Synthesis of Thermoresponsive, Catechol-Rich Poly(ethylene glycol) Brush Polymers for Attenuating Cellular Oxidative Stress.

Herein, we report a platform to integrate customizable quantities of catechol units into polymers by reacting caffeic acid carbonic anhydride with polymers having pendant amine groups. Brush poly(ethylene glycol)-caffeamide (PEG-CAF) copolymers based on oligo(ethylene glycol)methyl ether methacrylate (OEGMA500) were obtained with a catechol content of approximately 30, 40, and 50 mol % (vs OEGMA content). Owing to the hydrophobicity of the introduced CAF groups, the catechol copolymers exhibited cloud points in the range of 23-46 °C and were used to fabricate thermoresponsive FeIII metal-phenolic network capsules. Polymers with the highest CAF content (50 mol %) proved most effective for attenuating reactive oxygen species levels in vitro, in co-cultured fibroblasts, and breast cancer cells, even in the presence of an exogenous oxidant source. The reported approach to synthesize customizable catechol materials could be generalized to other amine-functional polymers, with potential biomedical applications such as adhesives or stimuli-responsive drug delivery systems.

Physiological and pharmacological impact of oxytocin on epididymal propulsion during the ejaculatory process in rodents and men.

During the emission phase of ejaculation, the sperm is driven from the cauda epididymidis, where it is stored, through the vas deferens by strong contractions. These contractions are thought of as being mainly induced by the sympathetic nervous system and the neurotransmitter noradrenaline. In the present study, we investigated the effect of oxytocin (suggested to exert effects during ejaculation as well) on defined segments of the rat and human epididymis using live imaging. Our results indicate that it is the very last part of the epididymis, segment 19 (S19) in rat and likewise segment 9 in human, which responds in a uniquely strong and rapid manner to oxytocin (similar to noradrenaline). Because of the complex nature of this contractile response, we developed an imaging analysis method, which allowed us to quantify multidirectional contractions and to display them using heat maps. The reaction of S19 to oxytocin was concentration-dependent and could be inhibited by pretreatment with oxytocin antagonists (atosiban and cligosiban), but not with an arginine vasopressin 1A antagonist (SR49059). In both rat and human tissue, pretreatment with the alpha-1 adrenoreceptor antagonist tamsulosin inhibited the response to noradrenaline, whereas the effect of oxytocin was unimpaired. Our data (from men and rodents) strongly suggest that the hormone oxytocin is involved in the ejaculatory process. Thus, oxytocin-based medications might be a promising non-adrenergic treatment option for ejaculatory disorders. Additionally, we propose that S19 could be an advantageous model (detecting very low concentrations of oxytocin) to test the bioactivity of new oxytocin agonists and oxytocin antagonists.

Superlattice ordering transitions driven by short-range structure in barium calcium carbonates.

Balcite (BaxCa1-xCO3) is a synthetic analog of rhombohedral carbonate minerals like calcite and dolomite that is disordered on both the cation and anion sublattices. Here, we show that multiple exotic superlattice structures, including a dolomite analog that we call balcomite, can form from balcite at elevated temperatures. The second-order balcite-to-balcomite conversion at temperatures between 150-600 °C is driven by the preference of barium and calcium for different oxygen coordination numbers and facilitated by local carbonate reorientation. At elevated pressure, further superlattice order arises from cation segregation in all three dimensions, producing a supercell with the same R3̄m symmetry as balcite but 6× larger. This highly ordered structure relaxes back to the balcomite structure upon returning to ambient conditions. None of the three naturally occurring polymorphs of Ba0.5Ca0.5CO3 (alstonite, paralstonite, barytocalcite) formed from balcite despite being putatively energetically favored. Instead, alstonite transformed to a balcomite-like structure via a first-order process after transiently converting to a paralstonite-like structure via a second-order process. Together, these results show that high temperature transformation pathways between structures in the barium calcium carbonate system can be driven by coarsening and are facilitated by similarity in short-range order, conceptually analogous to previously described low-temperature transformations. Many of the exotic high temperature carbonate structures are unstable, but may participate in transformation pathways between naturally observed metastable mineral phases, suggesting important roles for ephemeral phases in shaping past and current mineral distributions.

Ion exchange selectivity in clay is controlled by nanoscale chemical-mechanical coupling.

Ion exchange in nanoporous clay-rich media plays an integral role in water, nutrient, and contaminant storage and transport. In montmorillonite (MMT), a common clay mineral in soils, sediments, and muds, the swelling and collapse of clay particles through the addition or removal of discrete molecular layers of water alters cation exchange selectivities in a poorly understood way. Here, we show that ion exchange is coupled to the dynamic delamination and restacking of clay layers, which creates a feedback between the hydration state of the exchanging cation and the composition of the clay interlayer. Particles with different hydration states are distinct phases with unique binding selectivities. Surprisingly, equilibrium achieved through thermal fluctuations in cation concentration and hydration state leads to the exchange of both ions and individual MMT layers between particles, a process we image directly with high-resolution transmission electron microscopy at cryogenic conditions (cryo-TEM). We introduce an exchange model that accounts for the binding selectivities of different phases, which is likely applicable to many charged colloidal or macromolecular systems in which the structural conformation is correlated with the activities of water and counterions within spatially confined compartments.

Trisulfide-Bearing PEG Brush Polymers Donate Hydrogen Sulfide and Ameliorate Cellular Oxidative Stress.

A potential approach to combat cellular dysfunction is to manipulate cell communication and signaling pathways to restore physiological functions while protecting unaffected cells. For instance, delivering the signaling molecule H2S to certain cells has been shown to restore cell viability and re-normalize cell behavior. We have previously demonstrated the ability to incorporate a trisulfide-based H2S-donating moiety into linear polymers with good in vitro releasing profiles and demonstrated their potential for ameliorating oxidative stress. Herein, we report two novel series of brush polymers decorated with higher numbers of H2S-releasing segments. These materials contain two trisulfide-based monomers co-polymerized with oligo(ethylene glycol methyl ether methacrylate) via reversible addition-fragmentation chain-transfer polymerization. The macromolecules were characterized to have a range of trisulfide densities with similar, well-defined molecular weight distribution, good H2S-releasing profiles, and high cellular tolerance. Using an amperometric technique, the H2S liberated and total sulfide release were found to depend on concentrations and chemical nature of triggering molecules (glutathione and cysteine) and, importantly, the position of reactive groups within the brush structure. Notably, when introduced to cells at well-tolerated doses, two macromolecular donors which have the same proportion as of the H2S-donating monomer (30%) but differ in releasing moiety location show similar cellular H2S-releasing kinetics. These donors can restore reactive oxygen species levels to baseline values, when polymer pretreated cells are exposed to exogenous oxidants (H2O2). Our work opens up a new aspect in preparing H2S macromolecule donors and their application to arresting cellular oxidative cascades.

pH-Responsive Polymers for Improving the Signal-to-Noise Ratio of Hypoxia PET Imaging with [18 F]Fluoromisonidazole.

To improve the signal-to-noise ratio of hypoxia positron emission tomography (PET) imaging, stimuli-responsive polymers are designed for the delivery of the hypoxia PET tracer fluorine-18 labeled fluoromisonidazole ([18 F]FMISO). Linear poly(N-(2-(hydroxypropyl)methacrylamide)) polymers are functionalized with hydrazide linkers that form pH-sensitive acyl hydrazone bonds after their conjugation to an [18 F]FMISO ketone analogue. The release of the [18 F]FMISO ketone analogue from the polymers is considerably faster at a lower pH and its uptake is significantly higher in cancer cells growing under acidic conditions. Additionally, the retention of the PET tracer is significantly higher in hypoxic cells compared to normoxic cells. The delivery of a PET tracer using stimuli-responsive polymers may be an attractive strategy to improve signal-to-noise ratios in PET imaging.

The impact of size and charge on the pulmonary pharmacokinetics and immunological response of the lungs to PLGA nanoparticles after intratracheal administration to rats.

Polylactide-co-glycolide (PLGA) nanoparticles are one of the most commonly explored biodegradable polymeric drug carriers for inhaled delivery. Despite their advantages as inhalable nanomedicine scaffolds, we still lack a complete understanding of the kinetics and major pathways by which these materials are cleared from the lungs. This information is important to evaluate their safety over prolonged use and enable successful clinical translation. This study aimed to determine how the size and charge of 3H-labeled PLGA nanoparticles affect the kinetics and mechanisms by which they are cleared from the lungs and their safety in the lungs. The results showed that lung clearance kinetics and retention patterns were more significantly defined by particle size, whereas lung clearance pathways were largely influenced by particle charge. Each of the nanoparticles caused transient inflammatory changes in the lungs after a single dose that reflected lung retention times.

Controlling Nanomaterial Size and Shape for Biomedical Applications via Polymerization-Induced Self-Assembly.

Rapid developments in the polymerization-induced self-assembly (PISA) technique have paved the way for the environmentally friendly production of nanoparticles with tunable size and shape for a diverse range of applications. In this feature article, the biomedical applications of PISA nanoparticles and the substantial progress made in controlling their size and shape are highlighted. In addition to early investigations into drug delivery, applications such as medical imaging, tissue culture, and blood cryopreservation are also described. Various parameters for controlling the morphology of PISA nanoparticles are discussed, including the degree of polymerization of the macro-CTA and core-forming polymers, the concentration of macro-CTA and core-forming monomers, the solid content of the final products, the solution pH, the thermoresponsitivity of the macro-CTA, the macro-CTA end group, and the initiator concentration. Finally, several limitations and challenges for the PISA technique that have been recently addressed, along with those that will require further efforts into the future, will be highlighted.

Elucidating the Influences of Size, Surface Chemistry, and Dynamic Flow on Cellular Association of Nanoparticles Made by Polymerization-Induced Self-Assembly.

The size and surface chemistry of nanoparticles dictate their interactions with biological systems. However, it remains unclear how these key physicochemical properties affect the cellular association of nanoparticles under dynamic flow conditions encountered in human vascular networks. Here, the facile synthesis of novel fluorescent nanoparticles with tunable sizes and surface chemistries and their association with primary human umbilical vein endothelial cells (HUVECs) is reported. First, a one-pot polymerization-induced self-assembly (PISA) methodology is developed to covalently incorporate a commercially available fluorescent dye into the nanoparticle core and tune nanoparticle size and surface chemistry. To characterize cellular association under flow, HUVECs are cultured onto the surface of a synthetic microvascular network embedded in a microfluidic device (SynVivo, INC). Interestingly, increasing the size of carboxylic acid-functionalized nanoparticles leads to higher cellular association under static conditions but lower cellular association under flow conditions, whereas increasing the size of tertiary amine-decorated nanoparticles results in a higher level of cellular association, under both static and flow conditions. These findings provide new insights into the interactions between polymeric nanomaterials and endothelial cells. Altogether, this work establishes innovative methods for the facile synthesis and biological characterization of polymeric nanomaterials for various potential applications.

Bioconjugation and Fluorescence Labeling of Iron Oxide Nanoparticles Grafted with Bromomaleimide-Terminal Polymers.

Iron oxide nanoparticles have been widely applied in biomedical applications for their unique physical properties. Despite the relatively mature synthetic approaches for iron oxide nanoparticles, surface modification strategies for obtaining particles with satisfactory biofunctionality are still urgently needed to meet the challenge of nanomedicine. Herein, we report a surface modification and biofunctionalization strategy for iron oxide-based magnetic nanoparticles based on a dibromomaleimide (DBM)-terminated polymer with brushed polyethylene glycol (PEG) chains. PEG acrylate and phosphonate monomers, serving as antibiofouling and surface anchoring compartments for iron oxide nanoparticles, were incorporated utilizing a novel DBM containing reversible addition-fragmentation chain transfer (RAFT) agent. The particles prepared through this new surface architecture possessed high colloidal stability in a physiological buffer and the capacity of covalent conjugation with biomolecules for targeting. Cell tracking of the molecular probes was achieved concomitantly by exploiting DBM conjugation-induced fluorescence of the nanoparticles.

Exploiting Macromolecular Design To Optimize the Antibacterial Activity of Alkylated Cationic Oligomers.

There is growing interest in synthetic polymers which co-opt the structural features of naturally occurring antimicrobial peptides. However, our understanding of how macromolecular architecture affects antibacterial activity remains limited. To address this, we investigated whether varying architectures of a series of block and statistical co-oligomers influenced antibacterial and hemolytic activity. Cu(0)-mediated polymerization was used to synthesize oligomers constituting 2-(Boc-amino)ethyl acrylate units and either diethylene glycol ethyl ether acrylate (DEGEEA) or poly(ethylene glycol) methyl ether acrylate units with varying macromolecular architecture; subsequent deprotection produced primary amine functional oligomers. Further guanylation provided an additional series of antimicrobial candidates. Both chemical composition and macromolecular architecture were shown to affect antimicrobial activity. A broad spectrum antibacterial oligomer (containing guanidine moieties and DEGEEA units) was identified that possessed promising activity (MIC = 2 µg mL-1) toward both Gram-negative and Gram-positive bacteria. Bacterial membrane permeabilization was identified as an important contributor to the mechanism of action.

Hydrolyzable Poly[Poly(Ethylene Glycol) Methyl Ether Acrylate]-Colistin Prodrugs through Copper-Mediated Photoinduced Living Radical Polymerization.

Through the recently developed copper-mediated photoinduced living radical polymerization (CP-LRP), a novel and well-defined polymeric prodrug of the antimicrobial lipopeptide colistin has been developed. A colistin initiator (Boc5-col-Br2) was synthesized through the modification of colistin on both of its threonine residues using a cleavable initiator linker, 2-(2-bromo-2-methylpropanoyloxy) acetic acid (BMPAA), and used for the polymerization of acrylates via CP-LRP. Polymerization proceeds from both sites of the colistin initiator, and through the polymerization of poly(ethylene glycol) methyl ether acrylate (PEGA480), three water-soluble polymer-colistin conjugates (col-PPEGA, having degrees of polymerization of 5, 10, and 20) were achieved with high yield (conversion of ≥93%) and narrow dispersities (D < 1.3) in 2-4 h. Little or no effect on the structure and activity of the colistin was observed during the synthesis, and most of the active colistin can be recovered from the conjugates in vitro within 2 days. Furthermore, in vitro biological analyses including disk diffusion, broth microdilution, and time-kill studies suggested that all of the conjugates have the ability to inhibit the growth of multidrug-resistant (MDR) Gram-negative bacteria, of which col-PPEGA DP5 and DP10 showed similar or better antibacterial performance compared to the clinically relevant colistin prodrug CMS, indicating their potential as an alternative antimicrobial therapy. Moreover, considering the control over the polymerization, the CP-LRP technique has the potential to provide an alternative platform for the development of polymer bioconjugates.

Influence of Size and Shape on the Biodistribution of Nanoparticles Prepared by Polymerization-Induced Self-Assembly.

Polymerization-induced self-assembly (PISA) is a facile one-pot synthetic technique for preparing polymeric nanoparticles with different sizes and shapes for application in a variety of fields including nanomedicine. However, the in vivo biodistribution of nanoparticles obtained by PISA still remains unclear. To address this knowledge gap, we report the synthesis, cytotoxicity, and biodistribution in an in vivo tumor-bearing mouse model of polystyrene micelles with various sizes and polystyrene filomicelles with different lengths prepared by PISA. First, a library of nanoparticles was prepared comprised of poly(glycidyl methacrylate)-b-poly(oligo(ethylene glycol) methyl ether methacrylate)-b-polystyrene polymers, and their size and morphology were tuned by varying the polystyrene block length without affecting the surface chemistry. The 3H) ethanolamine, and a biodistribution study was carried out in nude mice bearing HT1080 tumor xenografts 48 h after intravenous delivery. In this model, we found that small spherical polystyrene core nanoparticles with a PEG corona (diameter 21 nm) have the highest tumor accumulation when compared to the larger spherical nanoparticles (diameter 33 nm) or rodlike (diameter 37 nm, contour length 350-500 nm) or wormlike counterparts (diameter 45 nm, contour length 1-2 µm). This finding has provided critical information on the biodistribution of polystyrene core nanoparticles with a PEG corona of different sizes and shapes prepared by the PISA technique and will inform their use in medical applications.

Star Polymers Reduce Islet Amyloid Polypeptide Toxicity via Accelerated Amyloid Aggregation.

Protein aggregation into amyloid fibrils is a ubiquitous phenomenon across the spectrum of neurodegenerative disorders and type 2 diabetes. A common strategy against amyloidogenesis is to minimize the populations of toxic oligomers and protofibrils by inhibiting protein aggregation with small molecules or nanoparticles. However, melanin synthesis in nature is realized by accelerated protein fibrillation to circumvent accumulation of toxic intermediates. Accordingly, we designed and demonstrated the use of star-shaped poly(2-hydroxyethyl acrylate) (PHEA) nanostructures for promoting aggregation while ameliorating the toxicity of human islet amyloid polypeptide (IAPP), the peptide involved in glycemic control and the pathology of type 2 diabetes. The binding of PHEA elevated the ß-sheet content in IAPP aggregates while rendering a new morphology of "stelliform" amyloids originating from the polymers. Atomistic molecular dynamics simulations revealed that the PHEA arms served as rodlike scaffolds for IAPP binding and subsequently accelerated IAPP aggregation by increased local peptide concentration. The tertiary structure of the star nanoparticles was found to be essential for driving the specific interactions required to impel the accelerated IAPP aggregation. This study sheds new light on the structure-toxicity relationship of IAPP and points to the potential of exploiting star polymers as a new class of therapeutic agents against amyloidogenesis.

Comb Poly(Oligo(2-Ethyl-2-Oxazoline)Methacrylate)-Peptide Conjugates Prepared by Aqueous Cu(0)-Mediated Polymerization and Reductive Amination.

The controlled synthesis of poly(oligo(2-ethyl-2-oxazoline)methacrylate) (P(OEtOxMA)) polymers by Cu(0)-mediated polymerization in water/methanol mixtures is reported. Utilizing an acetal protected aldehyde initiator for the polymerization, well-defined polymers are synthesized (>99% conversion, Ð < 1.25) with subsequent postpolymerization deprotection resulting in α-aldehyde end group containing comb polymers. These P(OEtOxMA) are subsequently site-specifically conjugated, via reductive amination, to a dipeptide (NH2 -Gly-Tyr-COOH) as a model peptide, prior to conjugation to the functional peptide oxytocin. The resulting oxytocin conjugates are evaluated in comparison to poly(oligo(ethylene glycol) methyl ether methacrylate) combs synthesized in the same manner for potential effects on thermal stability in comparison to the native peptide.

Multi-Step Crystallization of Barium Carbonate: Rapid Interconversion of Amorphous and Crystalline Precursors.

The direct observation of amorphous barium carbonate (ABC), which transforms into a previously unknown barium carbonate hydrate (herewith named gortatowskite) within a few hundred milliseconds of formation, is described. In situ X-ray scattering, cryo-, and low-dose electron microscopy were used to capture the transformation of nanoparticulate ABC into gortatowskite crystals, highly anisotropic sheets that are up to 1 µm in width, yet only about 10 nm in thickness. Recrystallization of gortatowskite to witherite starts within 30 seconds. We describe a bulk synthesis and report a first assessment of the composition, vibrational spectra, and structure of gortatowskite. Our findings indicate that transient amorphous and crystalline precursors can play a role in aqueous precipitation pathways that may often be overlooked owing to their extremely short lifetimes and small dimensions. However, such transient precursors may be integral to the formation of more stable phases.

Macromolecular Hydrogen Sulfide Donors Trigger Spatiotemporally Confined Changes in Cell Signaling.

Hydrogen sulfide (H2S) is involved in a myriad of cell signaling processes that trigger physiological events ranging from vasodilation to cell proliferation. Moreover, disturbances to H2S signaling have been associated with numerous pathologies. As such, the ability to release H2S in a cellular environment and stimulate signaling events is of considerable interest. Herein we report the synthesis of macromolecular H2S donors capable of stimulating cell signaling pathways in both the cytosol and at the cell membrane. Specifically, copolymers having pendent oligo(ethylene glycol) and benzonitrile groups were synthesized, and the benzonitrile groups were subsequently transformed into primary aryl thioamide groups via thionation using sodium hydrosulfide. These thioamide moieties could be incorporated into a hydrophilic copolymer or a block copolymer (i.e., into either the hydrophilic or hydrophobic domain). An electrochemical sensor was used to demonstrate release of H2S under simulated physiological conditions. Subsequent treatment of HEK293 cells with a macromolecular H2S donor elicited a slow and sustained increase in cytosolic ERK signaling, as monitored using a FRET-based biosensor. The macromolecular donor was also shown to induce a small, fast and sustained increase in plasma membrane-localized PKC activity immediately following addition to cells. Studies using an H2S-selective fluorescent probe in live cells confirmed release of H2S from the macromolecular donor over physiologically relevant time scales consistent with the signaling observations. Taken together, these results demonstrate that by using macromolecular H2S donors it is possible to trigger spatiotemporally confined cell signaling events. Moreover, the localized nature of the observed signaling suggests that macromolecular donor design may provide an approach for selectively stimulating certain cellular biochemical pathways.

Stability Enhancing N-Terminal PEGylation of Oxytocin Exploiting Different Polymer Architectures and Conjugation Approaches.

Oxytocin, a cyclic nine amino acid neurohypophyseal hormone therapeutic, is effectively used in the control of postpartum hemorrhaging (PPH) and is on the WHO List of Essential Medicines. However, oxytocin has limited shelf life stability in aqueous solutions, particularly at temperatures in excess of 25 °C and injectable aqueous oxytocin formulations require refrigeration (<8 °C). This is particularly problematic in the hot climates often found in many developing countries where daytime temperatures can exceed 40 °C and where reliable cold-chain storage is not always achievable. The purpose of this study was to develop N-terminal amine targeted PEGylation strategies utilizing both linear PEG and polyPEG "comb" polymers as an effective method for stabilizing solution formulations of this peptide for prolonged storage in the absence of efficient cold-chain storage. The conjugation chemistries investigated herein include irreversible amine targeted conjugation methods utilizing NHS ester and aldehyde reductive amination chemistry. Additionally, one reversible conjugation method using a Schiff base approach was explored to allow for the release of the native peptide, thus, ensuring that biological activity remains unaffected. The reversibility of this approach was investigated for the different polymer architectures, alongside a nonpolymer oxytocin analogue to monitor how pH can tune native peptide release. Elevated temperature degradation studies of the polymer conjugates were evaluated to assess the stability of the PEGylated analogues in comparison to the native peptide in aqueous formulations to mimic storage conditions in developing nations and regions where storage under appropriate conditions is challenging.

Disposition and safety of inhaled biodegradable nanomedicines: Opportunities and challenges.

The inhaled delivery of nanomedicines can provide a novel, non-invasive therapeutic strategy for the more localised treatment of lung-resident diseases and potentially also enable the systemic delivery of therapeutics that are otherwise administered via injection alone. However, the clinical translation of inhalable nanomedicine is being hampered by our lack of understanding about their disposition and clearance from the lungs. This review provides a comprehensive overview of the biodegradable nanomaterials that are currently being explored as inhalable drug delivery systems and our current understanding of their disposition within, and clearance from the lungs. The safety of biodegradable nanomaterials in the lungs is discussed and latest updates are provided on the impact of inflammation on the pulmonary pharmacokinetics of inhaled nanomaterials. Overall, the review provides an in-depth and critical assessment of the lung clearance mechanisms for inhaled biodegradable nanomedicines and highlights the opportunities and challenges for their translation into the clinic.