Metal-Assembled Collagen Peptide Microflorettes as Magnetic Resonance Imaging Agents.

Magnetic resonance imaging (MRI) is a medical imaging technique that provides detailed information on tissues and organs. However, the low sensitivity of the technique requires the use of contrast agents, usually ones that are based on the chelates of gadolinium ions. In an effort to improve MRI signal intensity, we developed two strategies whereby the ligand DOTA and Gd(III) ions are contained within Zn(II)-promoted collagen peptide (NCoH) supramolecular assemblies. The DOTA moiety was included in the assembly either via a collagen peptide sidechain (NHdota) or through metal-ligand interactions with a His-tagged DOTA conjugate (DOTA-His6). SEM verified that the morphology of the NCoH assembly was maintained in the presence of the DOTA-containing peptides (microflorettes), and EDX and ICP-MS confirmed that Gd(III) ions were incorporated within the microflorettes. The Gd(III)-loaded DOTA florettes demonstrated higher intensities for the T1-weighted MRI signal and higher longitudinal relaxivity (r1) values, as compared to the clinically used contrast agent Magnevist. Additionally, no appreciable cellular toxicity was observed with the collagen microflorettes loaded with Gd(III). Overall, two peptide-based materials were generated that have potential as MRI contrast agents.

Worm-Like Superparamagnetic Nanoparticle Clusters for Enhanced Adhesion and Magnetic Resonance Relaxivity.

Nanosized bioprobes that can highlight diseased tissue can be powerful diagnostic tools. However, a major unmet need is a tool with adequate adhesive properties and contrast-to-dose ratio. To this end, this study demonstrates that targeted superparamagnetic nanoprobes engineered to present a worm-like shape and hydrophilic packaging enhance both adhesion efficiency to target substrates and magnetic resonance (MR) sensitivity. These nanoprobes were prepared by the controlled self-assembly of superparamagnetic iron oxide nanoparticles (SPIONs) into worm-like superstructures using glycogen-like amphiphilic hyperbranched polyglycerols functionalized with peptides capable of binding to defective vasculature. The resulting worm-like SPION clusters presented binding affinity to the target substrate 10-fold higher than that of spherical ones and T2 molar MR relaxivity 3.5-fold higher than that of conventional, single SPIONs. The design principles discovered for these nanoprobes should be applicable to a range of other diseases where improved diagnostics are needed.

Hydrophilic packaging of iron oxide nanoclusters for highly sensitive imaging.

Superparamagnetic iron oxide nanoparticles (SPIONs) are used as imaging probes to provide contrast in magnetic resonance images. Successful use of SPIONs in targeted applications greatly depends on their ability to generate contrast, even at low levels of accumulation, in the tissue of interest. In the present study, we report that SPION nanoclusters packaged to a controlled size by a hyperbranched polyglycerol (HPG) can target tissue defects and have a high relaxivity of 719 mM(-1) s(-1), which was close to their theoretical maximal limit. The resulting nanoclusters were able to identify regions of defective vasculature in an ischemic murine hindlimb using MRI with iron doses that were 5-10 fold lower than those typically used in preclinical studies. Such high relaxivity was attributed to the molecular architecture of HPG, which mimics that of the water retentive polysaccharide, glycogen. The results of this study will be broadly useful in sensitive imaging applications.

Controlling the morphology of metal-promoted higher ordered assemblies of collagen peptides with varied core lengths.

Self-assembling peptides have become an important subclass of next-generation biomaterials. In particular, materials that mimic the properties of collagen have received considerable attention due to the unique properties of natural collagen. Previous peptide-based designs have been successful in generating structures with morphological properties that were primarily determined by the type of self-assembling mechanism. Herein we demonstrate the metal ion-promoted, supramolecular assembly of collagen-based peptide triple helices into distinct morphologies that are controlled by defining the number of Pro-Hyp-Gly repeating units. We synthesized and characterized collagen-based peptides that incorporated either 5, 7, 9, or 11 Pro-Hyp-Gly repeating units. We found that the number of repeating units, and the resulting stability of the collagen triple helix, is intimately linked with the types of assemblies formed. For instance, collagen peptides that did not form a stable triple helix, such as NCoH5, did not participate in supramolecular assembly with added metal ions. Collagen peptides that formed stable triple helices, such as NCoH11, resulted in microsaddle structures with metal-promoted assembly, whereas a highly cross-linked, three-dimensional mesh formed with NCoH7, albeit at a higher metal ion concentration. These data provide evidence that triple helix formation is required for efficient metal-triggered assembly to the observed microstructures.

Selective decoration and release of His-tagged proteins from metal-assembled collagen peptide microflorettes.

Materials that mimic the extracellular matrix may serve as ideal delivery vehicles for biopolymers with biomedical applications. Herein we investigate dual His-tagged protein modification and release of metal-triggered, collagen peptide microflorettes by taking advantage of unsatisfied metal/ligands on or within the microflorette structures. Using GFP and RFP as model proteins for visualization, microflorettes were treated with His-tagged proteins either during or after particle assembly. Fluorescence microscopy confirmed the essential role of the His-tag in protein functionalization of the florettes, and confocal microscopy demonstrated distinct labeling zones either within the core or on the surface of the particles depending on their mode of synthesis. The location of the His-tagged proteins within the microflorettes was found to strongly influence the rate of release of these proteins from the particles, with the surface-localized proteins demonstrating faster release in comparison to the core-localized proteins. We have demonstrated, therefore, dual His-tagged protein functionalization with spatial control within metal-triggered, collagen peptide microflorette structures, and temporally controlled release of these proteins into biological media.