Collagen-binding peptides for the enhanced imaging, lubrication and regeneration of osteoarthritic articular cartilage.

Treatments for osteoarthritis would benefit from the enhanced visualization of injured articular cartilage and from the targeted delivery of disease-modifying drugs to it. Here, by using ex vivo human osteoarthritic cartilage and live rats and minipigs with induced osteoarthritis, we report the application of collagen-binding peptides, identified via phage display, that are home to osteoarthritic cartilage and that can be detected via magnetic resonance imaging when conjugated with a superparamagnetic iron oxide. Compared with the use of peptides with a scrambled sequence, hyaluronic acid conjugated with the collagen-binding peptides displayed enhanced retention in osteoarthritic cartilage and better lubricated human osteoarthritic tissue ex vivo. Mesenchymal stromal cells encapsulated in the modified hyaluronic acid and injected intra-articularly in rats showed enhanced homing to osteoarthritic tissue and improved its regeneration. Molecular docking revealed WXPXW as the consensus motif that binds to the α1 chain of collagen type XII. Peptides that specifically bind to osteoarthritic tissue may aid the diagnosis and treatment of osteoarthritic joints.

A Polypeptide-Based, Membrane-Penetrating, Target-Specific Contrast Agent for Magnetic Resonance Molecular Imaging.

Molecular imaging based on magnetic resonance imaging (MRI) requires a contrast agent with high relaxivity and specificity. Much effort has been devoted to this goal over the past decades. In this work, we continue this endeavor by synthesizing an MRI contrast agent that can penetrate the cellular membrane and bind with specific proteins. It was characterized with one- and two-dimensional NMR spectroscopy, NMR micro-imaging, and mass spectroscopy. The target specificity has been further confirmed by both molecular dynamics simulation and micro-imaging on a living biological system. It is one of the largest of peptide-based bioactivated MRI contrast agents, and its relaxivity enhancement factor is among the highest of MRI contrast agents hitherto published. We envision interesting applications and extension of this smart MRI contrast agent with bio-specificity and high contrast for molecular imaging.

Anti-apoptotic BCL-2 regulation by changes in dynamics of its long unstructured loop.

BCL-2, a key protein in inhibiting apoptosis, has a 65-residue-long highly flexible loop domain (FLD) located on the opposite side of its ligand-binding groove. In vivo phosphorylation of the FLD enhances the affinity of BCL-2 for pro-apoptotic ligands, and consequently anti-apoptotic activity. However, it remains unknown as to how the faraway, unstructured FLD modulates the affinity. Here we investigate the protein-ligand interactions by fluorescence techniques and monitor protein dynamics by DEER and NMR spectroscopy tools. We show that phosphomimetic mutations on the FLD lead to a reduction in structural flexibility, hence promoting ligand access to the groove. The bound pro-apoptotic ligands can be displaced by the BCL-2-selective inhibitor ABT-199 efficiently, and thus released to trigger apoptosis. We show that changes in structural flexibility on an unstructured loop can activate an allosteric protein that is otherwise structurally inactive.

Engineered Paramagnetic Graphene Quantum Dots with Enhanced Relaxivity for Tumor Imaging.

Nano contrast agents (Nano CA) are nanomaterials used to increase contrast in the medical magnetic resonance imaging (MRI). However, the related relaxation mechanism of the Nano CA is not clear yet and little significant breakthrough in relaxivity enhancement has been achieved. Herein, a new hydrophilic Gd-DOTA complex functionalized with different chain length of PEG was synthesized and incorporated into graphene quantum dots (GQD) to obtain paramagnetic graphene quantum dots (PGQD). We performed a variable-temperature and variable-field intensity NMR study in aqueous solution on the water exchange and rotational dynamics of three different chain lengths of PGQD. The optimal GQD with paramagnetic chain length shows a great improvement in performance on 1H NMR relaxometric studies. In vitro results demonstrated that the relaxivity of the designed PGQD could be controlled by regulating the PEG length, and its relaxivity was ∼16 times higher than that of current commercial MRI contrast agents (e.g., Gd-DTPA), on a "per Gd" basis. The relaxivity of the Nano CA can be rationally tuned to obtain unmatched potentials in MR imaging, exemplified by preparation of the paramagnetic GQD with the enhanced T1 relaxivity. The fabricated PGQDs with suitable PEG length got the best relaxivity at 1.5 T. After intravenous injection, its feeding process by solid tumor could even be monitored by clinically used 1.5 T MRI scanners. This research will also provide an excellent platform for the design and synthesis of highly effective MR contrast agents.

Extending the Lifetime of Hyperpolarized Propane Gas through Reversible Dissolution.

Hyperpolarized (HP) propane produced by the parahydrogen-induced polarization (PHIP) technique has been recently introduced as a promising contrast agent for functional lung magnetic resonance (MR) imaging. However, its short lifetime due to a spin-lattice relaxation time T1 of less than 1 s in the gas phase is a significant translational challenge for its potential biomedical applications. The previously demonstrated approach for extending the lifetime of the HP propane state through long-lived spin states allows the HP propane lifetime to be increased by a factor of ∼3. Here, we demonstrate that a remarkable increase in the propane hyperpolarization decay time at high magnetic field (7.1 T) can be achieved by its dissolution in deuterated organic solvents (acetone-d6 or methanol-d4). The approximate values of the HP decay time for propane dissolved in acetone-d6 are 35.1 and 28.6 s for the CH2 group and the CH3 group, respectively (similar values were obtained for propane dissolved in methanol-d4), which are ∼50 times larger than the gaseous propane T1 value. Furthermore, we show that it is possible to retrieve HP propane from solution to the gas phase with the preservation of hyperpolarization.

Guizhi Fuling Wan as a Novel Agent for Intravesical Treatment for Bladder Cancer in a Mouse Model.

Alternative intravesical agents are required to overcome the side effects currently associated with the treatment of bladder cancer. This study used an orthotopic bladder cancer mouse model to evaluate Guizhi Fuling Wan (GFW) as an intravesical agent. The effects of GFW were compared with those of mitomycin-C (Mito-C) and bacille Calmette-Guérin (BCG). We began by evaluating the response of the mouse bladder cancer cell line MB49 to GFW treatment, with regard to cell viability, cell cycle progression and apoptosis. MB49 cells were subsequently implanted into the urothelial walls of the bladder in female C57BL/6 mice. The success of the model was confirmed by the appearance of hematuria and tumor growth in the bladder. Intravesical chemotherapy was administered in accordance with a published protocol. In vitro data revealed that GFW arrested MB49 cell cycle in the G0/G1 phase, resulting in the suppression of cell proliferation and induced apoptosis. One possible mechanism underlying these effects is an increase in intracellular reactive oxygen species (ROS) levels leading to the activation of ataxia telangiectasia-mutated (ATM)/checkpoint kinase 2 (CHK2) and ATM/P53 pathways, thereby mediating cell cycle progression and apoptosis, respectively. This mouse model demonstrates the effectiveness of GFW in the tumor growth, with results comparable to those achieved by using BCG and Mito-C. Furthermore, GFW was shown to cause only mild hematuria. The low toxicity of the compound was confirmed by a complete lack of lesions on bladder tissue, even after 10 consecutive treatments using high concentrations of GFW. These results demonstrate the potential of GFW for the intravesical therapy of bladder cancer.

Sensitive imaging of magnetic nanoparticles for cancer detection by active feedback MR.

PURPOSE: Sensitive imaging of superparamagnetic nanoparticles or aggregates is of great importance in MR molecular imaging and medical diagnosis. For this purpose, a conceptually new approach, termed active feedback magnetic resonance, was developed. METHODS: In the presence of the Zeeman field, a dipolar field is induced by the superparamagnetic nanoparticles or aggregates. Such dipolar field creates spatial and temporal (due to water diffusion) variations to the precession frequency of the nearby water 1 H magnetization. Sensitive imaging of magnetic nanoparticles or aggregates can be achieved by manipulating the intrinsic spin dynamics by selective self-excitation and fixed-point dynamics under active feedback fields. RESULTS: Phantom experiments of superparamagnetic nanoparticles; in vitro experiments of brain tissue with blood clots; and in vivo mouse images of colon cancers, with and without labeling by magnetic nanoparticles, suggest that this new approach provides enhanced, robust, and positive contrast in imaging magnetic nanoparticles or aggregates for cancer detection. CONCLUSION: The spin dynamics originated from selective self-excitation and fixed-point dynamics under active feedback fields have been shown to be sensitive to dipolar fields generated by magnetic nanoparticles. Magn Reson Med 74:33-41, 2015. © 2014 Wiley Periodicals, Inc.

Novel MRI contrast development by lock-in suppression.

PURPOSE: The goal of this study is to develop novel MR contrast by frequency lock-in technique. METHODS: An electronic feedback device that can control the frequency and bandwidth of the feedback RF field is presented. In this study, the effects of lock-in suppressed imaging are discussed both theoretically and experimentally. RESULTS: Two important imaging experiments were performed. The first experiment used magnetizations with the same central frequency but different frequency distributions and was compared with MR images obtained with T2 contrast agents. Lock-in suppressed images showed an improvement in contrast relative to the conventional imaging method. The second experiment used magnetizations with small shifts in frequency and a broad frequency distribution. This is helpful for differentiating between small structural variations in biological tissues. The contrast achieved in in vivo tumor imaging using the lock-in suppressed technique provide higher spatial resolutions and discriminate the regimes of necrosis and activation consistent with pathologic results. CONCLUSION: Lock-in suppressed imaging introduces a conceptually new approach to MRI. Heightened sensitivity to underlying susceptibility variations and their relative contribution to total magnetization may thus be achieved to yield new and enhanced contrast.

NMR relaxation study of water dynamics in superparamagnetic iron-oxide-loaded vesicles.

Superparamagnetic iron oxide (SPIO) nanoparticles have been introduced as contrast agents for clinical applications in magnetic resonance imaging. Recently, SPIO has been also used for tracking cells. However, NMR relaxation of water molecules behaves differently in a SPIO solution and SPIO-loaded cells. In this study, we used water-in-oil-in-water double emulsions to mimic cellular environments. The MR relaxation induced by the SPIO-loaded vesicles and SPIO solution indicates that T(2)* is sensitive to the iron concentration alone, and the behavior was very similar in both SPIO-loaded vesicles and SPIO solution. However, T(2) relaxation of water in SPIO-loaded vesicles was faster than that in a SPIO solution. In addition, the contribution of water inside and outside the vesicles was clarified by replacing H(2)O with D(2)O, and water inside the vesicles was found to cause a nonlinear iron concentration dependency. The studied dilution revealed that vesicle aggregation undergoes a structural transition upon dilution by a certain amount of water. R(2)* relaxation is sensitive to this structural change and shows an obvious nonlinear iron concentration dependency when the SPIO loading is sufficiently high. Random walk simulations demonstrated that in the assumed model, the vesicles aggregate structures causing the differences between R(2)* and R(2) relaxation of water in vesicles in the presence of SPIO particles.