A new class of polymer spherical nucleic acid (SNA) conjugates comprised of poly(lactic-co-glycolic acid) (PLGA) nanoparticle (NP) cores is reported. The nucleic acid shell that defines the PLGA-SNA exhibits a half-life of more than 2 h in fetal bovine serum. Importantly, the PLGA-SNAs can be utilized to encapsulate a hydrophobic model drug, coumarin 6, which can then be released in a polymer composition-dependent tunable manner, while the dissociation rate of the nucleic acid shell remains relatively constant, regardless of core composition. Like prototypical gold NP conjugate SNAs, PLGA-SNAs freely enter Raw-Blue cells and can be used to activate toll-like receptor 9 in a sequence- and dose-dependent manner. Taken together, the data show that this novel nanoconstruct provides a means for controlling the release kinetics of encapsulated cargos in the context of the SNA platform, which may be useful for developing combination therapeutics.

We have explored the relationship between conformational energetics and the protonation state of the Schiff base in retinal, the covalently bound ligand responsible for activating the G protein–coupled receptor rhodopsin, using quantum chemical calculations. Guided by experimental structural determinations and large-scale molecular simulations on this system, we examined rotation about each bond in the retinal polyene chain, for both the protonated and deprotonated states that represent the dark and photoactivated states, respectively. Particular attention was paid to the torsional degrees of freedom that determine the shape of the molecule, and hence its interactions with the protein binding pocket. While most torsional degrees of freedom in retinal are characterized by large energetic barriers that minimize structural fluctuations under physiological temperatures, the C6–C7 dihedral defining the relative orientation of the β-ionone ring to the polyene chain has both modest barrier heights, and a torsional energy surface that changes dramatically with protonation of the Schiff base. This surprising coupling between conformational degrees of freedom and protonation state is further quantified by calculations of the pKa as a function of the C6–C7 dihedral angle. Notably, pKa shifts of greater than two units arise from torsional fluctuations observed in molecular dynamics simulations of the full ligand-protein-membrane system. It follows that fluctuations in the protonation state of the Schiff base occur prior to forming the activated MII state. These new results shed light on important mechanistic aspects of retinal conformational changes that are involved in the activation of rhodopsin in the visual process.

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Abnormal scarring is a consequence of dysregulation in the wound healing process, with limited options for effective and noninvasive therapies. Given the ability of spherical nucleic acids (SNAs) to penetrate skin and regulate gene expression within, we investigated whether gold-core SNAs (AuSNAs) and liposome-core SNAs (LSNAs) bearing antisense oligonucleotides targeting transforming growth factor beta 1 (TGF-β1) can function as a topical therapy for scarring. Importantly, both SNA constructs appreciably downregulated TGF-β1 protein expression in primary hypertrophic and keloid scar fibroblasts in vitro. In vivo, topically applied AuSNAs and LSNAs downregulated TGF-β1 protein expression levels and improved scar histology as determined by the scar elevation index. These data underscore the potential of SNAs as a localized, self-manageable treatment for skin-related diseases and disorders that are driven by increased gene expression.

The study of materials is governed by the relationship between structure and function. A quintessential example of this phenomenon is deoxyribonucleic acid (DNA), in which molecules assemble in a particular pattern at the nanometer scale, organize into functional configurations at the micrometer scale, and consequently constitute the basis of life at the macroscale [1]. The visualization of nucleic acids and proteins is essential to our understanding of the human body; however, DNA in particular has additionally proven useful in applications beyond biology, such as the functionalization of particles for disease therapy or the patterning of functional substrates [2,3]. In order to characterize and fully control how DNA and other macromolecules may assist in these applications, we must visualize the distribution of DNA strands and observe the mechanisms by which they adhere to surfaces.The primary challenge in electron microscopy of soft materials is achieving reasonable contrast at electron doses low enough to avoid sample damage. Traditional staining methods to overcome signal-tonoise issues require heavy metal contrast agents, which are effective for visualizing the cell membrane due to strong lipid adherence but do not provide adequate staining of nuclear material. A DNA staining method recently developed by Ou et al. and leveraging a technique produced by Maranto (1982) [4,5], chromatin electron microscopy tomography (ChromEMT), allows for positive staining of DNA and other macromolecules in the nucleus of human cells. This method first labels cells with a fluorescent molecule that selectively binds to DNA, and then bathes the cells in diaminobenzidine (DAB). Upon photobleaching, excited fluorescent molecules generate a reactive oxygen species that catalyzes DAB polymerization localized to DNA. Then, cells are stained with osmium tetroxide (OsO 4 ), which binds to DAB with high affinity. The ChromEMT staining technique is tunable to different macromolecules by using appropriate selective fluorescent tags, and is not limited to material in cells [4].We extended ChromEMT to directly image DNA on the surface of polymer-core spherical nucleic acids (SNAs) [2], demonstrating the applicability of the method to isolated nucleic acid strands rather than macromolecules in cell nuclei. SNAs were deposited directly onto grids for transmission electron microscopy (TEM), and the ChromEMT staining procedure was performed by transfer of the grids into droplets of solution for the appropriate step. Four conditions were imaged using a Hitachi HD-2300 STEM in high angle annular dark field (HAADF) mode in order to determine the effectiveness of the staining procedure: 1) SNAs without a staining agent; 2) SNAs stained with OsO 4 ; 3) SNAs labeled with the fluorescent tag cyanine 5 (Cy5), washed in DAB and stained with OsO 4 ; and 4) SNAs labeled with Cy5, washed in DAB, photobleached, and stained with OsO 4 (the entirety of the ChromEMT method). All preparations and microscopy were performed at room temperature. Figures 1a-d s...