions and Implications: Our findings describe that β1AR blockade ameliorates cerebral damage by targeting neutrophils, identifying a novel therapeutic target to improve outcomes in patients with stroke. This therapeutic strategy is in the earliest stages of the translational pathway and should be further explored. This study received funding from the Instituto de Salud Carlos III (ISCIII; PI16/02110 to B.I. and PT20/00044 to M.D.), the European Regional Development Fund (ERDF) "A way of making Europe" the Comunidad de Madrid (S2017/BMD-3867 RENIM-CM to M.D. and B.I.), cofunding from European structural, and investment funds, and by Agencia Estatal de Investigación (PID2019-110369RB-I00 to B.I.). B.I. is a recipient of funding from the European Research Council (ERC) under the European Union Horizon 2020 Research and Innovation Programme (ERC-Consolidator Grant agreement No. 819775). E.O. is a recipient of funds from the Comunidad de Madrid Programa de Atracción de Talento (2017-T1/BMD-5185) and from a Ramón y Cajal grant (RYC2020-028884-I) funded by MCIN/AEI/10.13039/501100011033 and by "ESF Investing in your future." A.C.M. is the beneficiary of an FPU fellowship from the Ministerio de Ciencia e Innovación (FPU2017/01932). M.C.C. is the beneficiary of a Miguel Servet contract (MS16/00174). The CNIC is supported by the ISCIII, the Ministerio de Ciencia e Innovación and the Pro CNIC Foundation and is a Severo Ochoa Center of Excellence (CEX2020-001041-S).

Reperfusion therapy is the standard of care for ischaemic stroke; however, there is a need to identify new therapeutic targets able to ameliorate cerebral damage. Neutrophil βRats were subjected to middle cerebral artery occlusion-reperfusion to evaluate the effect on stroke of the selective β1AR blocker metoprolol (12.5 mg·kgMagnetic resonance imaging and histopathology analysis showed that pre-reperfusion i.v. metoprolol reduced infarct size. This effect was accompanied by reduced cytotoxic oedema at 24 h and vasogenic oedema at 7 days. Metoprolol-treated rats showed reduced brain neutrophil infiltration and those which infiltrated displayed a high proportion of anti-inflammatory phenotype (N2, YM1Our findings describe that β1AR blockade ameliorates cerebral damage by targeting neutrophils, identifying a novel therapeutic target to improve outcomes in patients with stroke. This therapeutic strategy is in the earliest stages of the translational pathway and should be further explored.

Stimuli-responsive nanomaterials are very attractive for biomedical applications. They can be activated through external stimuli or by the physico-chemical conditions present in cells or tissues. Here, we describe the preparation of hybrid iron oxide-manganese oxide core-satellite shell nanostructures that change their contrast mode in magnetic resonance imaging (MRI) from T2 to T1, after being internalized by cells. This occurs by the dissolution of the MnO2 of the shell, preserving intact the iron oxide at the core. First, we study the seeded-growth synthesis of iron oxide-manganese oxide nanoparticles studying the effect of varying the core size of the magnetic seeds and the concentration of the surfactant. This allows tuning the size and shape of the final hybrid nanostructure. Then, we show that the shell can be removed by a redox reaction with glutathione, which is naturally present inside the cells at much higher concentrations than outside the cells. Finally, the dissolution of the MnO2 shell and the change in the contrast mode is confirmed in cell cultures. After this process, the iron oxide nanoparticles at the core remain intact and are still active as heating mediators when an alternating magnetic field is applied., This work was supported by the Ministerio de Ciencia e Innovación (PID2019-106301RB-I00 to GS and PGC2018-096016-B-I00 to L.G.), Instituto de Salud Carlos III (PT20/00044; co‐funded by European Union, European Regional Development Fund, ERDF, “A way of making Europe”), Ministerio de Economía y Competitividad (SAF2017-87305-R), Comunidad de Madrid (fellowships PEJD-2016/IND-2293 and IND2017/IND-7809; projects B2017/BMD-3867; P2018/NMT-4321), European Commission H2020 programme (project NOCANTHER, grant agreement no. 685795), and Asociación Española Contra el Cáncer. IMDEA Nanociencia acknowledges support from the 'Severo Ochoa' Programme for Centres of Excellence in R&D (MINECO, Grant SEV-2016-0686). The CNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia e Innovación (MCIN) and the Pro CNIC Foundation. L.G. acknowledges financial support from the Ramón y Cajal program (RYC-2014-15512). N. L-G also thanks the Spanish Education Ministry for the funding (FPU18/02323). P. M-R- acknowledges the support from the ‘Severo Ochoa’ Programme for Centres of Excellence in R&D (MINECO, Grant SEV-2016-0686).

Existing clinical or microbiological scores are not sensitive enough to obtain prompt identification of patients at risk of complicated Clostridium difficile infection (CDI). Our aim was to use a CDI animal model to evaluate 2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography ([18F]FDG-PET) as a marker of severe course of infection. CDI was induced with cefoperazone for 10 days followed by clindamycin 1 day before C. difficile inoculation. Mice were divided into wild type (n = 6), antibiotic without infection (AC n = 4), h001-infected (n = 5, ribotype 001), and h027-infected (n = 5, ribotype 027). Two days after inoculation, [18F]FDG-PET was acquired. Weight, general animal condition, and survival were monitored daily for 9 days. h001 group showed symptoms for 4 days with 0 % mortality and a similar colon uptake than control animals (h001 0.52 ± 0.20, WT 0.42 ± 0.07, and AC 0.36 ± 0.06). The h027 group showed symptoms up to 7 days, with 66.7 % of mortality 4 days after infection, and significantly higher colon uptake (0.93 ± 0.38, p

E.X. and R.M. contributed equally to this work. Work partially supported by the Ministerio de Ciencia, Innovacion y Universidades (PID2019-106301RB-I00 and PID2019-105195RA-I00), by the Spanish Ministry of Economy and Competitiveness (MAT2017-85617-R, SEV-2016-0686), by the Comunidad de Madrid (RENIM-CM, B2017/BMD-3867, co-financed by the European Structural and Investment Fund; NANOMAGCOST-CM P2018/NMT-4321), by the European COST Actions CA17115 (MyWave) and CA17140 (Nano2Clinic), by the Spanish Scientific Network HiperNano (RED2018-102626-T) and by the European Commission Horizon 2020 project NanoTBTech (Grant Number: 801305). D.G.-C. acknowledges CAM for funding PEJ-2018-AI/IND-11245. A.B. acknowledges funding from Comunidad de Madrid through TALENTO grant ref. 2019-T1/IND-14014. E.X. is grateful for a Juan de la Cierva Formacion scholarship (FJC2018-036734-I). R.M. acknowledges the support of the European Commission through the European Union's Horizon 2020 research and innovation program under the Marie Skodowska-Curie Grant agreement N 797945 (LANTERNS). A. E. acknowledges the support from Comunidad de Madrid (Talento project 2018-T1/IND-1005) and from AECC (Ideas Semilla 2019 project). P.R.S. is grateful for a Juan de la Cierva Incorporacion scholarship (IJC2019-041915-I). Procedures involving animal experiments were approved by the regional authority for animal experimentation of the Comunidad de Madrid and were conducted in agreement with the Universidad Autonoma de Madrid Ethics Committee, in compliance with the European Union directives 63/2010UE and Spanish regulation RD 53/2013., Deliberate and local increase of the temperature within solid tumors represents an effective therapeutic approach. Thermal therapies embrace this concept leveraging the capability of some species to convert the absorbed energy into heat. To that end, magnetic hyperthermia (MHT) uses magnetic nanoparticles (MNPs) that can effectively dissipate the energy absorbed under alternating magnetic fields. However, MNPs fail to provide real-time thermal feedback with the risk of unwanted overheating and impeding on-the-fly adjustment of the therapeutic parameters. Localization of MNPs within a tissue in an accurate, rapid, and cost-effective way represents another challenge for increasing the efficacy of MHT. In this work, MNPs are combined with state-of-the-art infrared luminescent nanothermometers (LNTh; Ag2S nanoparticles) in a nanocapsule that simultaneously overcomes these limitations. The novel optomagnetic nanocapsule acts as multimodal contrast agents for different imaging techniques (magnetic resonance, photoacoustic and near-infrared fluorescence imaging, optical and X-ray computed tomography). Most crucially, these nanocapsules provide accurate (0.2 degrees C resolution) and real-time subcutaneous thermal feedback during in vivo MHT, also enabling the attainment of thermal maps of the area of interest. These findings are a milestone on the road toward controlled magnetothermal therapies with minimal side effects., Ministerio de Ciencia, Innovacion y Universidades PID2019-106301RB-I00 PID2019-105195RA-I00, Spanish Ministry of Economy and Competitiveness MAT2017-85617-R SEV-2016-0686, Comunidad de Madrid (RENIM-CM - European Structural and Investment Fund) B2017/BMD-3867 NANOMAGCOST-CM P2018/NMT-4321, Spanish Scientific Network HiperNano RED2018-102626-T, European Commission Horizon 2020 project NanoTBTech 801305, CAM PEJ-2018-AI/IND-11245, Comunidad de Madrid 2019-T1/IND-14014, Juan de la Cierva Formacion scholarship FJC2018-036734-I, European Commission through the European Union's Horizon 2020 research and innovation program under the Marie Skodowska-Curie Grant 797945, Juan de la Cierva Incorporacion scholarship IJC2019-041915-I, Comunidad de Madrid 2018-T1/IND-1005, AECC (Ideas Semilla 2019 project), European COST Action (MyWave) CA17115, European COST Action (Nano2Clinic) CA17140

Stem-cell-derived extracellular vesicles (EVs) have demonstrated multiple beneficial effects in preclinical models of cardiac diseases. However, poor retention at the target site may limit their therapeutic efficacy. Cardiac extracellular matrix hydrogels (cECMH) seem promising as drug-delivery materials and could improve the retention of EVs, but may be limited by their long gelation time and soft mechanical properties. Our objective was to develop and characterize an optimized product combining cECMH, polyethylene glycol (PEG), and EVs (EVs–PEG–cECMH) in an attempt to overcome their individual limitations: long gelation time of the cECMH and poor retention of the EVs. The new combined product presented improved physicochemical properties (60% reduction in half gelation time, p &lt, 0.001, and threefold increase in storage modulus, p &lt, 0.01, vs. cECMH alone), while preserving injectability and biodegradability. It also maintained in vitro bioactivity of its individual components (55% reduction in cellular senescence vs. serum-free medium, p &lt, 0.001, similar to EVs and cECMH alone) and increased on-site retention in vivo (fourfold increase vs. EVs alone, p &lt, 0.05). In conclusion, the combination of EVs–PEG–cECMH is a potential multipronged product with improved gelation time and mechanical properties, increased on-site retention, and maintained bioactivity that, all together, may translate into boosted therapeutic efficacy.

In both clinical and preclinical scenarios, 2-deoxy-2[18F]fluoro-D-glucose ([18F]FDG) is the radiotracer most widely used to study brain glucose metabolism with positron emission tomography (PET). In clinical practice, there is a worldwide standardized protocol for preparing patients for [18F]FDG-PET studies, which specifies the room lighting. However, this standard is typically not observed in the preclinical field, although it is well known that animal handling affects the biodistribution of [18F]FDG. The present study aimed to evaluate the effect of ambient lighting on brain [18F]FDG uptake in mice. Two [18F]FDG-PET studies were performed on each animal, one in light and one in dark conditions. Thermal video recordings were acquired to analyse animal motor activity in both conditions. [18F]FDGPET images were analysed with the Statistical Parametric Mapping method. The results showed that [18F]FDG uptake is higher in darkness than in light condition in mouse nucleus accumbens, hippocampus, midbrain, hindbrain, and cerebellum. The SPM analysis also showed an interaction between the illumination condition and the sex of the animal. Mouse activity was significantly different (p = 0.01) between light conditions (632 ± 215 s of movement) and dark conditions (989 ± 200 s), without significant effect of sex (p = 0.416). We concluded that room illumination conditions during [18F]FDG uptake in mice affected the brain [18F]FDG biodistribution. Therefore, we highlight the importance to control this factor to ensure more reliable and reproducible mouse brain [18F]FDG-PET results. This work was partially supported by the Comunidad de Madrid (S2017/BMD-3867 RENIMCM), and it was co-financed by the European Structural and Investment Fund. And by Ministerio de Ciencia e Innovación, Instituto de Salud Carlos III (PT20/00044), cofunded by European Union, European Regional Development Fund (ERDF),"A way of making Europe". The CNIC is supported by the ISCIII, the Ministerio de Ciencia e Innovación.

Exclusion of brain metastases from clinical trials is a major cause of the limited therapeutic options for this growing population of cancer patients. Here, we report a medium-throughput drug-screening platform (METPlatform) based on organotypic cultures that allows to evaluate inhibitors against metastases growing in situ. By applying this approach to brain metastasis, we identified several hits from a library of FDA approved inhibitors and others being tested in clinical trials. A blood-brain barrier permeable HSP90 inhibitor showed high potency against mouse and human brain metastases at clinically relevant stages of the disease, including a novel model of local relapse after neurosurgery. Furthermore, in situ proteomic analysis applied to organotypic cultures with metastases treated with the chaperone inhibitor revealed novel biomarkers in human brain metastasis and actionable mechanisms of resistance. Our work validates METPlatform as a potent resource for metastasis research integrating drug-screening and unbiased omic approaches that is fully compatible with human samples. We envision that METPlatform could be established as a clinically relevant strategy to personalize the management of metastatic disease in the brain and elsewhere.SummarySystemic spread of cancer continues to be the key aspect associated with lethality. In this publication, Zhu et al. describes a drug-screening platform specifically designed to study vulnerabilities of metastasis when colonizing secondary organs and demonstrates its value in difficult-to-treat brain metastasis using new models and patient-derived samples.

Cardiomyocytes are subjected to the intense mechanical stress and metabolic demands of the beating heart. It is unclear whether these cells, which are long-lived and rarely renew, manage to preserve homeostasis on their own. While analyzing macrophages lodged within the healthy myocardium, we discovered that they actively took up material, including mitochondria, derived from cardiomyocytes. Cardiomyocytes ejected dysfunctional mitochondria and other cargo in dedicated membranous particles reminiscent of neural exophers, through a process driven by the cardiomyocyte's autophagy machinery that was enhanced during cardiac stress. Depletion of cardiac macrophages or deficiency in the phagocytic receptor Mertk resulted in defective elimination of mitochondria from the myocardial tissue, activation of the inflammasome, impaired autophagy, accumulation of anomalous mitochondria in cardiomyocytes, metabolic alterations, and ventricular dysfunction. Thus, we identify an immune-parenchymal pair in the murine heart that enables transfer of unfit material to preserve metabolic stability and organ function. Video Abstract: [Figure presented] A system of macrophages in the heart supports cardiomyocyte health by phagocytosing exopher particles ejected from cardiomyocytes that contain defective mitochondria, among other cellular contents., This study was supported by Intramural grants from the Severo Ochoa program (IGP-SO); grants SAF2015-71878-REDT and SAF2014-56819-R from the Ministerio de Ciencia e Innovacion (MICINN) to A.C.; European Research Council grant EU-rhythmy (ERC-ADG-2014-ID:669387) to S.G.P., and MATRIX (ERC-COG-2018-ID: 819775) to B.I.; L.G.N. is supported by SIgN core funding from A∗STAR; grant BFU2016-75144-R from the Ministry of Science and Innovation to J.A.B,; grants PGC2018-096486-B-I00 and RD16/0011/0019 (ISCIII) from MICINN, TNE-17CVD04 from the Leducq Foundation, and S2017/BMD-3875 from the Comunidad de Madrid to M.T; intramural grant TPC/O-SO and grants SAF2015-65633-R, RTI2018-099357-B-I00, and HFSP (RGP0016/2018) to J.A.E.; intramural grant IGP-SO to J.A.-C. and A.H.; BIO2017-83640-P and RYC-2014-16604 to J.A-C; grants PRB3 (IPT17/0019-ISCIII-SGEFI/ERDF, ProteoRed) from the Carlos III Institute of Health and Fondo de Investigaciones Sanitarias, BIO2015-67580-P and PGC2018-097019-B-I00 from MICINN to J.V.; RTI2018-096068 from MICINN, AFM, MDA, LaCaixa-HR17-00040, UPGRADE-H2020-825825, and European Research Council (ERC-741538) to P.M.C.; S2017/BMD-3867 RENIM-CM from the Comunidad de Madrid and cofunded with European structural and investment funds to M.D.; 120/C/2015-20153032 from Fundació la Marató de TV3, SAF2015-65607-R and RTI2018-095497-B-I00 from MICINN, HR17_00527 from La Caixa Foundation, and TNE-18CVD04 from the Leducq Foundation to A.H.; C.V.R. is a Howard Hughes Medical Institute Faculty Scholar; J.A.N-A is supported by fellowship SVP-2014-068595, A.V.L.-V. by SVP-2013-068089, L.E.-M. by FJCI-2016-29384, and A.R.-P. by BES-2016-076635, all from MICINN; and the CNIC International Postdoctoral Program (EU grant agreement 600396 to D.J.S.). The CNIC is supported by the MICINN and the Pro-CNIC Foundation and is a Severo Ochoa Center of Excellence (MICINN award SEV-2015-0505).

The morphological changes that occur in the central nervous system of patients with severe acute intermittent porphyria (AIP) have not yet been clearly established. The aim of this work was to analyse brain involvement in patients with severe AIP without epileptic seizures or clinical posterior reversible encephalopathy syndrome, as well as in a mouse model receiving or not liver-directed gene therapy aimed at correcting the metabolic disorder. We conducted neuroradiologic studies in 8 severely affected patients (6 women) and 16 gender- and age-matched controls. Seven patients showed significant enlargement of the cerebral ventricles and decreased brain perfusion was observed during the acute attack in two patients in whom perfusion imaging data were acquired. AIP mice exhibited reduced cerebral blood flow and developed chronic dilatation of the cerebral ventricles even in the presence of slightly increased porphyrin precursors. While repeated phenobarbital-induced attacks exacerbated ventricular dilation in AIP mice, correction of the metabolic defect using liver-directed gene therapy restored brain perfusion and afforded protection against ventricular enlargement. Histological studies revealed no signs of neuronal loss but a denser neurofilament pattern in the periventricular areas, suggesting compression probably caused by imbalance in cerebrospinal fluid dynamics. In conclusion, severely affected AIP patients exhibit cerebral ventricular enlargement. Liver-directed gene therapy protected against the morphological consequences of the disease seen in the brain of AIP mice. The observational study was registered at Clinicaltrial.gov as NCT02076763.

Nonrevascularizable coronary artery disease is a frequent cause of hibernating myocardium leading to heart failure (HF). Currently, there is a paucity of therapeutic options for patients with this condition. There is a lack of animal models resembling clinical features of hibernating myocardium. Here we present a large animal model of hibernating myocardium characterized by serial multimodality imaging. Yucatan minipigs underwent a surgical casein ameroid implant around the proximal left anterior descending coronary artery (LAD), resulting in a progressive obstruction of the vessel. Pigs underwent serial multimodality imaging including invasive coronary angiography, cardiac magnetic resonance (CMR), and hybrid 18F-Fluorodeoxyglucose positron emission tomography-computed tomography (FDG-PET/CT). A total of 43 pigs were operated on and were followed for 120 ± 37 days with monthly multimodality imaging. 24 pigs (56%) died during the follow-up. Severe LAD luminal stenosis was documented in all survivors. In the group of 19 long-term survivors, 17 (90%) developed left ventricular systolic dysfunction [median LVEF of 35% (IQR 32.5-40.5%)]. In 17/17, at-risk territory was viable on CMR and 14 showed an increased glucose uptake in the at-risk myocardium on 18FDG-PET/CT. The present pig model resembles most of the human hibernated myocardium characteristics and associated heart failure (systolic dysfunction, viable myocardium, and metabolic switch to glucose). This human-like model might be used to test novel interventions for nonrevascularizable coronary artery disease and ischemia heart failure as a previous stage to clinical trials. his study has been partially funded by the Horizon 2020 European Research Area Network on Cardiovascular Diseases (ERA-CVD) Joint Transnational Call “AC16/00021: FAT-4HEART,” by the Spanish Society of Cardiology through a “Translational Research grant 2019,” and by the Instituto de Salud Carlos III (ISCIII) and the European Regional Development Fund (ERDF) through a FIS grant (Ref # PI16/02110). Imaging phenotyping was partially supported by the Comunidad de Madrid (S2017/BMD-3867 RENIM-CM) and cofunded with European structural and investment funds. The CNIC is supported by the ISCIII, the Ministerio de Ciencia e Innovación and the Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence (SEV-2015-0505). Sí

Myocardial uptake can hamper visualization of lung tumors, atherosclerotic plaques, and inflammatory diseases in 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) studies because it leads to spillover in adjacent structures. Several preparatory pre-imaging protocols (including dietary restrictions and drugs) have been proposed to decrease physiological [18F]FDG uptake by the heart, although their effect on tumor glucose metabolism remains largely unknown. The objective of this study was to assess the effects of a ketogenic diet (as an alternative protocol to fasting) on tumor glucose metabolism assessed by [18F]FDG positron emission tomography (PET) in a mouse model of lung cancer. PET scans were performed 60 min after injection of 18.5 MBq of [18F]FDG. PET data were collected for 45 min, and an x-ray computed tomograph (CT) image was acquired after the PET scan. A PET/CT study was obtained for each mouse after fasting and after the ketogenic diet. Quantitative data were obtained from regions of interest in the left ventricular myocardium and lung tumor. Three days on a ketogenic diet decreased mean standard uptake value (SUVmean) in the myocardium (SUVmean 0.95 ± 0.36) more than one night of fasting (SUVmean 1.64 ± 0.93). Tumor uptake did not change under either dietary condition. These results show that 3 days on high-fat diets prior to [18F]FDG-PET imaging does not change tumor glucose metabolism compared with one night of fasting, although high-fat diets suppress myocardial [18F]FDG uptake better than fasting.

Early diagnosis in Alzheimer's disease (AD), prior to the appearance of marked clinical symptoms, is critical to prevent irreversible neuronal damage and neural malfunction that lead to dementia and death. Therefore, there is an urgent need to generate new contrast agents which reveal by a noninvasive method the presence of some of the pathological signs of AD. In the present study, we demonstrate for the first time a new nanoconjugate composed of magnetic nanoparticles bound to an antiferritin antibody, which has been developed based on the existence of iron deposits and high levels of the ferritin protein present in areas with a high accumulation of amyloid plaques (particularly the subiculum in the hippocampal area) in the brain of a transgenic mouse model with five familial AD mutations. Both in vitro and after intravenous injection, functionalized magnetic nanoparticles were able to recognize and bind specifically to the ferritin protein accumulated in the subiculum area of the AD transgenic mice., Reina Sofia Foundation (to M.R.G.). This study was also funded by the Comunidad de Madrid (Neurotec-S2010/BMD-2460). Work at A.M.-S, laboratory was funded by MINECO (SAF2010-17167 and SAF2014-56101-R), Comunidad de Madrid (Neurostem-CM S2010-BMD-2336) and Instituto de Salud Carlos III

Background Extensive coronary artery disease without revascularization options results in ischemic hibernated myocardium and ultimately heart failure (HF). A metabolic reprogramming of the heart characterized by a shift from fatty acids (FFA) to glucose as the preferential metabolic substrate is frequent in HF and hibernated myocardium. Previous studies in mouse models of HF have shown that high fat diet (HFD) is able to reverse the metabolic reprogramming of the heart and improve cardiac function. Here we used a translational large animal model of ischemic HF evaluated by state-of-the-art imaging modalities Methods IHM was generated in Yucatan minipigs (N=50) by progressive stenosis of the proximal left anterior descending (LAD) after surgical insertion of a casein ameroid around the artery. Pigs underwent a serial multimodality imaging study including magnetic resonance imaging (MRI), multi-tracer PET/CT (18FDG & 14(R,S)-[18F]Fluoro-6-thia-heptadecanoic acid (18F-FTHA) 19F-palmitate in-house synthetized), and invasive coronary angiography (ICA). Once hibernated myocardium with metabolic switch was documented (complete occlusion of LAD, LVEF Results The first part of the study was dedicated to establish the model in 42 pigs. Pigs were followed-up for 170±24 days with monthly ICA, MRI and PET/CT. Severe LAD stenosis was documented in all pigs 3–4 weeks after surgery. Mortality during follow-up was 57% (n=24) (15 peri-procedure, 2 between day 1 and 7, seven between day 7 and 45). Among those surviving beyond the sixth week, 90% (16/18) developed hibernated myocardium (mean LVEF 36±10%, absence of transmural delayed enhancement, contractile reserve and metabolic switch on PET/CT). In the second part of the study, 5 long-term survivors were allocated to receive HFD (20% extra lard), while 3 pigs served as controls. Mean LVEF before HFD initiation was 35±8%, and 36±5% in controls. Three months HFD was associated with a LVEF improvement in all treated animals (mean LVEF at the end of follow-up was 45±9%). Control animals on regular diet did not show any change in LVEF (35±5% at the end of follow-up) Conclusions We present a large animal model of HF due hibernated myocardium by placing an ameroid around the LAD. Mortality of this model is high (approx. 55%) but long-term survivors resemble all features seen in patients: LAD progressive stenosis with final occlusion but viable myocardium, LVEF deterioration and metabolic switch Feeding pigs with HFD to force cardiac reutilization of FFA, results in a consistent and large LVEF improvement in all animals Cardiac metabolic switch might be implicated in the pathogenesis of systolic dysfunction in hibernated myocardium and is a potential target for nutritional approaches

Vascular stiffness is a major cause of cardiovascular disease during normal aging and in Hutchinson–Gilford progeria syndrome (HGPS), a rare genetic disorder caused by ubiquitous progerin expression. This mutant form of lamin A causes premature aging associated with cardiovascular alterations that lead to death at an average age of 14.6 years. We investigated the mechanisms underlying vessel stiffness in LmnaG609G/G609G mice with ubiquitous progerin expression, and tested the effect of treatment with nitrites. We also bred LmnaLCS/LCSTie2Cre+/tgand LmnaLCS/LCSSM22αCre+/tg mice, which express progerin specifically in endothelial cells (ECs) and in vascular smooth muscle cells (VSMCs), respectively, to determine the specific contribution of each cell type to vascular pathology. We found vessel stiffness and inward remodeling in arteries of LmnaG609G/G609G and LmnaLCS/LCSSM22αCre+/tg, but not in those from LmnaLCS/LCSTie2Cre+/tgmice. Structural alterations in aortas of progeroid mice were associated with decreased smooth muscle tissue content, increased collagen deposition, and decreased transverse waving of elastin layers in the media. Functional studies identified collagen (unlike elastin and the cytoskeleton) as an underlying cause of aortic stiffness in progeroid mice. Consistent with this, we found increased deposition of collagens III, IV, V, and XII in the media of progeroid aortas. Vessel stiffness and inward remodeling in progeroid mice were prevented by adding sodium nitrite in drinking water. In conclusion, LmnaG609G/G609G arteries exhibit stiffness and inward remodeling, mainly due to progerin-induced damage to VSMCs, which causes increased deposition of medial collagen and a secondary alteration in elastin structure. Treatment with nitrites prevents vascular stiffness in progeria, This study was supported by the Spanish Ministerio de Ciencia, Innovación y Universidades (MCIU, grants SAF2016‐79490‐R and SAF2016‐80305‐P), with co‐funding from the European Regional Development Fund (ERDF, “Una manera de hacer Europa”). RvK and RKA are supported by NIH grants AG047373, T32‐GM008076, F31HL142160 and NSF grant CMMI 1548571. L.d.C. was supported by a Jordi Soler postdoctoral grant from the Red de Investigación Cardiovascular (RETIC Program, Instituto de Salud Carlos III), and A.S.‐L. was supported by a Severo Ochoa predoctoral grant from the MCIU (SVP‐2014‐068334) and by a grant from Asociación Apadrina la Ciencia‐Ford España‐Ford Motor Company Fund. The CNIC is supported by the MCIU and the Pro‐CNIC Foundation, and is a Severo Ochoa Center of Excellence (SEV‐2015‐0505)

[Background] The Platform for Innovation in Medical and Health Technologies (ITEMAS) is a network of 66 healthcare centres focused on fostering innovation in medical and health technologies as an essential tool for increasing the sustainability of the Spanish healthcare system. The present research is focused on defining a formal representation that details the most relevant concepts associated with the creation and adoption of innovative medical technology in the Spanish healthcare system., [Methods] The methodology applied is based on the methontology process, including peer-review identification and selection of concepts from the ITEMAS innovation indicators and innovation management system standards. This stage was followed by an iterative validation process. Concepts were then conceptualised, formalised and implemented in an ontology., [Results] The ontology defined describes how relationships between employees, organisations, projects and ideas can be applied to generate results that are transferrable to the market, general public and scientific forums. Overall, we identified 136 concepts, 138 object properties and 30 properties in a five-level hierarchy. The ontology was tested and validated as an appropriate framework for calculating the ITEMAS innovation indicators., [Conclusions] The consensus concepts were expressed in the form of an ontology to be used as a single communication format between the members of the ITEMAS network. Healthcare centres can compare their innovation results and obtain a better understanding of their innovation context based on the reasoning techniques of artificial intelligence. As a result, they can benefit from advanced analytical capabilities to define the most appropriate innovation policies for each centre based on the common experience of the large number of healthcare centres involved. The results can be used to create a map of agents and knowledge to show capabilities, projects and services provided by each of the participating centres. The ontology could also be applied as an instrument to match needs with existing projects and capabilities from the community of organisations working in healthcare technology innovation., This research has been partly funded by the ITEMAS Platform (PT13/0006/0001, PT13/0006/0036 and PT17/0005/0036) by the Ministerio de Ciencia, Innovación y Universidades, Instituto de Salud Carlos III, both co-funded by European Regional Development Fund (ERDF), “A way of making Europe”.

Myocardial uptake can hamper visualization of lung tumors, atherosclerotic plaques, and inflammatory diseases in 2-deoxy-2-[PET scans were performed 60 min after injection of 18.5 MBq of [Three days on a ketogenic diet decreased mean standard uptake value (SUVmean) in the myocardium (SUVmean 0.95 ± 0.36) more than one night of fasting (SUVmean 1.64 ± 0.93). Tumor uptake did not change under either dietary condition.These results show that 3 days on high-fat diets prior to [

In cases where catheter-related candidemia (CRC) must be managed without catheter withdrawal, antifungal lock therapy using highly active anti-biofilm (HAAB) agents is combined with systemic treatment. However, the activity of HAAB agents has never been studied in in vivo models using bioluminescence. We assessed the efficacy of micafungin using a bioluminescent Candida albicans SKCA23-ACTgLuc strain in an animal model of CRC. We divided 33 female Wistar rats into five groups: sham (A), infected nontreated (B), treated with lock therapy (0.16 mg/ml) (C), systemically treated only (1 mg/kg) (D), and systemically treated+lock (E). Catheters were colonized 24 h before insertion into the femoral vein (day 0). Treatment started on day 1 and lasted 7 days, followed by 7 days of surveillance. Bioluminescence assays were carried out on days 1, 3, 5, and 14, together with daily monitoring of clinical variables. Postmortem microbiological cultures from the catheter and several tissue samples were also obtained. Overall, 28 rats (84.8%) completed the study. Group B animals showed significant weight loss at days 2, 4, and 5 compared with groups C and D (P < .05). In group B, no animals survived after day 7, 75% had CRC, and bioluminescence remained constant 5 days after catheter implantation. Positive catheter culture rates in groups C, D, and E were, respectively, 83.3%, 62.5%, and 25.0% (P = .15). Micafungin proved to be a HAAB agent when administered both systemically and in lock therapy in an animal model of CRC, although the bioluminescence signal persists after treatment. This persistence should be further analyzed.

// Michele Cioffi 1 , Sara M. Trabulo 1, 2 , Mireia Vallespinos 1 , Deepak Raj 2 , Tony Bou Kheir 2 , Meng-Lay Lin 2 , Julfa Begum 2 , Ann-Marie Baker 3 , Ala Amgheib 2 , Jaimy Saif 4 , Manuel Perez 5 , Joaquim Soriano 5 , Manuel Desco 6, 7, 8 , Maria Victoria Gomez-Gaviro 6, 7, 8 , Lorena Cusso 6, 7, 8 , Diego Megias 5 , Alexandra Aicher 1, 2 , Christopher Heeschen 1, 2 1 Stem Cells & Cancer Group, Spanish National Cancer Research Centre (CNIO), Madrid, Spain 2 Stem Cells in Cancer & Ageing, Barts Cancer Institute, Queen Mary University of London, London, UK 3 Centre for Tumour Biology, Barts Cancer Institute, Queen Mary University of London, London, UK 4 School of Clinical Sciences, University of Bristol, Bristol, UK 5 Confocal Microscopy Unit, Centro Nacional de Investigaciones Oncologicas, Madrid, Spain 6 Departamento de Ingenieria Biomedica e Ingenieria Aeroespacial, Universidad Carlos III de Madrid, Leganes, Spain 7 Instituto de Investigacion Sanitaria Gregorio Maranon, Madrid, Spain 8 Centro de Investigacion Biomedica en Red de Salud Mental (CIBERSAM), Madrid, Spain Correspondence to: Alexandra Aicher, email: aicher_a@yahoo.com Christopher Heeschen, email: c.heeschen@qmul.ac.uk Keywords: metastasis, CD274, bone marrow, stromal niche, MDSC Received: June 29, 2016 Accepted: January 10, 2017 Published: February 17, 2017 ABSTRACT The stromal microenvironment controls response to injury and inflammation, and is also an important determinant of cancer cell behavior. However, our understanding of its modulation by miRNA (miR) and their respective targets is still sparse. Here, we identified the miR-25-93-106b cluster and two new target genes as critical drivers for metastasis and immune evasion of cancer cells. Using miR-25-93-106b knockout mice or antagomiRs, we demonstrated regulation of the production of the chemoattractant CXCL12 controlling bone marrow metastasis. Moreover, we identified the immune checkpoint PD-L1 (CD274) as a novel miR-93/106b target playing a central role in diminishing tumor immunity. Eventually, upregulation of miR-93 and miR-106b via miR-mimics or treatment with an epigenetic reader domain (BET) inhibitor resulted in diminished expression of CXCL12 and PD-L1. These data suggest a potential new therapeutic rationale for use of BET inhibitors for dual targeting of cancers with strong immunosuppressive and metastatic phenotypes.

Noonan syndrome (NS) is an autosomal dominant genetic disorder characterized by short stature, craniofacial dysmorphism, and congenital heart defects. NS also is associated with a risk for developing myeloproliferative disorders (MPD), including juvenile myelomonocytic leukemia (JMML). Mutations responsible for NS occur in at least 11 different loci including KRAS. Here we describe a mouse model for NS induced by K-Ras(V14I), a recurrent KRAS mutation in NS patients. K-Ras(V14I)-mutant mice displayed multiple NS-associated developmental defects such as growth delay, craniofacial dysmorphia, cardiac defects, and hematologic abnormalities including a severe form of MPD that resembles human JMML. Homozygous animals had perinatal lethality whose penetrance varied with genetic background. Exposure of pregnant mothers to a MEK inhibitor rescued perinatal lethality and prevented craniofacial dysmorphia and cardiac defects. However, Mek inhibition was not sufficient to correct these defects when mice were treated after weaning. Interestingly, Mek inhibition did not correct the neoplastic MPD characteristic of these mutant mice, regardless of the timing at which the mice were treated, thus suggesting that MPD is driven by additional signaling pathways. These genetically engineered K-Ras(V14I)-mutant mice offer an experimental tool for studying the molecular mechanisms underlying the clinical manifestations of NS. Perhaps more importantly, they should be useful as a preclinical model to test new therapies aimed at preventing or ameliorating those deficits associated with this syndrome.

Heart failure is one of the major cardiovascular problems worldwide, with Dilated cardiomyopathy (DCM) of ischemic origin is the most frequent cause. Design of novel therapies rely on the development of relevant animal models close to human anatomy and physiology. Ischemic DCM was induced in

Purpose Self-gated cine sequences are a common choice for cardiac MRI in preclinical applications. The aims of our work were to apply the compressed sensing technique to IntraGateFLASH cardiac MRI studies on rats and to find the maximum acceleration factor achievable with this technique. Theory and Methods Our reconstruction method extended the Split Bregman formulation to minimize the total variation in both space and time. In addition, we analyzed the influence of the undersampling pattern on the acceleration factor achievable. Results Our results show that acceleration factors of up to 15 are achievable with our technique when appropriate undersampling patterns are used. The introduction of a time-varying random sampling clearly improved the efficiency of the undersampling schemes. In terms of computational efficiency, the proposed reconstruction method has been shown to be competitive as compared with the fastest methods found in the literature. Conclusion We successfully applied our compressed sensing technique to self-gated cardiac cine acquisition in small animals, obtaining an acceleration factor of up to 15 with almost unnoticeable image degradation. Magn Reson Med 72:369–380, 2014. © 2013 Wiley Periodicals, Inc.

Purpose: The cardiac gating signal (CGS) in positron emission tomography (PET) studies is usually obtained from an electrocardiography (ECG) monitor. In this work, we propose a method to obtain the CGS in small-animal PET using the acquired list-mode data without using any hardware or end-user input. Procedures: The CGS was obtained from the number of coincidences over time acquired in the lines-of-response connected with the cardiac region. This region is identified in the image as its value changes with frequencies in the range of 3 to 12 Hz. The procedure was tested in a study with 29 Wistar rats and 6 mice injected with 2-deoxy-2-[18F]fluoro-D-glucose, which underwent a 45-min single-bed list-mode PET scan of the heart syncronized with an ECG. The estimated signals and the reconstructed images using eight-gated frames were compared with the ones obtained using the ECG signal from the monitor. Results: The differences of the PET-based CGS with respect to the ECG relative to the duration of the heartbeats were 5.6 % in rats and 11.0 % in mice. The reconstructed gated images obtained from the proposed method do not differ qualitatively with respect to the ones obtained with the ECG. The quantitative analysis of both set of images were performed measuring the volume of the left ventricle (LV) of the rats in the end-of-systole and end-of-diastole phase. The differences found in these parameters between both methods were below 12.1 % in the ESV and 9.3 % in the EDV with a 95 % confidence interval, which are comparable to the accuracy (7 %) of the method used for segmenting the LV. Conclusion: The proposed method is able to provide a valid and accurate CGS in small-animal PET list-mode data. the ones obtained using the ECG signal from the monitor. This work was supported in part by Consejería de Educación, Juventud y Deporte de la Comunidad de Madrid (Spain) through the Madrid-MIT M+Visión Consortium, Comunidad de Madrid (S2013/MIT-3024 TOPUS-CM), UCM (Grupos UCM, 910059), CPAN (Consolider-Ingenio 2010, CSPD-2007-00042), RIC-RETIC network, Spanish MINECO (RD12/0042/0057),Ministerio de Ciencia e Innovación, Spanish Government (ENTEPRASE grant, PSE-300000-2009-5 and TEC2007-64731/TCM), and European Regional funds.

Single-photon emission computed tomography (SPECT) offers the possibility to study the biodistribution of 111In-labeled human mesenchymal stem cells (hMSCs). This functional information can be complemented by magnetic resonance imaging (MRI) that provides the anatomical context to the SPECT image. However, acquiring such studies with separate SPECT/MRI systems requires a multimodality registration in order to fuse both images. In this work, a special bed was designed based on three non-coplanar glass capillary tubes filled with a mixture of 99mTc and CuSO4, visible in both modalities. Moreover, it was necessary to design a semi-automated rigid registration procedure based on matching these rods by minimizing the distance between corresponding lines. The maximum target registration error achieved was 0.9 mm, in the range of SPECT resolution.

Homing is an inherent, complex, multistep process performed by cells such as human bone marrow mesenchymal stem cells (hMSCs) to travel from a distant location to inflamed or damaged tissue and tumors. This ability of hMSCs has been exploited as a tumor-targeting strategy in cell-based cancer therapy. The purpose of this study was to investigate the applicability of 111In-oxine for tracking hMSCs in vivo by combining single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI). 111In-labeled hMSCs (106 cells) were infused intraperitoneally in neuroblastoma-bearing mice, whereas a control group received a dose of free 111In-oxine. SPECT and MRI studies were performed 24 and 48 hours afterwards. Initially, the images showed similar activity in the abdomen in both controls and hMSC-injected animals. In general, abdominal activity decreases at 48 hours. hMSC-injected animals showed increased uptake in the tumor area at 48 hours, whereas the control group showed a low level of activity at 24 hours, which decreased at 48 hours. In conclusion, tracking 111In-labeled hMSCs combining SPECT and MRI is feasible and may be transferable to clinical research. The multimodal combination is essential to ensure appropriate interpretation of the images. Financial disclosure of authors: This work was funded in part by grants from Ministerio de Economía y Competitividad (PLE2009-0115), Red Tematica de Investigacion Cooperativa en Cancer (RTICC/ISCIII; RD12/0036/0027), the Madrid Regional Government (S-BIO-0204-2006–MesenCAM and P2010/BMD-2420-CellCAM), and the Ministerio de Ciencia e Innovación (CEN-20101014 and TEC-2010-21619-C04-01). Sí

Tumor-homing is a complex, multistep process used by many cells, like mesenchymal stem cells (MSCs), to travel from a distant location to a tumor. The purpose of this study is to investigate the applicability of 111In-oxine for tracking human MSCs in vivo with single photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI). 111In labeled hMSCs (106 cells) were infused intraperitoneally in neuroblastoma tumor-bearing mice, and SPECT/MRI images were performed 24 and 48 hour afterwards. High initial hMSC localization occurred in the abdomen cavity and 48 hour later hMSC-injected animals showed tumor uptake. MRI information was essential to properly define Regions of Interest on the images. Tracking 111In labeled hMSC combining SPECT and MRI is feasible and may be transferable to human studies.

Self-gated cine sequences are a common choice for cardiac MRI in preclinical applications. The aims of our work were to apply the compressed sensing technique to IntraGateFLASH cardiac MRI studies on rats and to find the maximum acceleration factor achievable with this technique.Our reconstruction method extended the Split Bregman formulation to minimize the total variation in both space and time. In addition, we analyzed the influence of the undersampling pattern on the acceleration factor achievable.Our results show that acceleration factors of up to 15 are achievable with our technique when appropriate undersampling patterns are used. The introduction of a time-varying random sampling clearly improved the efficiency of the undersampling schemes. In terms of computational efficiency, the proposed reconstruction method has been shown to be competitive as compared with the fastest methods found in the literature.We successfully applied our compressed sensing technique to self-gated cardiac cine acquisition in small animals, obtaining an acceleration factor of up to 15 with almost unnoticeable image degradation.

Reporter gene (RG) imaging of cell-based therapies provides a direct readout of therapeutic efficacy by assessing the fate of implanted cells. To permit long-term cellular imaging, RGs are traditionally required to be integrated into the cellular genome. This poses a potential safety risk and regulatory bottleneck for clinical translation as integration can lead to cellular transformation. To address this issue, we have developed non-integrative, replicating minicircles (MCs) as an alternative platform for safer monitoring of cells in living subjects. We developed both plasmids and minicircles containing the scaffold/matrix attachment regions (S/MAR) of the human interferon-beta gene, driven by the CMV promoter, and expressing the bioluminescence RG firefly luciferase. Constructs were transfected into breast cancer cells, and expanded S/MAR minicircle clones showed luciferase signal for greater than 3 months in culture and minicircles remained as episomes. Importantly, luciferase activity in clonal populations was slowly lost over time and this corresponded to a loss of episome, providing a way to reversibly label cells. To monitor cell proliferation in vivo, 1.5×10(6) cells carrying the S/MAR minicircle were implanted subcutaneously into mice (n = 5) and as tumors developed significantly more bioluminescence signal was noted at day 35 and 43 compared to day 7 post-implant (p

This work describes a new iterative method for extracting time-activity curves (TAC) from dynamic imaging studies using a priori information from generic models obtained from TAC templates. Analytical expressions of the TAC templates were derived from TACs obtained by manual segmentation of three 13NH3 pig studies (gold standard). An iterative method for extracting both ventricular and myocardial TACs using models of the curves obtained as an initial template was then implemented and tested. These TACs were extracted from masked and unmasked images; masking was applied to remove the lungs and surrounding non-relevant structures. The resulting TACs were then compared with TACs obtained manually; the results of kinetic analysis were also compared. Extraction of TACs for each region was sensitive to the presence of other organs (e.g., lungs) in the image. Masking the volume of interest noticeably reduces error. The proposed method yields good results in terms of TAC definition and kinetic parameter estimation, even when the initial TAC templates do not accurately match specific tracer kinetics. This work is supported by the following grants: RD07/0014/2009, Subprograma RETICS, Ministerio de Ciencia e Innovación. S2009/DPI-1802 (ARTEMIS), Comunidad de Madrid. CEN-20101014, Programa CENIT, CDTI, Ministerio de Ciencia e Innovación. European Commission, EFPIA, INNOVATIVE MEDICINE INITIATIVE (PredDICT-TB project, 115337-1) Publicado

Quantitative analysis of dynamic cardiac PET studies provides useful diagnostic information regarding different heart diseases. To correctly perform the kinetic analysis, tracer time-activity curves (TAC) must be precisely extracted from the chosen tissue on the imaging data. In this work we present an alternative clustering algorithm for segmentation of dynamic studies using a leader-follower clustering approach, in which the number of clusters is unknown.

In this work we evaluate a combination of a high-performance PET with a superconductive 3T MRI system. This PET/MRI system is implemented in a tandem configuration similar to some scanners used in clinical imaging but with both subsystems placed in proximity, what allows sharing a common animal holder that travels between both FOVs. The PET detector consists in two rings, each one having 18 DOI detectors, which provide a high sensitivity with a 1.0mm3 resolution at the centre and minimal degradation toward the edges of the 68mm diameter FOV. The MRI system is a compact, cryogen-free, auto-shielded and with a very tight fringe-field and no special room requirements. These characteristics enable the close location of the PET detector without deteriorating its performance. We have evaluated the system obtaining images of three mice in cardiology, oncology and neurology applications. All PET scans were performed 30m after intraperitoneal FDG injection of 25.2±5.8MBq and were reconstructed using a 3D-OSEM algorithm. For the cardiac MRI a gated FLASH sequence and a body coil were used. Tumor MRI was using an FSE T1w sequence and a body coil. Finally, brain MRI was done with an FSE T2w sequence. In all the cases registration of PET and MRI images was obtained following an equivalent protocol to the standard PET/CT preclinical system, and since the animals didn’t move from the bed the results were satisfactory. Attenuation correction was not applied to any image. The preliminary results from this prototype suggest that the tandem solution for preclinical PET/MRI is feasible and has the advantage of preserving the best image quality in both modalities without requiring any trade-off of specifications. The limitation of not being able to perform simultaneous acquisitions is compensated by the high quality results of studies that do not actually require such simultaneous data collection.