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The synthesis and characterization of a gadolinium-based contrast agent conjugated to 3-phosphoglycerate (Gd-3PG) are reported. The synthetic steps are optimized to incorporate a selective deprotection strategy for a primary tert-butyl dimethyl silyl (TBS) hydroxyl over a secondary one. The relaxivity of Gd-3PG shows characteristic improvement, likely due to secondary sphere effects and/or an increase in molecular weight (5.39 ± 0.14 mM−1 s−1 at 1.4 T and 37 °C). Michaelis–Menten enzymatic kinetics was measured on the modified 3PG-arm and it showed similar activity to the native 3PG metabolite, Km = 240 ± 30 μM. This agent and future versions of this type of GBCA, which are conjugated to glycolytic metabolites, were designed to monitor in vivo allosteric regulatory events in glycolytic processes. [ABSTRACT FROM AUTHOR]

Senescent cells play a vital role in the pathogenesis of musculoskeletal (MSK) diseases, such as chronic inflammatory joint disorders, rheumatoid arthritis (RA), and osteoarthritis (OA). Cellular senescence in articular joints represents a response of local cells to persistent stress that leads to cell-cycle arrest and enhanced production of inflammatory cytokines, which in turn perpetuates joint damage and leads to significant morbidities in afflicted patients. It has been recently discovered that clearance of senescent cells by novel "senolytic" therapies can attenuate the chronic inflammatory microenvironment of RA and OA, preventing further disease progression and supporting healing processes. To identify patients who might benefit from these new senolytic therapies and monitor therapy response, there is an unmet need to identify and map senescent cells in articular joints and related musculoskeletal tissues. To fill this gap, new imaging biomarkers are being developed to detect and characterize senescent cells in human joints and musculoskeletal tissues. This review article will provide an overview of these efforts. New imaging biomarkers for senescence cells are expected to significantly improve the specificity of state-of-the-art imaging technologies for diagnosing musculoskeletal disorders. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) play key roles in diverse biological processes, transport biomolecules between cells and have been engineered for therapeutic applications. A useful EV bioengineering strategy is to express engineered proteins on the EV surface to confer targeting, bioactivity and other properties. Measuring how incorporation varies across a population of EVs is important for characterising such materials and understanding their function, yet it remains challenging to quantitatively characterise the absolute number of engineered proteins incorporated at single‐EV resolution. To address these needs, we developed a HaloTag‐based characterisation platform in which dyes or other synthetic species can be covalently and stoichiometrically attached to engineered proteins on the EV surface. To evaluate this system, we employed several orthogonal quantification methods, including flow cytometry and fluorescence microscopy, and found that HaloTag‐mediated quantification is generally robust across EV analysis methods. We compared HaloTag‐labelling to antibody‐labelling of EVs using single vesicle flow cytometry, enabling us to measure the substantial degree to which antibody labelling can underestimate proteins present on an EV. Finally, we demonstrate the use of HaloTag to compare between protein designs for EV bioengineering. Overall, the HaloTag system is a useful EV characterisation tool which complements and expands existing methods. [ABSTRACT FROM AUTHOR]

Basal Cell Carcinoma (BCC) is the most commonly diagnosed cancer worldwide. While the survivability of BCC is high, many patients are excluded from clinically available treatments due to health risks or personal choice. Further, patients with advanced or metastatic disease have severely limited treatment options. The dysregulation of the Hedgehog (Hh) signaling cascade drives onset and progression of BCC. As such, the modulation of this pathway has driven advancements in BCC research. In this review, we focus firstly on inhibitors that target the Hh pathway as chemotherapeutics against BCC. Two therapies targeting Hh signaling have been made clinically available for BCC patients, but these treatments suffer from limited initial efficacy and a high rate of chemoresistant tumor recurrence. Herein, we describe more recent developments of chemical scaffolds that have been designed to hopefully improve upon the available therapeutics. We secondly discuss the history and recent efforts involving modulation of the Hh genome as a method of producing in vivo models of BCC for preclinical research. While there are many advancements left to be made towards improving patient outcomes with BCC, it is clear that targeting the Hh pathway will remain at the forefront of research efforts in designing more effective chemotherapeutics as well as relevant preclinical models. [ABSTRACT FROM AUTHOR]

Our lab has developed a new series of self‐immolative MR agents for the rapid detection of enzyme activity in mouse models expressing β‐galactosidase (β‐gal). We investigated two molecular architectures to create agents that detect β‐gal activity by modulating the coordination of water to GdIII. The first is an intermolecular approach, wherein we designed several structural isomers to maximize coordination of endogenous carbonate ions. The second involves an intramolecular mechanism for q modulation. We incorporated a pendant coordinating carboxylate ligand with a 2, 4, 6, or 8 carbon linker to saturate ligand coordination to the GdIII ion. This renders the agent ineffective. We show that one agent in particular (6‐C pendant carboxylate) is an extremely effective MR reporter for the detection of enzyme activity in a mouse model expressing β‐gal. [ABSTRACT FROM AUTHOR]

Our lab has developed a new series of self‐immolative MR agents for the rapid detection of enzyme activity in mouse models expressing β‐galactosidase (β‐gal). We investigated two molecular architectures to create agents that detect β‐gal activity by modulating the coordination of water to GdIII. The first is an intermolecular approach, wherein we designed several structural isomers to maximize coordination of endogenous carbonate ions. The second involves an intramolecular mechanism for q modulation. We incorporated a pendant coordinating carboxylate ligand with a 2, 4, 6, or 8 carbon linker to saturate ligand coordination to the GdIII ion. This renders the agent ineffective. We show that one agent in particular (6‐C pendant carboxylate) is an extremely effective MR reporter for the detection of enzyme activity in a mouse model expressing β‐gal. [ABSTRACT FROM AUTHOR]

Muscle stem cells are a potent cell population dedicated to efficacious skeletal muscle regeneration, but their therapeutic utility is currently limited by mode of delivery. We developed a cell delivery strategy based on a supramolecular liquid crystal formed by peptide amphiphiles (PAs) that encapsulates cells and growth factors within a muscle-like unidirectionally ordered environment of nanofibers. The stiffness of the PA scaffolds, dependent on amino acid sequence, was found to determine the macroscopic degree of cell alignment templated by the nanofibers in vitro. Furthermore, these PA scaffolds support myogenic progenitor cell survival and proliferation and they can be optimized to induce cell differentiation and maturation. We engineered an in vivo delivery system to assemble scaffolds by injection of a PA solution that enabled coalignment of scaffold nanofibers with endogenous myofibers. These scaffolds locally retained growth factors, displayed degradation rates matching the time course of muscle tissue regeneration, and markedly enhanced the engraftment of muscle stem cells in injured and noninjured muscles in mice. [ABSTRACT FROM AUTHOR]

Purpose To demonstrate a new MR imaging approach that unambiguously identifies and quantitates contrast agents based on intrinsic agent properties such as r1, r2 [ABSTRACT FROM AUTHOR]

Detection of protein expression by MRI requires a high payload of Gd(iii) per protein binding event. Presented here is a targeted AuDNA nanoparticle capable of delivering several hundred Gd(iii) chelates to the HaloTag reporter protein. Incubating this particle with HaloTag-expressing cells produced a 9.4 contrast-to-noise ratio compared to non-expressing cells. [ABSTRACT FROM AUTHOR]

Magnetic resonance (MR) imaging is advantageous because it concurrently provides anatomic, functional, and molecular information. MR molecular imaging can combine the high spatial resolution of this established clinical modality with molecular profiling in vivo. However, as a result of the intrinsically low sensitivity of MR imaging, high local concentrations of biological targets are required to generate discernable MR contrast. We hypothesize that the prostate-specific membrane antigen (PSMA), an attractive target for imaging and therapy of prostate cancer, could serve as a suitable biomarker for MR-based molecular imaging. We have synthesized three new high-affinity, low-molecular-weight GdIII-based PSMA-targeted contrast agents containing one to three GdIII chelates per molecule. We evaluated the relaxometric properties of these agents in solution, in prostate cancer cells, and in an in vivo experimental model to demonstrate the feasibility of PSMA-based MR molecular imaging. [ABSTRACT FROM AUTHOR]

Magnetic resonance (MR) imaging is advantageous because it concurrently provides anatomic, functional, and molecular information. MR molecular imaging can combine the high spatial resolution of this established clinical modality with molecular profiling in vivo. However, as a result of the intrinsically low sensitivity of MR imaging, high local concentrations of biological targets are required to generate discernable MR contrast. We hypothesize that the prostate-specific membrane antigen (PSMA), an attractive target for imaging and therapy of prostate cancer, could serve as a suitable biomarker for MR-based molecular imaging. We have synthesized three new high-affinity, low-molecular-weight GdIII-based PSMA-targeted contrast agents containing one to three GdIII chelates per molecule. We evaluated the relaxometric properties of these agents in solution, in prostate cancer cells, and in an in vivo experimental model to demonstrate the feasibility of PSMA-based MR molecular imaging. [ABSTRACT FROM AUTHOR]

Transcription factors are key regulators in both normal and pathological cell processes. Affecting the activity of these proteins is a promising strategy for understanding gene regulation and developing effective therapeutics. CoIII Schiff base complexes ([Co(acacen)(L)2]+ where L=labile axial ligands) have been shown to be potent inhibitors of a number of zinc metalloproteins including Cys2His2 zinc finger transcription factors. Inhibition by [Co(acacen)(L)2]+ of the target protein is believed to occur through a dissociative exchange of the labile axial ligands for histidine (His) residues essential for function. Here, we report a series of spectroscopic investigations with model peptides of zinc fingers that elucidate the interaction between [Co(acacen)(L)2]+ complexes and zinc finger transcription factors. Observed changes in NMR chemical shifts and 2D 1H-1H NOESY NMR spectra demonstrate the preference of [Co(acacen)(L)2]+ complexes to coordinate His residues over other amino acids. The conformation of [Co(acacen)(L)2]+ upon His coordination was characterized by 1H NMR spectroscopy, near-UV CD, and electronic absorption. These studies reveal that the resulting His-coordinated [Co(acacen)(L)2]+ complex possesses an octahedral structure. The effects of [Co(acacen)(L)2]+ complexes on the zinc-finger structure were assessed by the degree of hydrogen bonding (probed by 2D NMR spectroscopy) and secondary-structure profiles measured by far-UV CD. These structural studies demonstrate the ability of [Co(acacen)(L)2]+ complexes to disrupt the ββα structure of zinc fingers, resulting in primarily random-coil conformations. A mechanism is described wherein [Co(acacen)(L)2]+ complexes inhibit zinc finger transcription factor activity through selectively coordinating His residues in the zinc finger by dissociative ligand exchange and disrupting the ββα structural motif required for gene regulation. [ABSTRACT FROM AUTHOR]

Purpose: To minimize feature loss in T1- and T2-weighted MRI by merging multiple MR images acquired at different TR and TE to generate an image with increased dynamic range. Materials and Methods: High Dynamic Range (HDR) processing techniques from the field of photography were applied to a series of acquired MR images. Specifically, a method to parameterize the algorithm for MRI data was developed and tested. T1- and T2-weighted images of a number of contrast agent phantoms and a live mouse were acquired with varying TR and TE parameters. The images were computationally merged to produce HDR-MR images. All acquisitions were performed on a 7.05 T Bruker PharmaScan with a multi-echo spin echo pulse sequence. Results: HDR-MRI delineated bright and dark features that were either saturated or indistinguishable from background in standard T1- and T2-weighted MRI. The increased dynamic range preserved intensity gradation over a larger range of T1 and T2 in phantoms and revealed more anatomical features in vivo. Conclusions: We have developed and tested a method to apply HDR processing to MR images. The increased dynamic range of HDR-MR images as compared to standard T1- and T2-weighted images minimizes feature loss caused by magnetization recovery or low SNR. [ABSTRACT FROM AUTHOR]

A polymer‐caged nanobin (PCN) theranostic platform with a doxorubicin (DXR)‐loaded liposomal core and an acid‐sensitive polymer shell that is functionalized with herceptin and GdIII‐based magnetic resonance imaging (MRI) contrast agents is reported. In vitro testing reveals an 120‐fold improvement in cellular GdIII uptake in comparison with clinically approved DOTA‐GdIII, leading to significant T1 MRI contrast enhancement. [ABSTRACT FROM AUTHOR]

Interspin distances between 0.8 nm and 2.0 nm can be measured through the dipolar broadening of the continuous wave (cw) EPR spectrum of nitroxide spin labels at X-band (9.4 GHz, 0.35 T). We introduce Gd3+ as a promising alternative spin label for distance measurements by cw EPR above 7 Tesla, where the ☋−1/2〉 to ☋1/2〉 transition narrows below 1 mT and becomes extremely sensitive to dipolar broadening. To estimate the distance limits of cw EPR with Gd3+, we have measured spectra of frozen solutions of GdCl3 at 8.6 T (240 GHz) and 10 K at concentrations ranging from 50 mM to 0.1 mM, covering a range of average interspin distances. These experiments show substantial dipolar broadening at distances where line broadening cannot be observed with nitroxides at X-band. This data, and its agreement with calculated dipolar-broadened lineshapes, show Gd3+ to be sensitive to distances as long as ∼3.8 nm. Further, the linewidth of a bis-Gd3+ complex with a flexible ∼1.6 nm bridge is strongly broadened as compared to the mono-Gd3+ complex, demonstrating the potential for application to pairwise distances. Gd-DOTA-based chelates that can be functionalized to protein surfaces display linewidths narrower than aqueous GdCl3, implying they should be even more sensitive to dipolar broadening. Therefore, we suggest that the combination of tailored Gd3+ labels and high magnetic fields can extend the longest interspin distances measurable by cw EPR from 2.0 nm to 3.8 nm. cw EPR data at 260 K demonstrate that the line broadening remains clear out to similar average interspin distances, offering Gd3+ probes as promising distance rulers at temperatures higher than possible with conventional pulsed EPR distance measurements. [ABSTRACT FROM AUTHOR]

Objectives Patients with rheumatoid arthritis (RA) have a reduced life expectancy due to increased cardiovascular disease. The lack of a suitable animal model resembling both RA and atherosclerosis has hindered studies demonstrating a direct link between systemic infl ammation in RA and the development of atherosclerosis. Our objective was to overcome this barrier by generating an animal model (K/BxAg7) that spontaneously develops both RA-like disease and atherosclerosis. Methods Arthritis severity was evaluated using clinical indices and immunohistochemical staining of ankle joint specimens. Aortic atherosclerosis was delineated via Sudan IV staining and immunohistochemical analysis. Serum cholesterol and lipoprotein levels were measured using enzymatic assays. Serum levels of cytokines, chemokines and adipokines were determined by Luminex assays. Results K/BxAg7 mice developed a destructive arthropathy followed by prominent aortic atherosclerosis. These animals also displayed dyslipidaemia, characterised by reduced serum levels of total cholesterol and highdensity lipoprotein, and increased low-density lipoprotein (LDL)/vLDL compared with control mice. Further, there were higher levels of circulating infl ammatory mediators, such as interleukin-6, sRANKL and CCL5 in atherosclerotic K/BxAg7 mice compared with controls. Treatment with etanercept reduced arthritis and atherosclerosis development in K/BxAg7 mice. Conclusions K/BxAg7 mice recapitulate the same sequence of events occurring in patients with RA, namely an erosive, infl ammatory arthritis followed by atherosclerosis. These data suggest that the K/BxAg7 mouse is a novel system for investigating the interplay between systemic infl ammation occurring in RA and the development of atherosclerosis. [ABSTRACT FROM AUTHOR]

Snail family proteins are core EMT (epithelial-mesenchymal transition) regulatory factors that play essential roles in both development and disease processes and have been associated with metastasis in carcinomas. Snail factors are required for the formation of neural crest stem cells in most vertebrate embryos, as well as for the migratory invasive behavior of these cells. Snail factors have recently been linked to the formation of cancer stem cells, and expression of Snail proteins may be associated with tumor recurrence and resistance to chemotherapy and radiotherapy. We report that Co(III)-Ebox is a potent inhibitor of Snail- mediated transcriptional repression in breast cancer cells and in the neural crest of Xenopus. We further show that the activity of Co(III)-Ebox can be modulated by temperature, increasing the utility of this conjugate as a Snail inhibitor in model organisms. We exploit this feature to further delineate the requirements for Snail function during neural crest development, showing that in addition to the roles that Snail factors play in neural crest precursor formation and neural crest EMT/migration, inhibition of Snail function after the onset of neural crest migration leads to a loss of neural crest derived melanocytes. Co(III)-Ebox-mediated inhibition therefore provides a powerful tool for analysing the function of these core EMT factors with unparalleled temporal resolution. Moreover, the potency of Co(III)-Ebox as a Snail inhibitor in breast cancer cells suggests its potential as a therapeutic inhibitor of tumor progression and metastasis. [ABSTRACT FROM AUTHOR]

We report the first examples of amine-functionalized K2[OsII(bpy)(CN)4] (bpy = 2,2′-bipyridine) complexes. The tetracyanoosmate complexes were prepared by UV irradiation (λ = 254 nm) of K4[OsII(CN)6] and primary amine-functionalized bpy ligands in acidic aqueous media. The aqueous solution pH dependences of the spectroscopic and redox properties of 4,4′- and 5,5′-substituted complexes have been investigated. The pendant amine functional groups and coordinated cyanide ligands are basic sites that can be sequentially protonated, thereby allowing systematic tuning of electrochemical and optical spectroscopic properties. [ABSTRACT FROM AUTHOR]

The article discusses a study on the role of magnetic resonance imaging (MRI) in synthesizing ferritin-coated polystyrene beads. The researchers examined particles that are useful to study the effects of ferritin distribution, particle size and bead fraction on proton relaxation. Study revealed that ferritin-coated polystyrene beads were synthesized and characterized as long-term synthetic ferritin or hemosiderin mimics.

Despite recent advances in tissue engineering to regenerate biological function by combining cells with material supports, development is hindered by inadequate techniques for characterizing biomaterials in vivo. Magnetic resonance imaging is a tomographic technique with high temporal and spatial resolution and represents an excellent imaging modality for longitudinal noninvasive assessment of biomaterials in vivo. To distinguish biomaterials from surrounding tissues for magnetic resonance imaging, protein polymer contrast agents were developed and incorporated into hydrogels. In vitro and in vivo images of protein polymer hydrogels, with and without covalently incorporated protein polymer contrast agents, were acquired by magnetic resonance imaging. T1 values of the labeled gels were consistently lower when protein polymer contrast agents were included. As a result, the protein polymer contrast agent hydrogels facilitated fate tracking, quantification of degradation, and detection of immune response in vivo. For the duration of the in vivo study, the protein polymer contrast agent-containing hydrogels could be distinguished from adjacent tissues and from the foreign body response surrounding the gels. The hydrogels containing protein polymer contrast agent have a contrast-to-noise ratio 2-fold greater than hydrogels without protein polymer contrast agent. In the absence of the protein polymer contrast agent, hydrogels cannot be distinguished by the end of the gel lifetime. Magn Reson Med, 2010. © 2010 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

A chiral porphyrazine (pz), H2[pz(trans-A2B2)) (247), has been prepared that exhibits preferential in vivo accumulation inthe cells of tumors. Pz 247 exhibits near-infrared (NIR) emission with λ > 700 nm in the required wavelength range for maximum tissue penetration. When MDA-MB-231 breast tumor cells are treated with 247, the agent shows strong intracellular fluorescence with an emission maximum, 704 nm, which indicates that it localizes within a hydrophobic microenvironment. Pz 247 is shown to associate with the lipophilic core of LDL and undergo cellular entry primarily through receptor-mediated endocytosis accumulating in lysosomes. Preliminary in vivo studies show that 247 exhibits preferential accumulation and retention in the cells of MDA-MB231 tumors subcutaneously implanted in mice, thereby enabling NIR optical imaging with excellent contrast between tumor and surrounding tissue. The intensity of fluorescence from 247 within the tumor increases over time up to 48 h after injection presumably due to the sequestration of circulating 247/LDL complex by the tumor tissue. As the need for cholesterol, and thus LDL, is elevated in highly proliferative tumor cells over nontumorigenic cells, 247 has potential application for all such tumors. [ABSTRACT FROM AUTHOR]

A transition metal complex targeted for the inhibition of a subset of zinc finger transcription factors has been synthesized and tested in Xenopus Iaevis. A Co(lll) Schiff base complex modified with a 17-bp DNA sequence is designed to selectively inhibit Snail family transcription factors. The oligonucleotide-conjugated Co(lll) complex prevents Slug, Snail, and Sipi from binding their DNA targets whereas other transcription factors are still able to interact with their target DNA. The attachment of the oligonucleotide to the Co(lll) complex increases specificity 150-fold over the unconjugated complex. Studies demonstrate that neither the oligo, or the Co(lll) Schiff base complex alone, are sufficient for inactivation of Slug at concentrations that the conjugated complex mediates inhibition. Slug, Snail, and Sipi have been implicated in the regulation of epithelial-to-mesenchymal transition in development and cancer. A complex targeted to inactivate their transcriptional activity could prove valuable as an experimental tool and a cancer therapeutic. [ABSTRACT FROM AUTHOR]

We report the synthesis and characterization of polyvinyl alcohol (PVA) embolic particles modified with a clinically approved magnetic resonance (MR) contrast agent. PVA particles are used during transcatheter arterial embolization (TAE) procedures and this minimally invasive technique is a widely employed treatment for inoperable tumors. The PVA particles are injected into tumor vessels and prevent blood flow which results in tumor attenuation. An accurate assessment of the endpoint of embolization is critical to successful TAE procedures. Recent reports suggest that 20% of endpoint determination of TAE procedures by angiographic techniques are erroneous. Real time, in vivo imaging of the embolic particles would overcome this limitation. The contrast-modified PVA particles described here show an 80% decrease in T1 relaxation times compared to unmodified particles. Images of particles in capillary tubes of similar size to catheters used in TAE procedures are clearly visible by MRI. Magn Reson Med 59:898-902, 2008. © 2008 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Zinc(II) plays a vital role in normal cellular function as an essential component of numerous enzymes, transcription factors, and synaptic vesicles. While zinc can be linked to a variety of physiological processes, the mechanisms of its cellular actions are less discernible. Here, we have synthesized and tested a Zn(II)-activated magnetic resonance imaging (MRI) contrast agent in which the coordination geometry of the complex rearranges upon binding of Zn(II). In the absence of Zn(II) water is restricted from binding to a chelated Gd(III) ion by coordinating acetate arms resulting in a low relaxivity of 2.33 mM-1·s-1 at 60 MHz. Upon addition of Zn(II) the relaxivity of the Gd(III)-Zn(II) complex increases to 5.07 mM-1·s-1 and is consistent with one water molecule bound to Gd(III). These results were confirmed by nuclear magnetic relaxation dispersion analysis. There was no observed change in relaxivity of the Gd(III) complex when physiologically competing cations Ca(II) and Mg(II) were added. A competitive binding assay gave a dissociation constant of 2.38 × 10-4 M for the Gd(III)-Zn(II) complex. In vitro magnetic resonance images confirm that Zn(II) concentrations as low as 100 µM can be detected by using this contrast agent. [ABSTRACT FROM AUTHOR]

The in vivo monitoring of cell survival and migration will be essential to the translation of cell-based therapies from the laboratory to clinical studies. The pre-labeling of cells with magnetic resonance imaging (MRI) contrast agents renders them visible in vivo for serial cellular imaging. However, little is known about the impact of the presence of these metal particles inside transplanted cells. The use of the bimodal contrast agent GRID made it possible to demonstrate by means of fluorescent microscopy and inductively coupled plasma mass spectrometry (ICP-MS) that, after 16 h of incubation (without the use of a transfection agent), neural stem cells (NSCs) were saturated and no longer incorporated particles. With this maximal uptake, no significant effect on cell viability was observed. However, a significant decrease in proliferation was evident in cells that underwent 24 h of labeling. A significant increase in reactive oxygen species was observed for all GRID labeling, with a very significant increase with 24 h of labeling. GRID labeling did not affect cell motility in comparison with PKH26-labeled NSCs in a glioma-based migration assay and also allowed differentiation into all major cell types of the brain. GRID-labeled cells induced a signal change of 47% on T2 measurements and allows a detection of cell clusters of ∼220 cells/µl. Further in vivo testing will be required to ensure that cell labeling with gadolinium-based MRI contrast agents does not impair their ability to repair. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The inability to transduce cellular membranes is a limitation of current magnetic resonance imaging probes used in biologic and clinical settings. This constraint confines contrast agents to extracellular and vascular regions of the body, drastically reducing their viability for investigating processes and cycles in developmental biology. Conversely, a contrast agent with the ability to permeate cell membranes could be used in visualizing cell patterning, cell fate mapping, gene therapy, and, eventually, noninvasive cancer diagnosis. Therefore, we describe the synthesis and quantitative imaging of four contrast agents with the capability to cross cell membranes in sufficient quantity for detection. Each agent is based on the conjugation of a Gd(III) chelator with a cellular transduction moiety. Specifically, we coupled Gd(III)-diethylenetriaminepentaacetic acid DTPA and Gd(III)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid with an 8-amino acid polyarginine oligomer and an amphipathic stilbene molecule, 4-amino-4′-(N,N-dimethylamino) stilbene. The imaging modality that provided the best sensitivity and spatial resolution for direct detection of the contrast agents is synchrotron radiation x-ray fluorescence (SR-XRF). Unlike optical microscopy, SR-XRF provides two-dimensional images with resolution 10³ better than 153Gd gamma counting, without altering the agent by organic fluorophore conjugation. The transduction efficiency of the intracellular agents was evaluated by T1 analysis and inductively coupled plasma mass spectrometry to determine the efficacy of each chelate-transporter combination. [ABSTRACT FROM AUTHOR]

High-resolution in vivo imaging of gene expression is not possible in opaque animals by existing techniques. Here we present a new approach for obtaining such images by magnetic resonance imaging (MRI) using an MRI contrast agent that can indicate reporter gene expression in living animals. We have prepared MRI contrast agents in which the access of water to the first coordination sphere of a chelated paramagnetic ion is blocked with a substrate that can be removed by enzymatic cleavage. Following cleavage, the paramagnetic ion can interact directly with water protons to increase the MR signal. Here, we report an agent where galactopyranose is the blocking group. This group renders the MRI contrast agent sensitive to expression of the commonly used marker gene,b-galactosidase. To cellular resolution, regions of higher intensity in the MR image correlate with regions expressing marker enzyme. These results offer the promise of in vivo mapping of gene expression in transgenic animals and validate a general approach for constructing a family of MRI contrast agents that respond to biological activity. [ABSTRACT FROM AUTHOR]

Focuses on a study which examined the impact of the metmyoglobin on the Schiff-base complex `Co(acetylacetonate-ethlylenediimine)(NH3)2]+.' Details on a folded protein which contained six Co(III) complexes; Definition of partially folded proteins; Methodology used to conduct the study; Results of the study.