Dustin Osborne

Synergistic effects of leucine and resveratrol on insulin sensitivity and fat metabolism in adipocytes and mice

BackgroundSirtuins are important regulators of glucose and fat metabolism, and sirtuin activation has been proposed as a therapeutic target for insulin resistance and diabetes. We have shown leucine to increase mitochondrial biogenesis and fat oxidation via Sirt1 dependent pathways. Resveratrol is a widely recognized activator of Sirt; however, the biologically-effective high concentrations used in cell and animal studies are generally impractical or difficult to achieve in humans. Accordingly, we sought to determine whether leucine would exhibit synergy with low levels of resveratrol on sirtuin-dependent outcomes in adipocytes and in diet-induced obese (DIO) mice.Methods3T3-L1 mouse adipocytes were treated with Leucine (0.5 mM), β-hydroxy-β-methyl butyrate (HMB) (5 μM) or Resveratrol (200 nM) alone or in combination. In addition, diet-induced obese mice were treated for 6-weeks with low (2 g/kg diet) or high (10 g/kg diet) dose HMB, Leucine (24 g/kg diet; 200% of normal level) or low (12.5 mg/kg diet) or high (225 mg/kg diet) dose resveratrol, alone or as combination with leucine-resveratrol or HMB-resveratrol.ResultsFatty acid oxidation, AMPK, Sirt1 and Sirt3 activity in 3T3-L1 adipocytes and in muscle cells, were significantly increased by the combinations compared to the individual treatments. Similarly, 6-week feeding of low-dose resveratrol combined with either leucine or its metabolite HMB to DIO mice increased adipose Sirt1 activity, muscle glucose and palmitate uptake (measured via PET/CT), insulin sensitivity (HOMAIR), improved inflammatory stress biomarkers (CRP, IL-6, MCP-1, adiponectin) and reduced adiposity comparable to the effects of high dose resveratrol, while low-dose resveratrol exerted no independent effect.ConclusionThese data demonstrate that either leucine or its metabolite HMB may be combined with a low concentration of resveratrol to exert synergistic effects on Sirt1-dependent outcomes; this may result in more practical dosing of resveratrol in the management of obesity, insulin-resistance and diabetes.

LaPO4 nanoparticles doped with actinium-225 that partially sequester daughter radionuclides.

Nanoscale materials have been envisioned as carriers for various therapeutic drugs, including radioisotopes. Inorganic nanoparticles (NPs) are particularly appealing vehicles for targeted radiotherapy because they can package several radioactive atoms into a single carrier and can potentially retain daughter radioisotopes produced by in vivo generators such as actinium-225 ((225)Ac, t(1/2) = 10 d). Decay of this radioisotope to stable bismuth-209 proceeds through a chain of short-lived daughters accompanied by the emission of four α-particles that release >27 MeV of energy. The challenge in realizing the enhanced cytotoxic potential of in vivo generators lies in retaining the daughter nuclei at the therapy site. When (225)Ac is attached to targeting agents via standard chelate conjugation methods, all of the daughter radionuclides are released after the initial α-decay occurs. In this work, (225)Ac was incorporated into lanthanum phosphate NPs to determine whether the radioisotope and its daughters would be retained within the dense mineral lattice. Further, the (225)Ac-doped NPs were conjugated to the monoclonal antibody mAb 201B, which targets mouse lung endothelium through the vasculature, to ascertain the targeting efficacy and in vivo retention of radioisotopes. Standard biodistribution techniques and microSPECT/CT imaging of (225)Ac as well as the daughter radioisotopes showed that the NPs accumulated rapidly in mouse lung after intravenous injection. By showing that excess, competing, uncoupled antibodies or NPs coupled to control mAbs are deposited primarily in the liver and spleen, specific targeting of NP-mAb 201B conjugates was demonstrated. Biodistribution analysis showed that ∼30% of the total injected dose of La((225)Ac)PO(4) NPs accumulated in mouse lungs 1 h postinjection, yielding a value of % ID/g >200. Furthermore, after 24 h, 80% of the (213)Bi daughter produced from (225)Ac decay was retained within the target organ and (213)Bi retention increased to ∼87% at 120 h. In vitro analyses, conducted over a 1 month interval, demonstrated that ∼50% of the daughters were retained within the La((225)Ac)PO(4) NPs at any point over that time frame. Although most of the γ-rays from radionuclides in the (225)Ac decay chain are too energetic to be captured efficiently by SPECT detectors, appropriate energy windows were found that provided dramatic microSPECT images of the NP distribution in vivo. We conclude that La((225)Ac)PO(4)-mAb 201B conjugates can be targeted efficiently to mouse lung while partially retaining daughter products and that targeting can be monitored by biodistribution techniques and microSPECT imaging.

Radioembolization and the dynamic role of 90Y PET/CT

Before the advent of tomographic imaging, it was postulated that decay of 90 Y to the 0+ excited state of 90Zr may result in emission of a positron–electron pair. While the branching ratio for pair-production is small (~32 × 10−6), PET has been successfully used to image 90 Y in numerous recent patients and phantom studies. 90 Y PET imaging has been performed on a variety of PET/CT systems, with and without time-of-flight (TOF) and/or resolution recovery capabilities as well as on both bismuth-germanate and lutetium yttrium orthosilicate (LYSO)-based scanners. On all systems, resolution and contrast superior to bremsstrahlung SPECT has been reported. The intrinsic radioactivity present in LYSO-based PET scanners is a potential limitation associated with accurate quantification of 90 Y. However, intrinsic radioactivity has been shown to have a negligible effect at the high activity concentrations common in 90 Y radioembolization. Accurate quantification is possible on a variety of PET scanner models, with or without TOF, although TOF improves accuracy at lower activity concentrations. Quantitative 90 Y PET images can be transformed into 3-dimensional (3D) maps of absorbed dose based on the premise that the 90 Y activity distribution does not change after infusion. This transformation has been accomplished in several ways, although the most common is with the use of 3D dose-point-kernel convolution. From a clinical standpoint, 90 Y PET provides a superior post-infusion evaluation of treatment technical success owing to its improved resolution. Absorbed dose maps generated from quantitative PET data can be used to predict treatment efficacy and manage patient follow-up. For patients who receive multiple treatments, this information can also be used to provide patient-specific treatment-planning for successive therapies, potentially improving response. The broad utilization of 90 Y PET has the potential to provide a wealth of dose–response information, which may lead to development of improved radioembolization treatment-planning models in the future.

Comparative Analysis of Peptide p5 and Serum Amyloid P Component for Imaging AA Amyloid in Mice Using Dual-Isotope SPECT

PurposeI-labeled human serum amyloid P component (SAP) is used clinically only in the UK for imaging visceral amyloidosis to assist with diagnosis, disease staging, and monitoring response to therapy. We compare a new amyloid-reactive probe, peptide p5, with SAP for imaging amyloidosis.ProceduresDual-energy SPECT/CT images were acquired of 125I-labeled SAP and 99mTc-labeled p5 in mice with systemic AA amyloidosis (n = 3). Twelve organs and tissues were harvested for radiotracer biodistribution assessment and for micro-autoradiographic analysis.ResultsI-SAP and 99mTc-p5 localized equivalently in amyloid deposits in liver (∼10% injected dose (ID)/g) whereas 125I-SAP was twofold higher in the spleen (∼20% ID/g; 99mTc-p5, ∼10% ID/g). In contrast, 99mTc-p5 was bound to pancreatic and intestinal amyloid approximately fivefold more efficiently as evidenced in biodistribution data.ConclusionsRadiolabeled p5 is an effective amyloid-imaging radiotracer as compared to SAP in the murine model of amyloidosis and may be rapidly translated for imaging patients with visceral amyloidosis in the USA.

Guidance for Methods Descriptions Used in Preclinical Imaging Papers

Preclinical molecular imaging is a rapidly growing field, where new imaging systems, methods, and biological findings are constantly being developed or discovered. Imaging systems and the associated software usually have multiple options for generating data, which is often overlooked but is essential when reporting the methods used to create and analyze data. Similarly, the ways in which animals are housed, handled, and treated to create physiologically based data must be well described in order that the findings be relevant, useful, and reproducible. There are frequently new developments for metabolic imaging methods. Thus, specific reporting requirements are difficult to establish; however, it remains essential to adequately report how the data have been collected, processed, and analyzed. To assist with future manuscript submissions, this article aims to provide guidelines of what details to report for several of the most common imaging modalities. Examples are provided in an attempt to give comprehensive, succinct descriptions of the essential items to report about the experimental process.

A Routine PET/CT Protocol with Streamlined Calculations for Assessing Cardiac Amyloidosis Using 18F-Florbetapir

Introduction Cardiac amyloidosis is a rare condition characterized by the deposition of well-structured protein fibrils, proteoglycans, and serum proteins as amyloid. Recent work has shown that it may be possible to use 18F-Florbetapir to image cardiac amyloidosis. Current methods for assessment include invasive biopsy techniques. This work enhances foundational work by Dorbala et al. by developing a routine imaging and analysis protocol using 18F-Florbetapir for cardiac amyloid assessment. Methods Eleven patients, three healthy controls and eight myloid positive patients, were imaged using 18F-Florbetapir to assess cardiac amyloid burden. Four of the patients were also imaged using 82Rb-Chloride to evaluate possible 18F-Florbetapir retention because of reduced myocardial blood flow. Quantitative methods using modeling, SUVs and SUV ratios were used to define a new streamlined clinical imaging protocol that could be used routinely and provide patient stratification. Results Quantitative analysis of 18F-Florbetapir cardiac amyloid data were compiled from a 20-min listmode protocol with data histogrammed into two static images at 0–5, 10–15, or 15–20 min. Data analysis indicated the use of SUVs or ratios of SUVs calculated from regions draw in the septal wall were adequate in identification of all healthy controls from amyloid positive patients in this small cohort. Additionally, we found that it may be possible to use this method to differentiate patients suffering from AL vs. TTR amyloid. Conclusion This work builds on the seminal work by Dorbala et al. by describing a short 18F-Florbetapir imaging protocol that is suitable for routine clinical use and uses a simple method for quantitative analysis of cardiac amyloid disease.

Development and Validation of a Complete GATE Model of the Siemens Inveon Trimodal Imaging Platform

This article presents and validates a newly developed GATE model of the Siemens Inveon trimodal imaging platform. Fully incorporating the positron emission tomography (PET), single-photon emission computed tomography (SPECT), and computed tomography (CT) data acquisition subsystems, this model enables feasibility studies of new imaging applications, the development of reconstruction and correction algorithms, and the creation of a baseline against which experimental results for real data can be compared. Model validation was based on comparing simulation results against both empirical and published data. The PET modality was validated using the NEMA NU-4 standard. Validations of SPECT and CT modalities were based on assessment of model accuracy compared to published and empirical data on the platform. Validation results show good agreement between simulation and empirical data of approximately ± 5%.

Dynamic PET and SPECT imaging with radioiodinated, amyloid-reactive peptide p5 in mice: A positive role for peptide dehalogenation

Dynamic molecular imaging provides bio-kinetic data that is used to characterize novel radiolabeled tracers for the detection of disease. Amyloidosis is a rare protein misfolding disease that can affect many organs. It is characterized by extracellular deposits composed principally of fibrillar proteins and hypersulfated proteoglycans. We have previously described a peptide, p5, which binds preferentially to amyloid deposits in a murine model of reactive (AA) amyloidosis. We have determined the whole body distribution of amyloid by molecular imaging techniques using radioiodinated p5. The loss of radioiodide from imaging probes due to enzymatic reaction has plagued the use of radioiodinated peptides and antibodies. Therefore, we studied iodine-124-labeled p5 by using dynamic PET imaging of both amyloid-laden and healthy mice to assess the rates of amyloid binding, the relevance of dehalogenation and the fate of the radiolabeled peptide. Rates of blood pool clearance, tissue accumulation and dehalogenation of the peptide were estimated from the images. Comparisons of these properties between the amyloid-laden and healthy mice provided kinetic profiles whose differences may prove to be indicative of the disease state. Additionally, we performed longitudinal SPECT/CT imaging with iodine-125-labeled p5 up to 72h post injection to determine the stability of the radioiodinated peptide when bound to the extracellular amyloid. Our data show that amyloid-associated peptide, in contrast to the unbound peptide, is resistant to dehalogenation resulting in enhanced amyloid-specific imaging. These data further support the utility of this peptide for detecting amyloidosis and monitoring potential therapeutic strategies in patients.

Reduction of Patient Anxiety in PET/CT Imaging by Improving Communication Between Patient and Technologist

Patients experience anxiety during imaging procedures because of the confined space, uncertainty about the procedure, worry about the results, and other concerns. When a patient experiences anxiety during PET/CT imaging, the quality of the scan can be affected in several ways. Current patient–technologist communication is limited in PET/CT because the technologist must be separated from the patient during the course of the imaging workflow. This study investigated the use of a call device enabling rapid communication to reduce patient anxiety. Methods: Clinical patients with various oncologic indications and undergoing 18F-FDG PET/CT imaging were asked to participate in anxiety surveys under several conditions. Metrics were tracked regarding the survey results for comparison between groups and survey conditions. During the course of this study, 2 patient surveys were used. One of the patient populations was asked to fill out a survey on personal perceptions of the use of such a device, with questions related to their comfort with the device and the degree to which they perceived the device to reduce their anxiety. The 2 remaining populations were given a standard Spielberger State Anxiety survey for anxiety assessments against control populations. Results: Perception survey results indicated that 75% of the respondents experienced a reduction in anxiety and that 84% would request this type of device for other procedures. A correlation was observed between improved patient–technologist communication and perceived feelings of safety, with identical percentages of positive responses. Although responses were mostly positive, 18.8% did not perceive any reduction in anxiety, and the same number indicated they would not use the system in the future. For those patients given the standard Spielberger State Anxiety survey, a statistically significant reduction in anxiety was observed (P < 0.05) in those patients given a call device. Reductions in anxiety were observed for all patient populations, including first-time and repeated-imaging patients. Conclusion: Patient anxiety can be reduced through the use of a tangible device that improves communication between the patient and the imaging staff. Reducing anxiety may have a positive effect on imaging, because involuntary motion may be reduced and there may be improvement in the patients’ comfort and in their overall experience with the imaging procedure.

Clinical workflow considerations for implementation of continuous-bed-motion PET/CT

Within the last 3 y, a new type of technology has emerged for PET imaging that uses a continuous-bed-motion (CBM) acquisition. For technologists, this type of acquisition requires some modifications of the standard approach to PET protocols and imaging workflows. There are several key aspects of CBM that technologists need to learn and understand when transitioning from traditional step-and-shoot PET imaging to this new technology, including differences in acquisition type, image quality, and protocol setup as well as the impact that CBM can have on workflow. This article explains how CBM differs from step and shoot and focuses on the issues critical for technologists to know when first using this technology.