Utilization of nanoparticles as X‐ray contrast agents for diagnostic imaging applications

Among all the diagnostic imaging modalities, X‐ray imaging techniques are the most commonly used owing to their high resolution and low cost. The improvement of these techniques relies heavily on the development of novel X‐ray contrast agents, which are molecules that enhance the visibility of internal structures within the body in X‐ray imaging. To date, clinically used X‐ray contrast agents consist mainly of small iodinated molecules that might cause severe adverse effects (e.g. allergies, cardiovascular diseases and nephrotoxicity) in some patients owing to the large and repeated doses that are required to achieve good contrast. For this reason, there is an increasing interest in the development of alternative X‐ray contrast agents utilizing elements with high atomic numbers (e.g. gold, bismuth, ytterbium and tantalum), which are well known for exhibiting high absorption of X‐rays. Nanoparticles (NPs) made from these elements have been reported to have better imaging properties, longer blood circulation times and lower toxicity than conventional iodinated X‐ray contrast agents. Additionally, the combination of two or more of these elements into a single carrier allows for the development of multimodal and hybrid contrast agents. Herein, the limitations of iodinated X‐ray contrast agents are discussed and the parameters that influence the efficacy of X‐ray contrast agents are summarized. Several examples of the design and production of both iodinated and iodine‐free NP‐based X‐ray contrast agents are then provided, emphasizing the studies performed to evaluate their X‐ray attenuation capabilities and their toxicity in vitro and in vivo. Copyright © 2014 John Wiley & Sons, Ltd.     

Selective X‐ray contrast enhancement of the spleen of living mice mediated by gold nanorods

Gold nanomaterials (AuNPs) represent a promising new class of contrast agents for X‐ray computed tomographic (CT) imaging in both research and clinical settings. These materials exhibit superior X‐ray absorption properties compared with other iodinated agents, and thus require lower injection doses. Gold is nonimmunogenic and therefore contributes to safety profile in living specimens. Unfortunately, most reports on the use of AuNPs as X‐ray CT enhancers only demonstrate marginal enhancement of the intended anatomical structure. In this study, we demonstrate the dramatic properties of gold nanorods (GNR) to serve as robust X‐ray CT contrast‐enhancing agent for selective imaging of the spleen. These organ‐specific uptake properties were delineated by performing longitudinal CT imaging of living mice that were dosed with GNR at 2 day intervals. Rapid uptake in spleen was noted within 12 h of first systemic administration with a change in contrast enhancement of 90 Hounsfield units (ΔHU = 90) and with two subsequent injections a total contrast enhancement of over 200 HU was observed. The resulting images provide excellent contrast that will enable the detailed anatomical visualization and study of a range of pre‐clinical models of spleen disease including infection and cancer. Copyright © 2014 John Wiley & Sons, Ltd.     

Tumor imaging in small animals with a combined micro‐CT/micro‐DSA system using iodinated conventional and blood pool contrast agents

X‐ray based micro‐computed tomography (CT) and micro‐digital subtraction angiography (DSA) are important non‐invasive imaging modalities for following tumorogenesis in small animals. To exploit these imaging capabilities further, the two modalities were combined into a single system to provide both morphological and functional data from the same tumor in a single imaging session. The system is described and examples are given of imaging implanted fibrosarcoma tumors in rats using two types of contrast media: (a) a new generation of blood pool contrast agent containing iodine with a concentration of 130 mg/mL (Fenestra? VC, Alerion Biomedical, San Diego, CA, USA) for micro‐CT and (b) a conventional iodinated contrast agent (Isovue®‐370 mg/mL iodine, trademark of Bracco Diagnostics, Princeton, NJ, USA) for micro‐DSA. With the blood pool contrast agent, the 3D vascular architecture is revealed in exquisite detail at 100 µm resolution. Micro‐DSA images, in perfect registration with the 3D micro‐CT datasets, provide complementary functional information such as mean transit times and relative blood flow through the tumor. This imaging approach could be used to understand tumor angiogenesis better and be the basis for evaluating anti‐angiogenic therapies. Copyright © 2006 John Wiley & Sons Ltd.     

Managing children with acute non‐traumatic limp: the utility of clinical findings,laboratory inflammatory markers and X‐rays

Objectives: To examine the utility of clinical findings, laboratory markers and X‐ray radiographs (X‐ray) in the assessment of children presenting with an acute non‐traumatic limp. Methods: A retrospective review of all children who received hip X‐rays over a 2 year period in the Children's Emergency Department, Starship Children's Hospital, Auckland, New Zealand. Children were identified from the radiology database and clinical notes reviewed. Children aged 0–12 years old were included if the limp was acute (less than 2 weeks of duration) with no history of trauma. X‐rays were reported by a consultant paediatric radiologist. Univariate and multivariate analysis was performed to determine predictors of osteomyelitis and septic arthritis. Receiver operator curves were used to assess the optimum cut‐off points for C reactive protein (CRP), erythrocyte sedimentation rate (ESR) and white cell count (WCC). Results: A total of 350 patients were enrolled. There were 21 (6%) abnormal X‐rays . Fever, non‐weight bearing, raised white cell count, raised erythrocyte sedimentation rate and raised CRP were all associated with increased risk of septic hip or osteomyelitis. The optimum inflammatory marker cut‐off was a CRP of 12 with a sensitivity of 87% and specificity of 91%. Conclusion: In acute non‐traumatic limp, X‐rays of the hips diagnose slipped upper femoral epiphysis, as such they should be routinely used from the age of 9 years upwards. Below this age they are of little value. Inflammatory markers have utility in risk‐stratifying children and selecting a group in whom to proceed with definitive tests to exclude osteomyelitis or septic hip. Children with a short history and minimal symptoms can be managed with appropriate follow up and no investigations.     

X‐ray Radiation Causes Electromagnetic Interference in Implantable Cardiac Pacemakers

Background: X‐rays are not thought to cause electromagnetic interference (EMI) in implantable cardiac pacemakers. However, x‐ray radiation during computed tomography (CT) scanning has been reported to cause EMI in some implantable cardiac pacemakers. The objectives of this study were to identify the location within the pacemakers where x‐ray radiation causes EMI and to investigate the association of EMI with the x‐ray radiation conditions. Methods: We verified the location where x‐ray radiation caused EMI using a CT scanner and conventional radiographic x‐ray equipment. An inhibition test and an asynchronous test were performed using five types of implantable cardiac pacemakers. Results: X‐ray radiation inhibited the pacing pulses of four types of implantable cardiac pacemakers when the body of each implantable cardiac pacemaker, containing a complementary metal‐oxide semiconductor (CMOS), was scanned using a CT scanner. We confirmed that x‐ray‐induced EMI depends on the x‐ray radiation conditions, that is, the tube voltage, tube current, x‐ray dose, and direction of x‐ray radiation, as well as the sensing thresholds of the implantable cardiac pacemakers. Conclusions: X‐ray radiation caused EMI in some implantable cardiac pacemakers, probably because the CMOS component was irradiated. The occurrence of EMI depended on the pacemaker model, sensing threshold of the pacemaker, and x‐ray radiation conditions. (PACE 2010; 33:1174–1181)     

β‐Cell subcellular localization of glucose‐stimulated Mn uptake by X‐ray fluorescence microscopy: implications for pancreatic MRI

Manganese (Mn) is a calcium (Ca) analog that has long been used as a magnetic resonance imaging (MRI) contrast agent for investigating cardiac tissue functionality, for brain mapping and for neuronal tract tracing studies. Recently, we have extended its use to investigate pancreatic β‐cells and showed that, in the presence of MnCl₂, glucose‐activated pancreatic islets yield significant signal enhancement in T₁‐weigheted MR images. In this study, we exploited for the first time the unique capabilities of X‐ray fluorescence microscopy (XFM) to both visualize and quantify the metal in pancreatic β‐cells at cellular and subcellular levels. MIN‐6 insulinoma cells grown in standard tissue culture conditions had only a trace amount of Mn, 1.14 ± 0.03 × 10^?11 µg/µm², homogenously distributed across the cell. Exposure to 2 m m glucose and 50 µ m MnCl₂ for 20 min resulted in nonglucose‐dependent Mn uptake and the overall cell concentration increased to 8.99 ± 2.69 × 10^?11 µg/µm². When cells were activated by incubation in 16 m m glucose in the presence of 50 µ m MnCl₂, a significant increase in cytoplasmic Mn was measured, reaching 2.57 ± 1.34 × 10^?10 µg/µm². A further rise in intracellular concentration was measured following KCl‐induced depolarization, with concentrations totaling 1.25 ± 0.33 × 10^?9 and 4.02 ± 0.71 × 10^?10 µg/µm² in the cytoplasm and nuclei, respectively. In both activated conditions Mn was prevalent in the cytoplasm and localized primarily in a perinuclear region, possibly corresponding to the Golgi apparatus and involving the secretory pathway. These data are consistent with our previous MRI findings, confirming that Mn can be used as a functional imaging reporter of pancreatic β‐cell activation and also provide a basis for understanding how subcellular localization of Mn will impact MRI contrast. Copyright © 2011 John Wiley & Sons, Ltd.     

64CuS‐labeled nanoparticles: a new sentinel‐lymph‐node‐mapping agent for PET–CT and photoacoustic tomography

Determining sentinel lymph node (SLN) status is critical to cancer staging and treatment decisions. Currently, in clinical practice, ^99mTc‐radiocolloid‐mediated planar scintigraphy and single‐photon emission computed tomography (SPECT) are used to guide the biopsy and resection of SLNs. Recently, an emerging technique that combines positron emission tomography (PET) and photoacoustic tomography (PAT; PET–PAT) may offer accurate information in detecting SLNs. Herein, we report a kind of ⁶⁴CuS‐labeled nanoparticle (⁶⁴CuS‐NP) for the detection of SLNs with PET–PAT. We subcutaneously injected ⁶⁴CuS‐NPs into the rats’ forepaw pads. After 24 h, the rats’ first draining axillary lymph nodes (i.e. the SLNs) could be clearly visualized with micro‐PET (μPET)–CT. Rats were sacrificed after μPET–CT imaging, their axillary lymph nodes were surgically identified, and then PAT was employed to discover ⁶⁴CuS‐NP‐avid SLNs, which were embedded inside tissues. Biodistribution, autoradiography, and copper staining analyses confirmed the SLNs’ high uptake of ⁶⁴CuS‐NPs. Our study indicates that ⁶⁴CuS‐NPs are a promising dual‐function agent for both PET–CT and PAT and could be used with multi‐modal imaging strategies such as PET–PAT to identify SLNs in a clinical setting. Copyright © 2016 John Wiley & Sons, Ltd.     

In‐vivo high‐resolution X‐ray microtomography for liver and spleen tumor assessment in mice

The present study sought to validate the use of glycery1‐2‐oley‐1,3‐bis‐[7‐(3‐amino‐2,4,6‐triiodophenyl)‐ heptanoate] (DHOG) contrast agent for mouse spleen tumor and liver metastasis imaging by high‐resolution X‐ray microtomography. Three groups of female nude mice were compared: controls (n = 5), and mice injected with 2.5 × 10⁶ STC1 tumor cells in the spleen, imaged at 15 days (group G15, n = 5) and at 30 days (group G30, n = 5, of which one died before imaging). Micro‐CT scans (X‐ray voltage, 50 kVp; anode current, 200 µA; exposure time, 632 ms; 180 rotational steps resulting in 35 µm isotropic spatial resolution) were acquired at 0, 0.75, 2 and 4 h after i.v. injection of DHOG. CT number (Hounsfield units: HU) and contrast‐to‐noise ratios (CNR) were determined in three organs. Statistical analysis was performed by Mann–Whitney U‐test. Contrast enhancement in normal spleen and liver increased, respectively to 1020 ± 159 and 351 ± 27 HU over baseline at 4 h, and 482 ± 3 and 203 ± 14 HU on day 6 after a single contrast injection. Automated three‐dimensional reconstruction and modeling of the spleen provided accurate and quantifiable images. Spleen tumor and liver metastases did not take up DHOG, making them detectable in contrast to the increased signal in normal tissue. The smallest liver metastasis detected measured 0.3 mm in diameter. High‐resolution X‐ray micro‐CT in living mice using DHOG contrast agent allowed visualization and volume quantification of normal spleen and of spleen tumor and its liver metastases. Copyright © 2007 John Wiley & Sons, Ltd.     

Serine‐ and mannose‐modified liposomal contrast agent for computed tomography: evaluation of the enhancement in rabbit liver VX‐2 tumor model

Kupffer cell imaging is a powerful tool for the detection of liver cancer. This diagnostic procedure depends on the faculty of the reticuloendothelial system (RES) which takes up foreign bodies, including small particles. The current study aimed to develop a novel RES targeting liposomal contrast agent that functionalized with serine or mannose, the moiety specifically binding to a corresponding receptor on phagocytic cells. Liposomes loaded with non‐ionic X‐ray contrast media, Iohexol, were prepared by supercritical carbon dioxide reverse‐phase evaporation method and were intravenously injected to healthy rabbits in order to evaluate the liver parenchymal enhancement in X‐ray computed tomography (CT). From 10 to 40 min after injection, the mean enhancement value of the liver parenchyma approached 45 and 34 Hounsfield units (HU) when serine‐modified iodinated liposomal contrast agent (ILCA) and mannose‐modified ILCA were applied, respectively. The tumor‐to‐liver contrast values were also evaluated after the administration of the prepared ILCA to rabbits with VX‐2 carcinoma. For serine‐modified ILCA, tumor‐to‐liver contrast was 82 HU at 1 min and >24 HU at 10–40 min; for mannose‐modified ILCA, the values were 58 HU at 0.5 min and >21 HU at 10–40 min. These vales estimated from the region of intrest and the imaging figures of liver indicate the potential of ILCA for clinical use. Copyright © 2010 John Wiley & Sons, Ltd.     

Multimodal‐3D imaging based on μMRI and μCT techniques bridges the gap with histology in visualization of the bone regeneration process

Bone repair/regeneration is usually investigated through X‐ray computed microtomography (μCT) supported by histology of extracted samples, to analyse biomaterial structure and new bone formation processes. Magnetic resonance imaging (μMRI) shows a richer tissue contrast than μCT, despite at lower resolution, and could be combined with μCT in the perspective of conducting non‐destructive 3D investigations of bone. A pipeline designed to combine μMRI and μCT images of bone samples is here described and applied on samples of extracted human jawbone core following bone graft. We optimized the coregistration procedure between μCT and μMRI images to avoid bias due to the different resolutions and contrasts. Furthermore, we used an Adaptive Multivariate Clustering, grouping homologous voxels in the coregistered images, to visualize different tissue types within a fused 3D metastructure. The tissue grouping matched the 2D histology applied only on 1 slice, thus extending the histology labelling in 3D. Specifically, in all samples, we could separate and map 2 types of regenerated bone, calcified tissue, soft tissues, and/or fat and marrow space. Remarkably, μMRI and μCT alone were not able to separate the 2 types of regenerated bone. Finally, we computed volumes of each tissue in the 3D metastructures, which might be exploited by quantitative simulation. The 3D metastructure obtained through our pipeline represents a first step to bridge the gap between the quality of information obtained from 2D optical microscopy and the 3D mapping of the bone tissue heterogeneity and could allow researchers and clinicians to non‐destructively characterize and follow‐up bone regeneration.     

Use of perfluorocarbon nanoparticles for non‐invasive multimodal cell tracking of human pancreatic islets

In vivo imaging of engraftment and immunorejection of transplanted islets is critical for further clinical development, with ¹H MR imaging of superparamagnetic iron oxide‐labeled cells being the current premier modality. Using perfluorocarbon nanoparticles, we present here a strategy for non‐invasive imaging of cells using other modalities. To this end, human cadaveric islets were labeled with rhodamine‐perfluorooctylbromide (PFOB) nanoparticles, rhodamine‐perfluoropolyether (PFPE) nanoparticles or Feridex® as control and tested in vitro for cell viability and c‐peptide secretion for 1 week. ¹⁹F MRI, computed tomography (CT) and ultrasound (US) imaging was performed on labeled cell phantoms and on cells following transplantation beneath the kidney capsule of mice and rabbits. PFOB and PFPE‐labeling did not reduce human islet viability or glucose responsiveness as compared with unlabeled cells or SPIO‐labeled cells. PFOB‐ and PFPE‐labeled islets were effectively fluorinated for visualization by ¹⁹F MRI. PFOB‐labeled islets were acoustically reflective for detection by US imaging and became sufficiently brominated to become radiopaque allowing visualization with CT. Thus, perfluorocarbon nanoparticles are multimodal cellular contrast agents that may find applications in real‐time targeted delivery and imaging of transplanted human islets or other cells in a clinically applicable manner using MRI, US or CT imaging. Copyright © 2011 John Wiley & Sons, Ltd.     

Reporter gene imaging using radiographic contrast from nonradioactive iodide sequestered by the sodium–iodide symporter

The hypothesis that the human sodium–iodide symporter, NIS, can be used to detect NIS expression using standard radiological techniques was tested using adenoviral transduced NIS expression in human tumor xenografts grown in mice and in a naive dog prostate. Nonradioactive iodide was administered systemically to animals that 1–3 days previously had received a local injection of a replication‐competent adenovirus expressing NIS under the control of the CMV promoter. The distribution of radiopacity was assessed in mouse tumors using micro‐CT and a clinical X‐ray machine and in the prostate of an anesthetized dog using a clinical spiral CT. Iodide sequestration and NIS expression were measured using X‐ray spectrochemical analysis and fluorescence microscopy, respectively. Radiographic contrast due to NIS gene expression that was observed indicates the technique has potential for use in preclinical rodent tumor studies but probably lacks sensitivity for human use. Copyright © 2007 John Wiley & Sons, Ltd.     

A novel Tc‐99 m and fluorescence labeled peptide as a multimodal imaging agent for targeting angiogenesis in a murine tumor model

The serine–aspartic acid–valine (SDV) peptide binds specifically to integrin α_Vβ₃. In the present study, we successfully developed a TAMRA–GHEG–ECG–SDV peptide labeled with both Tc‐99 m and TAMRA to target the integrin α_Vβ₃ of tumor cells; furthermore, we evaluated the diagnostic performance of Tc‐99 m TAMRA–GHEG–ECG–SDV as a dual‐modality imaging agent for tumor of the murine model. TAMRA–GHEG–ECG–SDV was synthesized using Fmoc solid‐phase peptide synthesis. Radiolabeling of TAMRA–GHEG–ECG–SDV with Tc‐99 m was done using ligand exchange methods. Labeling stability and cytotoxicity studies were performed. Gamma camera imaging, biodistribution and ex vivo imaging studies were performed in murine models with HT‐1080 and HT‐29 tumors. A tumor tissue slide was prepared and analyzed using confocal microscopy. After radiolabeling procedures with Tc‐99 m, the Tc‐99 m TAMRA–GHEG–ECG–SDV complexes were prepared in high yield (>99%). In the gamma camera imaging study, a substantial uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV into HT‐1080 tumor (integrin α_Vβ₃ positive) and low uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV into HT‐29 tumor (integrin α_Vβ₃ negative) were demonstrated. A competition study revealed that HT‐1080 tumor uptake was effectively blocked by the co‐injection of an excess concentration of SDV. Specific uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV was confirmed by biodistribution, ex vivo imaging and confocal microscopy studies. Our in vivo and in vitro studies revealed substantial uptake of Tc‐99 m TAMRA–GHEG–ECG–SDV in the integrin α_Vβ₃‐positive tumor. Tc‐99 m TAMRA–GHEG–ECG–SDV could be a good candidate for a dual‐modality imaging agent targeting tumor angiogenesis. Copyright © 2016 John Wiley & Sons, Ltd.     

Body composition in Italian and Danish women

The objective of this cross‐sectional study was to compare the body composition and fat distribution measured by dual energy X‐ray absorptiometry (DPX, Lunar) in different age decades of age‐matched Danish and Italian women. The subjects comprised 133 healthy Italian women (age 20–60 years) age‐matched to a representative sub‐sample of healthy Danish women (n=375). Total and abdominal body fat tissue mass were measured by dual‐energy X‐ray absorptiometry. Italian women were shorter and fatter compared with age‐matched Danish women, but in middle‐age, had a less abdominal fat distribution. There was no difference in total body bone mineral density.     

Biodistribution and evaluation of 131I‐labeled neuropilin‐binding peptide for targeted tumor imaging

Neuropilin‐1 (NRP‐1) is overexpressed in several kinds of cancer cell and contributes to tumor aggressiveness. Recently, the arginine/lysine‐rich peptide with C‐terminal motifs (R/K)XX(R/K) indicated promising penetrating and transporting capability into NRP‐1 positive cancer cells. In the present study, we describe a ¹³¹I‐labeled C‐end rule motif peptide conjugate, Tyr–tLyp‐1, for NRP‐1 positive tumor targeting and imaging properties. Briefly, a truncated Lyp‐1 peptide was designed to expose its C‐end motif and conjugated to tyrosine for radiolabeling after structural modification. The peptide indicated specific binding to A549 cancer cells at 2 μM concentration, and its binding was dependent on NRP‐1 expression and could be inhibited by other NRP‐1‐binding peptides. In vivo imaging of ¹³¹I‐labeled Tyr–tLyp‐1peptide showed that a subcutaneous A549 xenograft tumor could be visualized using a SPECT/CT scanner. The tumor uptake of ¹³¹I‐Tyr–tLyp‐1 was 4.77 times higher than the uptake in muscles by SPECT/CT software quantification at 6 h post injection. Together, this study indicated that truncated Lyp‐1 peptide could specifically localize in NRP‐1 positive tumors and successfully mediate the ¹³¹I radionuclide diagnosis, indicating promising targeted imaging capability for NRP‐1 positive tumors. Copyright © 2016 John Wiley & Sons, Ltd.     

Magnetite loaded Polypeptide‐PLGA multifunctional microbubbles for dual‐mode US/MR imaging

Magnetite loaded Polypeptide‐PLGA multifunctional microbubbles (Fe₃O₄/Polypeptide‐PLGA MMBs) that show superparamagnetic properties were prepared by a modified double emulsion method and employed as imaging agent for dual‐mode Ultrasound/Magnetic resonance (US/MR) imaging of prostatic cancer. The successful synthesis of MMBs was determined by Fourier Transform Infrared Spectrometer (FTIR), X‐ray diffraction (XRD), Transmission Electron Microscope (TEM), Scanning Electron Microscope (SEM), Atomic Absorption Spectroscopy (AAS) and vibrating sample magnetometer (VSM). The as‐prepared MMBs had a diameter of 700 nm and were quite safe as confirmed by MTT assays. Prussian Blue Staining showed that targeted Fe₃O₄/Polypeptide‐PLGA MMBs enhanced the cellular uptake efficiency. In cell attachment study, adherence of MMBs was significantly higher to LNCaP cells compared with negative control PC3 cells. The in vitro results demonstrated that these MMBs could enhance both US and MR imaging of prostatic cancer. Copyright © 2015 John Wiley & Sons, Ltd.     

Influence of different iodinated contrast media on the induction of DNA double‐strand breaks after in vitro X‐ray irradiation

The objective of this work was to examine differences in DNA double‐strand break induction in peripheral blood lymphocytes after in vitro X‐ray irradiation between iodinated contrast agents. Four different iodinated X‐ray contrast agents – three of them with two different iodine concentrations – and mannitol (negative control; concentration of 150 mg mannitol per ml blood) were pipetted into blood samples so that there was a concentration of 0, 7.5 or 15 mg of iodine per ml blood in the samples. Negative controls without contrast medium (0 mg of iodine per ml blood) were also processed for every irradiation dose. The tubes were exposed to 0, 20 or 500 mGy in vitro X‐ray irradiation. After that, the lymphocytes were separated by using density‐gradient centrifugation. Fluorescence microscopy was applied to determine the average number of γH2AX‐foci per lymphocyte in the presence or absence of different contrast media or mannitol. Differences in the number of γH2AX‐foci were statistically analysed by one‐way ANOVA and post‐hoc Tukey's honestly significant difference test. Iodinated contrast agents led to a statistically significant increase in DNA double‐strand breaks after in vitro irradiation. This effect increased statistically significant with rising radiation dose and appeared independent of the contrast agent used (iopromid, iodixanol, iomeprol, iopamidol). A statistically significant difference in DNA damage between the different tested contrast agents was not found. Therefore, the increase in DNA double‐strand breaks depends solely on the amount of iodine applied. For evaluation of clinical consequences, our findings could be tested in further animal studies. Copyright © 2014 John Wiley & Sons, Ltd.     

Sensitivity‐enhanced chemical exchange saturation transfer (CEST) MRI with least squares optimization of Carr Purcell Meiboom Gill multi‐echo echo planar imaging

Chemical exchange saturation transfer (CEST) imaging is a novel MRI technique that is sensitive to biomolecules, local pH and temperature, and offers considerable advantages for in vivo applications. However, the magnitude of CEST effect for dilute CEST agents undergoing slow or intermediate chemical exchange is typically small, requiring the use of signal averaging to enhance its sensitivity. Given that T₂‐induced signal loss can be normalized by asymmetry analysis, the magnitude of CEST effect is independent of echo time. Therefore, CEST MRI with multi‐echo echo planar imaging (EPI) readout should yield the same CEST effect as conventional single echo acquisition. Importantly, CEST multi‐echo (CESTme) EPI images can be averaged to enhance CEST MRI sensitivity. The goal of this study was to validate CESTme EPI using a creatine–agarose gel CEST phantom with similar T₂ as biological tissue. Using least‐squares optimization, we found that the sensitivity of CESTme sequence was significantly higher than that obtained by conventional single echo CEST‐EPI acquisition. Specifically, signal‐to‐noise ratio and contrast‐to‐noise ratio from the proposed CESTme EPI were approximately equivalent to that obtained by doubling the number of signal averages of the standard single echo CEST MRI sequence. In summary, our results demonstrated CESTme EPI for sensitivity‐enhanced CEST imaging. Copyright © 2014 John Wiley & Sons, Ltd.     

Functional association of phosphoinositide‐3‐kinase with platelet glycoprotein Ibα, the major ligand‐binding subunit of the glycoprotein Ib–IX–V complex

Summary. Background: The adhesion receptor glycoprotein (GP)Ib–IX–V, which binds von Willebrand factor (VWF) and other ligands, initiates platelet activation and thrombus formation at arterial shear rates, and may control other vascular processes, such as coagulation, inflammation, and platelet‐mediated tumor metastasis. The cytoplasmic C‐terminal domain of the ligand‐binding GPIbα subunit contains binding sites for filamin (residues 561–572, critically Phe568/Trp570), 14‐3‐3ζ (involving phosphorylation sites Ser587/590 and Ser609), and the phosphoinositide‐3‐kinase (PI3‐kinase) regulatory subunit, p85. Objectives: We previously showed that, as compared with wild‐type receptor, deleting the contiguous sequence 580–590 or 591–610, but not upstream sequences, of GPIbα expressed as a GPIb–IX complex in Chinese hamster ovary cells inhibited VWF‐dependent Akt phosphorylation, which is used as a read‐out for PI3‐kinase activity. Pulldown experiments using glutathione‐S‐transferase (GST)–p85 or GST–14‐3‐3ζ constructs, and competitive inhibitors of 14‐3‐3ζ binding, suggested an independent association of 14‐3‐3ζ and PI3‐kinase with GPIbα. The objective of this study was to analyze a further panel of GPIbα deletion mutations within residues 580–610. Results: We identified a novel deletion mutant, Δ591–595, that uniquely disrupts 14‐3‐3ζ binding but retains the functional p85/PI3‐kinase association. Deletion of other sequences within the 580–610 region were less discriminatory, and either partially affected p85/PI3‐kinase and 14‐3‐3ζ binding (Δ580–585, Δ586–590, Δ596–600, Δ601–605), or strongly inhibited binding of both proteins (Δ606–610). Conclusions: Together, these findings have significant implications for interpreting the functional role of p85 and/or 14‐3‐3ζ in GPIb‐dependent signaling or platelet functional studies involving truncation of the C‐terminal residues in cell‐based assays and mouse models. The Δ591–595 mutation provides another strategy for determining the function of GPIbα‐associated 14‐3‐3ζ by selective disruption of 14‐3‐3ζ but not p85/PI3‐kinase binding.     

Additional value of B‐flow imaging in arterial wall calcifications

The accuracy of color Doppler ultrasonography (CDU) is hampered by the presence of arterial wall calcifications. B‐flow imaging may overcome these limitations. We present a case of a severely calcified stenosis of the right common femoral artery (CFA) diagnosed with the aid of B‐flow imaging. Both the CT angiography scan and CDU were limited by the presence of diffuse dense arterial calcifications. B‐flow imaging showed a >75% stenosis of the CFA. B‐flow imaging appears to improve the accuracy of CDU in the presence of calcified stenosis of the CFA. It is of clinical relevance to improve the duplex sonographic accuracy, ideally reducing the need for other imaging modalities prior to surgery. © 2017 Wiley Periodicals, Inc. J Clin Ultrasound 46 :136–139, 2018