Surface engineered polymersomes for enhanced modulation of dendritic cells during cardiovascular immunotherapy.

The principle cause of cardiovascular disease (CVD) is atherosclerosis, a chronic inflammatory condition characterized by immunologically complex fatty lesions within the intima of arterial vessel walls. Dendritic cells (DCs) are key regulators of atherosclerotic inflammation, with mature DCs generating pro-inflammatory signals within vascular lesions and tolerogenic DCs eliciting atheroprotective cytokine profiles and regulatory T cell (Treg) activation. Here, we engineered the surface chemistry and morphology of synthetic nanocarriers composed of poly(ethylene glycol)-b-poly(propylene sulfide) copolymers to selectively target and modulate DCs by transporting the anti-inflammatory agent 1, 25-Dihydroxyvitamin D3 (aVD) and ApoB-100 derived antigenic peptide P210. Polymersomes decorated with an optimized surface display and density for a lipid construct of the P-D2 peptide, which binds CD11c on the DC surface, significantly enhanced the cytosolic delivery and resulting immunomodulatory capacity of aVD in vitro. Intravenous administration of the optimized polymersomes achieved selective targeting of DCs in atheroma and spleen compared to all other cell populations, including both immune and CD45- cells, and locally increased the presence of tolerogenic DCs and cytokines. aVD-loaded polymersomes significantly inhibited atherosclerotic lesion development in high fat diet-fed ApoE-/- mice following 8 weeks of administration. Incorporation of the P210 peptide generated the largest reductions in vascular lesion area (~33%, p<0.001), macrophage content (~55%, p<0.001), and vascular stiffness (4.8-fold). These results correlated with an ~6.5-fold increase in levels of Foxp3+ regulatory T cells within atherosclerotic lesions. Our results validate the key role of DC immunomodulation during aVD-dependent inhibition of atherosclerosis and demonstrate the therapeutic enhancement and dosage lowering capability of cell-targeted nanotherapy in the treatment of CVD.

Measuring ligand-receptor binding kinetics and dynamics using k-space image correlation spectroscopy.

Accurate measurements of kinetic rate constants for interacting biomolecules are crucial for understanding the mechanisms underlying intracellular signalling pathways. The magnitude of binding rates plays a very important molecular regulatory role which can lead to very different cellular physiological responses under different conditions. Here, we extend the k-space image correlation spectroscopy (kICS) technique to study the kinetic binding rates of systems wherein: (a) fluorescently labelled, free ligands in solution interact with unlabelled, diffusing receptors in the plasma membrane and (b) systems where labelled, diffusing receptors are allowed to bind/unbind and interconvert between two different diffusing states on the plasma membrane. We develop the necessary mathematical framework for the kICS analysis and demonstrate how to extract the relevant kinetic binding parameters of the underlying molecular system from fluorescence video-microscopy image time-series. Finally, by examining real data for two model experimental systems, we demonstrate how kICS can be a powerful tool to measure molecular transport coefficients and binding kinetics.

Reversible self-assembly of superstructured networks.

Soft structures in nature, such as protein assemblies, can organize reversibly into functional and often hierarchical architectures through noncovalent interactions. Molecularly encoding this dynamic capability in synthetic materials has remained an elusive goal. We report on hydrogels of peptide-DNA conjugates and peptides that organize into superstructures of intertwined filaments that disassemble upon the addition of molecules or changes in charge density. Experiments and simulations demonstrate that this response requires large-scale spatial redistribution of molecules directed by strong noncovalent interactions among them. Simulations also suggest that the chemically reversible structures can only occur within a limited range of supramolecular cohesive energies. Storage moduli of the hydrogels change reversibly as superstructures form and disappear, as does the phenotype of neural cells in contact with these materials.