Photothermal-Enhanced Modulation of Cellular Membrane Potential Using Long-Wavelength-Activated Gold Nanoflowers.

In mammalian cells, plasma membrane potential plays vital roles in both physiology and pathology and it is controlled by a network of membrane-resident ion channels. There is considerable interest in the use of nanoparticles (NPs) to control biological functions, including the modulation of membrane potential. The photoexcitation of gold NPs (AuNPs) tethered close to the plasma membrane has been shown to induce membrane depolarization via localized heating of the AuNP surface coupled with the opening of voltage-gated sodium channels. Previous work has employed spherical AuNPs (AuNS) with absorption in the 500-600 nm range for this purpose. However, AuNP materials with absorption at longer wavelengths [e.g., near-infrared (NIR)] would enable greater tissue penetration depth in vivo. We show here the use of new anisotropic-shaped AuNPs [gold nanoflowers (AuNFs)] with broad absorption spanning into the NIR part of the spectrum (∼650-1000 nm). The AuNFs are directly synthesized with bidentate thiolate ligands, which preserves the AuNF's shape and colloidal stability, while facilitating conjugation to biomolecules. We describe the characterization of the AuNF particles and demonstrate that they adhere to the plasma membrane when bioconjugated to PEGylated cholesterol (PEG-Chol) moieties. The AuNF-PEG-Chol mediated the depolarization of rat adrenal medulla pheochromocytoma (PC-12) neuron-like cells more effectively than AuNS-PEG-Chol and unconjugated AuNS and AuNF when photoexcited at ∼561 or ∼640 nm. Importantly, AuNF induction of depolarization had no impact on cellular viability. This work demonstrates anisotropic AuNFs as an enabling nanomaterial for use in cellular depolarization and the spatiotemporal control of cellular activity.

Multivalent Display of Erythropoietin on Quantum Dots Enhances Aquaporin-4 Expression and Water Transport in Human Astrocytes In Vitro.

In mammalian cells, growth factor-induced intracellular signaling and protein synthesis play a critical role in cellular physiology and homeostasis. In the brain's glymphatic system (GS), the water-conducting activity of aquaporin-4 (AQPN-4) membrane channels (expressed in polarized fashion on astrocyte end-feet) mediates the clearance of wastes through the convective transport of fluid and solutes through the perivascular space. The glycoprotein erythropoietin (EPO) has been shown to induce the astrocyte expression of AQPN-4 via signaling through the EPO receptor and the JAK/STAT signaling pathway. Here, we self-assemble EPO in a multivalent fashion onto the surface of semiconductor quantum dots (QDs) (driven by polyhistidine-based self-assembly) to drive the interaction of the bioconjugates with EPOR on human astrocytes (HA). This results in a 2-fold augmentation of JAK/STAT signaling activity and a 1.8-fold enhancement in the expression of AQPN-4 in cultured primary HA compared to free EPO. This translates into a 2-fold increase in the water transport rate in HA cells as measured by the calcein AM water transport assay. Importantly, EPO-QD-induced augmented AQPN-4 expression does not elicit any deleterious effect on the astrocyte viability. We discuss our results in the context of the implications of EPO-nanoparticle (NP) bioconjugates for use as research tools to understand the GS and their potential as therapeutics for the modulation of GS function. More generally, our results illustrate the utility of NP bioconjugates for the controlled modulation of growth factor-induced intracellular signaling.

Determining the Cytosolic Stability of Small DNA Nanostructures In Cellula.

DNA nanostructures have proven potential in biomedicine. However, their intracellular interactions─especially cytosolic stability─remain mostly unknown and attempts to discern this are confounded by the complexities of endocytic uptake and entrapment. Here, we bypass the endocytic uptake and evaluate the DNA structural stability directly in live cells. Commonly used DNA structures─crosshairs and a tetrahedron─were labeled with a multistep Förster resonance energy transfer dye cascade and microinjected into the cytosol of transformed and primary cells. Energy transfer loss, as monitored by fluorescence microscopy, reported the structure's direct time-resolved breakdown in cellula. The results showed rapid degradation of the DNA crosshair within 20 min, while the tetrahedron remained consistently intact for at least 1 h postinjection. Nuclease assays in conjunction with a current understanding of the tetrahedron's torsional rigidity confirmed its higher stability. Such studies can inform design parameters for future DNA nanostructures where programmable degradation rates may be required.

Melanin Produced by the Fast-Growing Marine Bacterium Vibrio natriegens through Heterologous Biosynthesis: Characterization and Application.

Melanin is a pigment produced by organisms throughout all domains of life. Due to its unique physicochemical properties, biocompatibility, and biostability, there has been an increasing interest in the use of melanin for broad applications. In the vast majority of studies, melanin has been either chemically synthesized or isolated from animals, which has restricted its use to small-scale applications. Using bacteria as biocatalysts is a promising and economical alternative for the large-scale production of biomaterials. In this study, we engineered the marine bacterium Vibrio natriegens, one of the fastest-growing organisms, to synthesize melanin by expressing a heterologous tyrosinase gene and demonstrated that melanin production was much faster than in previously reported heterologous systems. The melanin of V. natriegens was characterized as a polymer derived from dihydroxyindole-2-carboxylic acid (DHICA) and, similarly to synthetic melanin, exhibited several characteristic and useful features. Electron microscopy analysis demonstrated that melanin produced from V. natriegens formed nanoparticles that were assembled as "melanin ghost" structures, and the photoprotective properties of these particles were validated by their protection of cells from UV irradiation. Using a novel electrochemical reverse engineering method, we observed that melanization conferred redox activity to V. natriegens Moreover, melanized bacteria were able to quickly adsorb the organic compound trinitrotoluene (TNT). Overall, the genetic tractability, rapid division time, and ease of culture provide a set of attractive properties that compare favorably to current E. coli production strains and warrant the further development of this chassis as a microbial factory for natural product biosynthesis.IMPORTANCE Melanins are macromolecules that are ubiquitous in nature and impart a large variety of biological functions, including structure, coloration, radiation resistance, free radical scavenging, and thermoregulation. Currently, in the majority of investigations, melanins are either chemically synthesized or extracted from animals, which presents significant challenges for large-scale production. Bacteria have been used as biocatalysts to synthesize a variety of biomaterials due to their fast growth and amenability to genetic engineering using synthetic biology tools. In this study, we engineered the extremely fast-growing bacterium V. natriegens to synthesize melanin nanoparticles by expressing a heterologous tyrosinase gene with inducible promoters. Characterization of the melanin produced from V. natriegens-produced tyrosinase revealed that it exhibited physical and chemical properties similar to those of natural and chemically synthesized melanins, including nanoparticle structure, protection against UV damage, and adsorption of toxic compounds. We anticipate that producing and controlling melanin structures at the nanoscale in this bacterial system with synthetic biology tools will enable the design and rapid production of novel biomaterials for multiple applications.

Gold-Nanoparticle-Mediated Depolarization of Membrane Potential Is Dependent on Concentration and Tethering Distance from the Plasma Membrane.

The photoactivation of plasma-membrane-tethered gold nanoparticles (AuNPs) for the photothermally driven depolarization of membrane potential has recently emerged as a new platform for the controlled actuation of electrically active cells. In this report, we characterize the relationship between AuNP concentration and AuNP-membrane separation distance with the efficiency of photoactivated plasma membrane depolarization. We show in differentiated rat pheochromocytoma (PC-12) cells that AuNPs capped with poly(ethylene glycol) (PEG)-cholesterol ligands localize to the plasma membrane and remain resident for up to 1 h. The efficiency of AuNP-mediated depolarization is directly dependent on the concentration of the NPs on the cell surface. We further show that the efficiency of AuNP-mediated photothermal depolarization of membrane potential is directly dependent on the tethering distance between the AuNP and the plasma membrane, which we control by iteratively tuning the length of the PEG linker. Importantly, the AuNP conjugates do not adversely affect cell viability under the photoactivation conditions required for membrane depolarization. Our results demonstrate the fine control that can be elicited over AuNP bioconjugates and establishes principles for the rational design of functional nanomaterials for the control of electrically excitable cells.

Recent Progress in Bioconjugation Strategies for Liposome-Mediated Drug Delivery.

In nanoparticle (NP)-mediated drug delivery, liposomes are the most widely used drug carrier, and the only NP system currently approved by the FDA for clinical use, owing to their advantageous physicochemical properties and excellent biocompatibility. Recent advances in liposome technology have been focused on bioconjugation strategies to improve drug loading, targeting, and overall efficacy. In this review, we highlight recent literature reports (covering the last five years) focused on bioconjugation strategies for the enhancement of liposome-mediated drug delivery. These advances encompass the improvement of drug loading/incorporation and the specific targeting of liposomes to the site of interest/drug action. We conclude with a section highlighting the role of bioconjugation strategies in liposome systems currently being evaluated for clinical use and a forward-looking discussion of the field of liposomal drug delivery.

Hybrid Liquid Crystal Nanocarriers for Enhanced Zinc Phthalocyanine-Mediated Photodynamic Therapy.

Current challenges in photodynamic therapy (PDT) include both the targeted delivery of the photosensitizer (PS) to the desired cellular location and the maintenance of PS efficacy. Zinc phthalocyanine (ZnPc), a macrocyclic porphyrin and a potent PS for PDT, undergoes photoexcitation to generate reactive singlet oxygen that kills cells efficiently, particularly when delivered to the plasma membrane. Like other commonly employed PS, ZnPc is highly hydrophobic and prone to self-aggregation in aqueous biological media. Further, it lacks innate subcellular targeting specificity. Cumulatively, these attributes pose significant challenges for delivery via traditional systemic drug delivery modalities. Here, we report the development and characterization of a liquid crystal nanoparticle (LCNP)-based formulation for the encapsulation and targeted tethering of ZnPc to the plasma membrane bilayer. ZnPc was coloaded with the organic fluorophore, perylene (PY), in the hydrophobic polymeric matrix of the LCNP core. PY facilitated the fluorescence-based tracking of the LCNP carrier while also serving as a Förster resonance energy transfer (FRET) donor to the ZnPc acceptor. This configuration availed efficient singlet oxygen generation via enhanced excitation of ZnPc from multiple surrounding PY energy donors. When excited in a FRET configuration, cuvette-based assays revealed that singlet oxygen generation from the ZnPc was ∼1.8-fold greater and kinetically 12 times faster compared to when the ZnPc was excited directly. The specific tethering of the LCNPs to the plasma membrane of HEK 293 T/17 and HeLa cells was achieved by surface functionalization of the NPs with PEGylated cholesterol. In HeLa cells, LCNPs coloaded with PY and ZnPc, when photoexcited in a FRET configuration, mediated 70% greater cell killing compared to LCNPs containing ZnPc alone (direct excitation of ZnPc). This was attributed to a significant increase of the oxidative stress in the cells during the PDT. Overall, this work details the ability of the LCNP platform to facilitate (1) the specific tethering of the PY-ZnPc FRET pair to the plasma membrane and (2) the FRET-mediated, augmented singlet oxygen generation for enhanced PDT relative to the direct excitation of ZnPc alone.

Lipid Raft-Mediated Membrane Tethering and Delivery of Hydrophobic Cargos from Liquid Crystal-Based Nanocarriers.

A main goal of bionanotechnology and nanoparticle (NP)-mediated drug delivery (NMDD) continues to be the development of novel biomaterials that can controllably modulate the activity of the NP-associated therapeutic cargo. One of the desired subcellular locations for targeted delivery in NMDD is the plasma membrane. However, the controlled delivery of hydrophobic cargos to the membrane bilayer poses significant challenges including cargo precipitation and lack of specificity. Here, we employ a liquid crystal NP (LCNP)-based delivery system for the controlled partitioning of a model dye cargo from within the NP core into the plasma membrane bilayer. During synthesis of the NPs, the water-insoluble model dye cargo, 3,3'-dioctadecyloxacarbocyanine perchlorate (DiO), was efficiently incorporated into the hydrophobic LCNP core as confirmed by multiple spectroscopic analyses. Conjugation of a PEGylated cholesterol derivative to the NP surface (DiO-LCNP-PEG-Chol) facilitated the localization of the dye-loaded NPs to lipid raft microdomains in the plasma membrane in HEK 293T/17 cell. Analysis of DiO cellular internalization kinetics revealed that when delivered as a LCNP-PEG-Chol NP, the half-life of DiO membrane residence time (30 min) was twice that of free DiO (DiO(free)) (15 min) delivered from bulk solution. Time-resolved laser scanning confocal microscopy was employed to visualize the passive efflux of DiO from the LCNP core and its insertion into the plasma membrane bilayer as confirmed by Förster resonance energy transfer (FRET) imaging. Finally, the delivery of DiO as a LCNP-PEG-Chol complex resulted in the attenuation of its cytotoxicity; the NP form of DiO exhibited ∼30-40% less toxicity compared to DiO(free). Our data demonstrate the utility of the LCNP platform as an efficient vehicle for the combined membrane-targeted delivery and physicochemical modulation of molecular cargos using lipid raft-mediated tethering.

Biological evaluation of liposome-encapsulated hemoglobin surface-modified with a novel PEGylated nonphospholipid amphiphile.

Traumatic injury is often associated with hemorrhagic shock. Liposome-encapsulated hemoglobin (LEH) is being developed as an artificial oxygen carrier to address post-hemorrhage oxygen and volume deficit. Here, we report a new composition of LEH based on the use of polyethylene glycol (PEG2K ) conjugated with nonphospholipid hexadecylcarbamoylmethylhexadecanoate (HDAS) to modify the surface of LEH particles. LEH was manufactured by the high-pressure homogenization method using dipalmitoylphosphatidylcholine (∼38 mol%), cholesterol (∼38 mol%), HDAS (∼20 mol%), and highly purified stroma-free human hemoglobin. HDAS-PEG2K was postinserted into the resultant LEH to generate HDAS-PEG2K -LEH. We investigated the potential immune response to HDAS-PEG2K -LEH in a mice model. At the same time, the preparation was tested in a rat model to study the effect of repeated HDAS-PEG2K -LEH injection over 4 weeks. We found that HDAS-PEG2K modification substantially reduced the circulating levels of anaphylatoxins C3a and C5a, as well as plasma levels of thromboxane B2, in mice. Repeated injections of HDAS-PEG2K -LEH in rats did not appear to alter its clearance profile after 4 weeks of treatment. No antibody response against human hemoglobin or PEG was detected in rat plasma. Histological observations of lung, liver, spleen, and kidney were not significantly different between saline-treated rats and HDAS-PEG2K -LEH-treated rats. Immunohistochemical staining for rat heme oxygenase-1 (HO-1) did not show induced expression of HO-1 in these organs. These results suggest that the new surface modification of LEH is immune-neutral and does not adversely affect histology even after repeated administration.

Special Issue "Nanoparticle-Mediated Drug Delivery, Imaging, and Control of Cellular Functions".

Over the past several decades, nanoparticles (NPs) have shown promising capabilities in the field of medicine for their applications as vehicles for targeted drug delivery [...].

Liquid Crystal Nanoparticle Conjugates for Scavenging Reactive Oxygen Species in Live Cells.

The elevated intracellular production of or extracellular exposure to reactive oxygen species (ROS) causes oxidative stress to cells, resulting in deleterious irreversible biomolecular reactions (e.g., lipid peroxidation) and disease progression. The use of low-molecular weight antioxidants, such as 4-amino-2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO), as ROS scavengers fails to achieve the desired efficacy because of their poor or uncontrolled cellular uptake and off-target effects, such as dysfunction of essential redox homeostasis. In this study, we fabricated a liquid crystal nanoparticle (LCNP) conjugate system with the fluorescent dye perylene (PY) loaded in the interior and poly (ethylene glycol) (PEG) decorated on the surface along with multiple molecules of TEMPO (PY-LCNP-PEG/TEMPO). PY-LCNP-PEG/TEMPO exhibit enhanced cellular uptake, and efficient ROS-scavenging activity in live cells. On average, the 120 nm diameter PY-LCNPs were conjugated with >1800 molecules of TEMPO moieties on their surface. PY-LCNP-PEG/TEMPO showed significantly greater reduction in ROS activity and lipid peroxidation compared to free TEMPO when the cells were challenged with ROS generating agents, such as hydrogen peroxide (H2O2). We suggest that this is due to the increased local concentration of TEMPO molecules on the surface of the PY-LCNP-PEG/TEMPO NPs, which efficiently bind to the plasma membrane and enter cells. Overall, these results demonstrate the enhanced capability of TEMPO-conjugated LCNPs to protect live cells from oxidative stress by effectively scavenging ROS and reducing lipid peroxidation.

Nanoparticle-based delivery of nitric oxide for therapeutic applications.

Nitric oxide (NO), a low molecular weight signaling molecule, plays critical roles in both cellular health and disease. There is continued interest in new modalities for the controlled therapeutic delivery of NO to cells and tissues. The physicochemical properties of NO (including its short half-life and on-demand synthesis at the point of function), however, pose considerable challenges for its specific and efficient delivery. Recently, a number of nanoparticle (NP)-based systems are described that address some of these issues by taking advantage of the unique attributes of the NP carrier to effect efficient NO delivery. This review highlights the progress that has been made over the past 5 years in the use of various constructs for the therapeutic delivery of NO.

This review details progress made over the past 5 years in the implementation of various nanoparticle (NP) bioconjugates for the therapeutic delivery of nitric oxide. Various NP formulations including liposomes, polymeric NPs, and hard NPs such as AuNPs and upconversion NPs are covered and we discuss the inherent advantages and challenges in using these materials for the controlled delivery of nitric oxide to cells and tissues.

Quercetin Decreases Corneal Haze In Vivo and Influences Gene Expression of TGF-Beta Mediators In Vitro.

We have previously reported the flavonoid, quercetin, as a metabolic regulator and inhibitor of myofibroblast differentiation in vitro. Our current study evaluated the effects of topical application of quercetin on corneal scar development using two different animal models followed by RNA analysis in vitro. Wild-type C57BL/6J mice were anesthetized and the corneal epithelium and stroma were manually debrided, followed by quercetin (0.5, 1, 5, or 50 mM) or vehicle application. Corneal scarring was assessed for 3 weeks by slit lamp imaging and clinically scored. In a separate animal study, six New Zealand White rabbits underwent lamellar keratectomy surgery, followed by treatment with 5 mM quercetin or vehicle twice daily for three days. Stromal backscattering was assessed at week 3 by in vivo confocal microscopy. In mice, a single dose of 5 mM quercetin reduced corneal scar formation. In rabbits, stromal backscattering was substantially lower in two out of three animals in the quercetin-treated group. In vitro studies of human corneal fibroblasts showed that quercetin modulated select factors of the transforming growth factor-ß (TGF-ß) signaling pathway. These results provide evidence that quercetin may inhibit corneal scarring. Further studies in a larger cohort are required to validate the efficacy and safety of quercetin for clinical applications.

Cobalt(III)- and rhodium(III)-porphyrin complexes for the effective degradation of fentanyl: Biological inactivation and mechanistic insights.

Cobalt(III) and rhodium(III) complexes containing the water-soluble porphyrin ligand meso-tri(4-sulfonatophenyl)mono(4-carboxyphenyl)porphine (C1S3TPP), [Rh(C1S3TPP)]Nax•nH2O (1) and [Co(C1S3TPP)]Nax•nH2O (2) were prepared from the direct reaction of free porphyrin and metal chloride salts in refluxing MeOH/DMF or EtOH/H2O. Compounds 1 and 2 were characterized using UV-vis and 1H NMR spectroscopies, and high-resolution mass spectrometry. Cell culture based assays of opioid receptor activation showed that while the rhodium complex reduced fentanyl opioid activity 113-fold to an IC50 value of 1.7 µM, the cobalt complex reduced fentanyl activity by 160-fold to an IC50 value of 2.4 µM. An oxidative mechanism for fentanyl breakdown is proposed.

Sensing Nitric Oxide in Cells: Historical Technologies and Future Outlook.

Nitric oxide (NO) is a critical cell signaling molecule with important roles in both normal cellular physiology and pathology. Over the past 20 years, multiple sensing modalities have been developed for the intracellular synthesis (endogenous) and release (exogenous) of NO. In this review, we survey the historical progression of NO sensing platforms, highlight the current state of the art, and offer a forward-looking view of how we expect the field of NO sensing to develop in the context of recent advances in bio-nanotechnology and nanoscale cellular biosensors.

Solvent-assisted optical modulation of FRET-induced fluorescence for efficient conjugated polymer-based DNA detection.

The solvent effects were studied in fluorescence resonance energy transfer (FRET) from a cationic polyfluorene copolymer (FHQ, FPQ) to a fluorescein (Fl)-labelled oligonucleotide (ssDNA-Fl). Upon addition of dimethyl sulfoxide (DMSO), the optical properties of polymers and the probe dye were substantially modified and the FRET-induced PL signal was enhanced 3.8-37 times, relative to that in phosphate buffer solution (PBS). The hydrophobic interaction between polymers and ssDNA-Fl is expected to decrease in the presence of DMSO, which induces the weaker polymer/ssDNA-Fl complexation with longer intermolecular donor-acceptor separation and perturbs the competition between the FRET and PL quenching processes such as photo-induced charge transfer. The gradual decrease in Fl PL quenching with increasing the DMSO content was investigated by measuring the Stern-Volmer quenching constants (3.3-4.2 × 10(6) M(-1) in PBS, 0.56-1.1 × 10(6) M(-1) in 80 vol% DMSO) and PL lifetime of the excited Fl* in polymer/ssDNA-Fl (600 ps in PBS and 2120 ps in 80 vol% DMSO for FHQ/ssDNA-Fl) in PBS/DMSO mixtures. The substantially reduced PL quenching would amplify the resulting FRET Fl signal. The signal amplification in real DNA detection was also demonstrated with fluorescein-labelled PNA (probe PNA) in the presence of a complementary target DNA and noncomplementary DNA in aqueous DMSO solutions. This approach suggests a simple way of modifying the fine-structure of polymer/ssDNA-Fl and improving the detection sensitivity in conjugated polymer-based FRET bioassays.

Anionic Conjugated Polyelectrolytes for FRET-based Imaging of Cellular Membrane Potential.

We report a Förster resonance energy transfer (FRET)-based imaging ensemble for the visualization of membrane potential in living cells. A water-soluble poly(fluorene-cophenylene) conjugated polyelectrolyte (FsPFc10) serves as a FRET donor to a voltage-sensitive dye acceptor (FluoVolt™ ). We observe FRET between FsPFc10 and FluoVolt™ , where the enhancement in FRET-sensitized emission from FluoVolt™ is measured at various donor/acceptor ratios. At a donor/acceptor ratio of 1, the excitation of FluoVolt™ in a FRET configuration results in a three-fold enhancement in its fluorescence emission (compared to when it is excited directly). FsPFc10 efficiently labels the plasma membrane of HEK 293T/17 cells and remains resident with minimal cellular internalization for ~ 1.5 h. The successful plasma membrane-associated colabeling of the cells with the FsPFc10-FluoVolt™ donor-acceptor pair is confirmed by dual-channel confocal imaging. Importantly, cells labeled with FsPFc10 show excellent cellular viability with no adverse effect on cell membrane depolarization. During depolarization of membrane potential, HEK 293T/17 cells labeled with the donor-acceptor FRET pair exhibit a greater fluorescence response in FluoVolt™ emission relative to when FluoVolt™ is used as the sole imaging probe. These results demonstrate the conjugated polyelectrolyte to be a new class of membrane labeling fluorophore for use in voltage sensing schemes.

Nanoparticle-Mediated Visualization and Control of Cellular Membrane Potential: Strategies, Progress, and Remaining Issues.

The interfacing of nanoparticle (NP) materials with cells, tissues, and organisms for a range of applications including imaging, sensing, and drug delivery continues at a rampant pace. An emerging theme in this area is the use of NPs and nanostructured surfaces for the imaging and/or control of cellular membrane potential (MP). Given the important role that MP plays in cellular biology, both in normal physiology and in disease, new materials and methods are continually being developed to probe the activity of electrically excitable cells such as neurons and muscle cells. In this Review, we highlight the current state of the art for both the visualization and control of MP using traditional materials and techniques, discuss the advantageous features of NPs for performing these functions, and present recent examples from the literature of how NP materials have been implemented for the visualization and control of the activity of electrically excitable cells. We conclude with a forward-looking perspective of how we expect to see this field progress in the near term and further into the future.

Counterion effects on fluorescence energy transfer in conjugated polyelectrolyte-based DNA detection.

Cationic poly[9,9'-bis[6''-(N,N,N-trimethylammonium)hexyl]fluorene-co-alt-phenylene]s with five different counterions (CIs) were synthesized and studied as fluorescence resonance energy transfer (FRET) donors (D) to dye-labeled DNA (FRET acceptor, A). The polymers with different CIs show the same pi-conjugated electronic structure with similar absorption (lambda(abs) = approximately 380 nm) and photoluminescence (lambda(PL) = approximately 420 nm) emission spectra in water. The CIs accompanying the polymer chain are expected to affect the D/A complexation and modify the D-A intermolecular separation by acting as a spacer. Polymers with different CIs function differently as FRET excitation donors to fluorescein (Fl)-labeled single-stranded DNA (ssDNA-Fl). The FRET-induced Fl emission was enhanced significantly by the larger CI-exchanged polymers. The polymers with the CIs of tetrakis(1-imidazolyl)borate (FPQ-IB) and tetraphenylborate (FPQ-PB) showed a 2-4-fold enhancement in the FRET-induced signal compared with the polymer with bromide (FPQ-BR). The delayed FRET signal saturation and low association constants (K(a)) with ssDNA-Fl (3.53 x 10(6) M(-1) for FPQ-BR and 1.80 x 10(6) M(-1) for FPQ-PB) were measured for the polymers with larger CIs. The delayed acceptor saturation strengthens the antenna effect and reduces self-quenching of Fl by increasing the polymer concentration near Fl. The weak polymer/ssDNA-Fl association reduces the amount of energy-wasting charge transfer by increasing D-A intermolecular separation. The combined effects lead to increase the overall FRET-induced signal.

Active Cellular and Subcellular Targeting of Nanoparticles for Drug Delivery.

Nanoparticle (NP)-mediated drug delivery (NMDD) for active targeting of diseases is a primary goal of nanomedicine. NPs have much to offer in overcoming the limitations of traditional drug delivery approaches, including off-target drug toxicity and the need for the administration of repetitive doses. In the last decade, one of the main foci in NMDD has been the realization of NP-mediated drug formulations for active targeted delivery to diseased tissues, with an emphasis on cellular and subcellular targeting. Advances on this front have included the intricate design of targeted NP-drug constructs to navigate through biological barriers, overcome multidrug resistance (MDR), decrease side effects, and improve overall drug efficacy. In this review, we survey advancements in NP-mediated drug targeting over the last five years, highlighting how various NP-drug constructs have been designed to achieve active targeted delivery and improved therapeutic outcomes for critical diseases including cancer, rheumatoid arthritis, and Alzheimer's disease. We conclude with a survey of the current clinical trial landscape for active targeted NP-drug delivery and how we envision this field will progress in the near future.