Polycation Complexes Research Articles

Self-assembled thiophanate-methyl/star polycation complex prevents plant cell-wall penetration and fungal carbon utilization during cotton infection by Verticillium dahliae

No effective fungicides are available for the management of Verticillium dahliae, which causes vascular wilt disease. In this study, a star polycation (SPc)-based nanodelivery system was used for the first time to develop a thiophanate-methyl (TM) nanoagent for the management of V. dahliae. SPc spontaneously assembled with TM through hydrogen bonding and Van der Waals forces to decrease the particle size of TM from 834 to 86 nm. Compared to TM alone, the SPc-loaded TM further reduced the colony diameter of V. dahliae to 1.12 and 0.64 cm, and the spore number to 1.13 × 108 and 0.72 × 108 cfu/mL at the concentrations of 3.77 and 4.71 mg/L, respectively. The TM nanoagents disturbed the expression of various crucial genes in V. dahliae, and contributed to preventing plant cell-wall degradation and carbon utilization by V. dahliae, which mainly impaired the infective interaction between pathogens and plants. TM nanoagents remarkably decreased the plant disease index and the fungal biomass in the root compared to TM alone, and its control efficacy was the best (61.20 %) among the various formulations tested in the field. Furthermore, SPc showed negligible acute toxicity toward cotton seeds. To the best of our knowledge, this study is the first to design a self-assembled nanofungicide that efficiently inhibits V. dahliae growth and protects cotton from the destructive Verticillium wilt.

Revealing the Mechanism of Bilayer Heterogeneous Polyelectrolytes to Suppress the Self‐Discharge of Symmetric Supercapacitors

The electrochemical supercapacitors with high power density, long cycle life, and excellent safety represent one of the most promising energy storage devices for flexible and portable electronics, but their spontaneously rapid drop of open‐circuit voltage (self‐discharge) greatly limits their wide applications. Herein, a series of bilayer heterogeneous polyelectrolyts (BHPs) consisting of a polyanion complex and a polycation complex are designed, to regulate the self‐discharge performance of supercapacitors. The BHP‐based supercapacitors possess comparable energy storage properties with those of devices based on traditional homogeneous polymer electrolyte, but exhibit a unique and noteworthy suppressed effect on the self‐discharge performance of devices. The experimental results and theoretical simulation reveal that the zeta potential difference between the used polyanion/polycation complexes has effect on the self‐discharge rate of BHP‐based supercapacitor, and the electrostatic interaction between polyelectrolytes and the mobile counterions also greatly affects the self‐discharge performance of devices. Herein, the effect of molecular structures and their interaction of polyelectrolyte complexes in BHPs on the electrochemical performance of the resultant supercapacitors are systemmatically investigated, which provides a general strategy to design novel polymer electrolytes to build high‐performance flexible supercapacitors with long self‐discharge time.

A novel bilayer heterogeneous poly(ionic liquid) electrolyte for high-performance flexible supercapacitors with ultraslow self-discharge.

Flexible supercapacitors with high power density and long cyclic stability represent a promising candidate to be used as power supplies for portable electronics, but often suffer from the disadvantages of a limited working voltage and rapid self-discharge (spontaneous drop of open-circuit voltage). Here, we design a bilayer heterogeneous poly(ionic liquid) electrolyte (BHPE) consisting of a polycation complex and a polyanion complex with different zeta potentials to suppress the self-discharge of flexible symmetric supercapacitors. The resultant BHPE-based supercapacitors using active carbon/carbon nanotube composite electrodes exhibit a high working potential of 3.0 V and an energy density of 33 W h kg-1, which are comparable with those of devices obtained by using a homogeneous poly(ionic liquid) electrolyte (HPE). More significantly, the developed BHPE-based supercapacitor charged under forward bias exhibits a self-discharge time of 23.2 h, which is at least twice that of the device charged under reverse bias and is also much superior to those of HPE-based supercapacitors. The BHPE-based supercapacitors also possess excellent mechanical flexibility and stability, due to the stabilized interface contact between two layers of poly(ionic liquid)s.

Targeting nucleic acid-based therapeutics to tumors: Challenges and strategies for polyplexes.

The current medical reality of cancer gene therapy is reflected by more than ten approved products on the global market, including oncolytic and other viral vectors and CAR T-cells as ex vivo gene-modified cell therapeutics. The development of synthetic antitumoral nucleic acid therapeutics has been proceeding at a lower but steady pace, fueled by a plethora of alternative nucleic acid platforms (from various antisense oligonucleotides, siRNA, microRNA, lncRNA, sgRNA, to larger mRNA and DNA) and several classes of physical and chemical delivery technologies. This review summarizes the challenges and strategies for tumor-targeted nucleic acid delivery. Focusing primarily on polyplexes (polycation complexes) as nanocarriers, delivery options across multiple barriers into tumor cells are illustrated.

Weak polyanion and strong polycation complex based membranes: Linking aqueous phase separation to traditional membrane fabrication

In recent work, Aqueous Phase Separation (APS) based on pH change induced polyelectrolyte complexation has shown great potential for the preparation of sustainable polymeric membranes with tunable structures. Unfortunately, thus far this has only been possible with a single polyelectrolyte combination. In this work, we demonstrate that this APS approach extends beyond a single system by preparing sustainable membranes from polyelectrolyte complexes (PECs) of the weak polyanion poly(acrylic acid) (PAA) and the strong polycation poly(diallyldimethylammonium chloride) (PDADMAC). PE solutions are mixed in an acidic medium where PAA is uncharged, and then this mixture is cast and immersed in a coagulation bath at a pH where PAA becomes charged and able to form a PEC with the oppositely charged PDADMAC. Since this process includes both phase separation and PE complexation, it is expected that membrane structure and performance is influenced by a combination of many factors. Casting solution pH, PAA molecular weight, and coagulation bath pH all directly affect the phase separation behavior of PAA/PDADMAC complexes in ways similar to conventional nonsolvent induced phase separation (NIPS). In addition, coagulation bath salinity and PE mixing ratio influence the complexation behavior. Through tuning of all these parameters it is possible to create a wide variety of membrane structures, ranging from nodular symmetrically porous membranes, to asymmetric membranes with cellular pores and in some cases dense top layers. The nodular membranes show good performance as microfiltration membranes with excellent oil retention (>95% for 3–4 µm droplets) and good water permeances. However for the cellular membranes, filtration led to collapse of the porous structure, emphasizing the importance of PE selection for membrane applications.

Scale-up synthesis of a polymer designed for protein therapy

Here we report optimization and scale-up of a biodegradable polycation, poly(ethylene argininylaspartate diglyceride) (PEAD) for the purpose of clinical translation. This polycation complexes heparin to form a coacervate. The resultant complex coacervate acts as a vehicle for controlled delivery of therapeutic proteins and through the bioactivity of heparin, greatly increases the half lives of many therapeutic proteins. In an effort to translate this drug delivery platform commercially and clinically, it is necessary to consider the scale of the synthesis, the environmental impact and the cost of the production. Therefore, we substituted the more expensive and hazardous solvents in the reactions, optimized the reaction parameters for a higher yield, scaled up the synthesis capacity up to hundreds of grams per batch and reduced the polymer cost by nearly 90%. The resultant PEAD bears more positive charges than the counterpart made using our previous method, and thus less polymer is needed to complex heparin for coacervation. The material demonstrated good cytocompatibility with human umbilical vein endothelial cells at concentrations up to 5 mg/ml, nearly 500-times more cytocompatible than the commercially resourced polyethyleneimine. Basic fibroblast growth factor was loaded in the coacervate with loading efficiency above 99%. The cargo steadily released approximately 1% over 37 days in vitro without an initial burst, indicating a highly stable coacervate for sustained release. This study paves the way for the translation of this delivery platform and may inform the scale-up of similar polymers.

Effect of pDNA Complex Particle Size on Gene Expression: Difference between In Vitro and In Vivo Experiments

Effect of pDNA Complex Particle Size on Gene Expression: Difference between In Vitro and In Vivo Experiments We have developed a novel gene-transfection system comprising very small (70-150 nm) plasmid/polyethyleneimine/hyaluronan (HA) ternary complexes having negative surface charge, and reduced nonspecific interactions with bio-components. They showed high in vivo gene expression efficiency. However, in in vitro transfection experiments, such small ternary complexes often demonstrated rather much lower gene expression than the conventional DNA/ polycation complex. In this study, we investigated the influence of the complex particle size on gene expression efficiency, focusing on the difference between in vitro and in vivo transfection. Size of the DNA complex particles strongly depended on the concentration of the mixing solutions, and the higher concentration resulted in the larger particles. As for the in vitro transfection, DNA/ polyethyleneimine binary complexes could readily be bound to the cells through the electrostatic attractions, regardless of their size. Smaller binary complexes particles showed higher gene expression than the larger ones, likely due to their high internalization efficiency. On the other hand, small DNA/polyethyleneimine/HA ternary complexes having negative surface charge kept dispersing in the medium away from the cells, and demonstrated lower in vitro gene transfection efficiency than larger ones. In regards to in vivo transfection after intravenous injection, only small ternary complexes showed high gene expression, because reduced nonspecific interactions and small size of the complex particles are required to be distributed in the body, and accumulated in tumor. The superiority and inferiority of the vectors for in vivo transfection cannot be always correctly predicted by the results of in vitro transfection.

Chemistry specificity of DNA-polycation complex salt response: a simulation study of DNA, polylysine and polyethyleneimine.

In this work, the chemistry specific stability determining factors of DNA-polycation complexes are examined by performing all-atom molecular dynamics simulations. To this end, we conduct a systematic variation of polycation line charge through polyethyleneimine (PEI) protonation and polycation chemistry via comparison with poly-l-lysine (PLL). Our simulations show that increasing line charge of the polycation alone does not lead to more salt tolerant complexes. Instead, the effective charge compensation by the polycation correlates with the increased stability of the complex against additional salt. The salt stability of PEI-DNA complexes also links to the proton sponge property of weak polycations, commonly assumed to be behind the effectivity of PEI as a gene delivery vector. Examination of the complexes reveals the mechanism behind this behaviour; more Cl(-) ions are attracted by the protonated complexes but, in contrast to the common depiction of the proton sponge behaviour, the ion influx does not cause swelling of the complex structure itself. However, PEI protonation leads to release of PEI while DNA remains tightly bound to the complex. Jointly, these findings shed light on the stability determining factors of DNA-polycation complexes, raise charge distribution as an important stability determining contributor, and indicate that the effectivity of PEI in gene delivery is likely to result from the freed PEI facilitating gene transfection.

Phase separation of siRNA–polycation complex and its effect on transfection efficiency

Polycations and cationic lipids have been widely employed as non-viral vectors for siRNA delivery. However, the stability of siRNA–polycation complexes is still rarely reported. Using highly charged dendrimer poly(amidoamine) (PAMAM) as an example, we studied the complexation of siRNA with a polycation by laser light scattering and atomic force microscopy. PAMAM forms a complex of more than 100 nm in diameter with siRNA right after mixing. However, the complex is not stable. It experiences an interesting kinetic process of aggregation, followed by phase separation. The rate of phase separation is related to the chemical composition of the polycations. Gene expressions on A549 cells and Hela cells demonstrate that the transfection efficiency decreases with the kinetics of the complexes. Cross-linking of the primary amino group of the PAMAM stabilizes the siRNA complex and causes the transfection results to be steady with time.

Structure, Dynamics, and Energetics of siRNA−Cationic Vector Complexation: A Molecular Dynamics Study

The design and synthesis of safe and efficient nonviral vectors for gene delivery has attracted significant attention in recent years. Previous experiments have revealed that the charge density of a polycation (the carrier) plays a crucial role in complexation and the release of the gene from the complex in the cytosol. In this work, we adopt an atomistic molecular dynamics simulation approach to study the complexation of short strand duplex RNA with six cationic carrier systems of varying charge and surface topology. The simulations reveal detailed molecular-level pictures of the structures and dynamics of the RNA-polycation complexes. Estimates for the binding free energy indicate that electrostatic contributions are dominant followed by van der Waals interactions. The binding free energy between the 8(+)polymers and the RNA is found to be larger than that of the 4(+)polymers, in general agreement with previously published data. Because reliable binding free energies provide an effective index of the ability of the polycationic carrier to bind the nucleic acid and also carry implications for the process of gene release within the cytosol, these novel simulations have the potential to provide us with a much better understanding of key mechanistic aspects of gene-polycation complexation and thereby advance the rational design of nonviral gene delivery systems.

In vivo magnetic resonance imaging of endogenous neuroblasts labelled with a ferumoxide–polycation complex

Neurogenesis occurs at the subependymal zone (SEZ) of the adult brain. Neural progenitor cells give rise to neuroblasts, which migrate to the olfactory bulb (OB) via the rostral migratory stream (RMS). Development of methods capable of labelling and tracking these cells in vivo would be of great benefit to the understanding of neuroblast migration away from the SEZ under normal and pathological conditions. In this study, we demonstrate that endogenous neuroblasts can be labelled in vivo with an MRI contrast agent and that they can be visualised using MRI. We compared two labelling strategies: intraventricular injection of the ferumoxide Endorem, with or without the transfection agent protamine sulphate. Administration of Endorem alone resulted in its distribution outside of the ventricle and into the periventricular space after 48 h. In contrast, we observed that intraventricular injection of Endorem complexed to protamine sulphate – forming the FePro complex – is restricted to the ventricular walls after 48 h. The FePro complex successfully labelled Doublecortin + neuroblasts in vivo up to 28 days post-injection. FePro-labelled neuroblasts in the RMS could be visualised using MRI in vivo and ex vivo on a 2.35 T MRI system, and FePro-labelled cells were identified in the OB on a 9.4 T MRI system. This study demonstrates the feasibility of in vivo imaging of endogenous neuroblast migration using MRI.

Application of Polyammonium Cations to Enzyme-immobilized Electrode: Voltammetric Behavior of Polycation-hexacyanoferrate Anion Complexes and Applicability as Electron-Transfer Mediator

Cyclic voltammograms of the aqueous solution containing polyammonium cations and the [Fe(CN)6]4-. anion at a plastic-formed carbon or platinum electrode were recorded. Polyammonium salts comprising quaternary ammonium in the main chain or sub chain and those comprising primary ammonium were tested. Under certain conditions, well-developed anodic and cathodic peak currents with the midpoint potentials different from that when the polyammonium cation is absent, that is, the midpoint potential of [Fe(CN)6]3- + e- = [Fe(CN)6]4- were exhibited, indicating that the [Fe(CN)6]4-/3- anionic species associate with the polyammonium cations tested to form the polycation-[Fe(CN)6]4-/3- complex species. The polycation complex species can be easily localized around the electrode surface with a dialysis membrane. An application of the polycation complex species as an electron-transfer mediator in the catalytic electrode with a redox enzyme was examined.

Revascularization of rat fasciocutaneous flap using CROSSEAL® with VEGF protein or plasmid DNA expressing VEGF

Fasciocutaneous tissue transfer is a common reconstructive procedure. Revascularization of flap tissue is an important component of tissue healing. Gene therapy offers an avenue through which the period of pedicle vascular dependency can be reduced. Rat fasciocutaneous flaps were elevated and a two-hour ischemic time induced. Polycation complex (jet PEI) and human fibrin sealant CROSSEAL was applied between flap and underlying abdominal tissues. Group 1 (six rats) was the control; Group 2 (seven rats) had vascular endothelial growth factor (VEGF) protein applied; Group 3 (seven rats) had plasmid DNA expressing VEGF applied. Vascular pedicles were ligated on postoperative day 5, percentage flap survival evaluated on day 7. All flaps survived initial ischemia. Mean +/- SD percentage area of the flap that survived was 28.1 +/- 12.4 (Group 1), 71.6 +/- 16.2 (Group 2), and 77.5 +/- 12.7 (Group 3) (P < 0.001, Group 1-3, 2-3). No differences were observed between Groups 2 and 3. Locally administered VEGF protein or plasmid DNA expressing VEGF enhanced survival of fasciocutaneous flaps.

A convenient and adjustable surface-modified complex containing poly-L-glutamic acid conjugates as a vector for gene delivery

In order to quantify the amount of ligands or poly(ethylene glycol) (PEG) on each vector, here we developed a system in which poly-L-glutamic acid (PLG) was used as surface modification loading backbone, to which one PEG (MW 5000, 10000, 20000) or epidermal growth factor (EGF) was linked. The PLG conjugates can electro-statically adsorb upon DNA/ polycation complex with positive charge, and, the amount of EGF or PEG on the surface of complexes could be varied. We have made a series of complexes containing the various PLG conjugates and examined their physicochemical properties, and made a comparison of properties and transfection efficiency between these complexes. EGF- and PEG-modified complexes showed 10-25-folds higher cell transfection efficiency than unmodified complexes in medium with or without serum.

Photoinduced in situ formation of various F‐actin assemblies with a photoresponsive polycation

We developed a novel in situ method for the control of F-actin assembly by using a synthetic photoresponsive polycation. The photoresponsive polycation mainly comprises a water-soluble cationic monomer and also contains a small amount of the monomer of a triphenylmethane leucohydroxide derivative (20 mol %), which is a well-known photochromic molecule that can be cationized in aqueous solution by ultra violet (UV) irradiation, thereby causing an increase in the total charge on the photoresponsive polycation. Thus, by exposure to UV radiation in aqueous solution, F-actin and the photoresponsive polycation start assembling into F-actin/photoresponsive polycation complexes of various morphologies such as bundles, coils, and networks, depending upon the concentrations of both the F-actin and salt. Further, localized UV irradiation can be applied in order to control the local formation of F-actin/photoresponsive polycation complexes. Thus, this technique provides a novel method for the spatiotemporal control of F-actin assembly and can be applied to investigate the unknown characteristics of F-actin.

Factors Controlling Particle Size during Nebulization of DNA–Polycation Complexes

Pulmonary gene therapy has the potential to treat or cure respiratory diseases such as cystic fibrosis. Much work has focused on the delivery of genes to the lung using viral vectors with varying degrees of success. Viral vectors are problematic and undesirable for use in the lung because they can provoke an acute immune response. This study has focused on the characterization of nonviral, polymer-based gene vectors for use with nebulizers. Calf thymus DNA has been used as a model, and was complexed with each of the three polycations; 22 kDa linear polyethyleneimine, 25 kDa branched polyethyleneimine, and 29.5 kDa polylysine using water, glucose solution, and phosphate-buffered saline (PBS) as carrier liquids. Fourier transform infrared spectroscopy has shown that the DNA retains the B form during the complex formation. The complexes prepared at N:P ratios of 10, have been nebulized using a vibrating plate nebulizer and the particle size and Zeta potentials measured before and after nebulization. The particle size distributions of the DNA complexes prepared in water and glucose solution were unimodal before and after nebulization with a small increase in particle size following nebulization. Choice of complexing polymer is shown to have only a small effect on particle size with the dominant effect coming from the ionic character of the dispersion fluid. Complexes prepared in PBS, although originally unimodal, showed pronounced agglomeration on nebulization. With all polymers in water or glucose solution, the Zeta potential increases after nebulization, but with PBS as the carrier liquid the potential falls and is clearly associated with the observed agglomeration. Gel electrophoresis shows that the complexing polymers protect the DNA through the nebulization process in all cases.

Dissociation of DNA–polycation complexes by polyanions and polyampholytes

Both polyanions and polyampholytes cause the dissociation of DNA–polycation complexes in experiments. To elucidate their mechanisms, Monte Carlo simulations have been conducted on a simple model, with DNA modeled as an infinite charged cylinder and other polyions treated as charged Shish–Kebab rods. Our results show that a highly charged polyanion is required to separate a polycation from a DNA. However, for a diblocked polyampholyte, its net dipole induces a higher probability to bridge a DNA and a polycation. Thus, the loosening mechanisms are found to be markedly different between polyanions and polyampholytes.

Study of Interpolymer Complexes of Oppositely Charged Macromolecules with Different Affinity to Solvent

We consider the new type of water-soluble stoichiometric interpolymer polyelectrolyte complexes (IPEC) consisting of macromolecules having different affinities to solvent and different densities of ionic groups along the chain. It was proposed that complexes of such macromolecules have a core−shell structure. The inner part of complex with a radius r contains monomer units of both macromolecules, while exterior part with a radius R consists exclusively of monomer units of hydrophilic macromolecules. With such complex organization, the external charged hydrophilic shell effectively protects the complex from precipitation and fusion with other complex assemblies. These theoretical conclusions allow to explain the experimental data on the stability of IPECs containing poly(l-lysine), poly(l-lysine citramide), poly(l-lysine citramide imide), and poly(acrylic acid), and can be of interest in the field of DNA−polycation complexes aimed at transfecting genes to cell nuclei.

Condensation of nonstochiometric DNA/polycation complexes by divalent cations

This study found that divalent cations induced the further condensation of partially condensed DNA within nonstochiometric polycation complexes. The addition of a few mmol of a divalent cation such as calcium reduced by half the inflection point at which DNA became fully condensed by poly-L-lysine (PLL) and a variety of other polycations. The effect on DNA condensation was initially observed using a new method, which is based on the concentration-dependent self-quenching of fluorescent moieties (e.g., rhodamine) covalently linked to the DNA backbone at relatively high densities. Additional analyses, which employed ultracentrifugation, dynamic light scattering, agarose gel electrophoresis, and atomic force microscopy, confirmed the effect of divalent cations. These results provide an additional accounting of the process by which divalent cations induce greater chromatin compaction that is based on the representation of chromatin fibers as a nonstoichiometric polyelectrolyte complex. They also offer a new approach to assemble nonviral vectors for gene therapy.

DNA–polycation complexes Effect of polycation structure on physico-chemical and biological properties

The purpose of the study was to investigate the influence of cationic polymer structure on the formation of DNA-polycation complexes and their transfection activity. Primary, tertiary, and quaternary polyamines with molecular masses ranging from 8000 to 200,000 were investigated. DNA-cationic polymer interaction was characterized by low gradient viscometry, dynamic light scattering, circular dichroism, UV spectrometry, flow birefringence, DNA electrophoresis, and electron microscopy. Transfection activity of the complexes was evaluated by the expression of reporter gene (beta-galactosidase) and using synthetic FITC-labelled oligonucleotides. Complex formation was found to be dependent on the structure and molecular weight of the polymer and the ionic strength of the solution. Secondary DNA structure in complexes was not disrupted, and DNA was protected from protonation. Cell lines of different origin were used for testing of transfection activity of the complexes. The sensitivity of the cells to transfection was established to be highly dependent on the cell line. DNA-polycation complexes are non-toxic according to MTT. Polyallylamine, and polydimethylaminoethylmethacrylate were found to be the most promising polycations for gene delivery. Transfection efficacy of their complexes with DNA to T-98G cells reaches up to 90-100%. It was found that optimal molecular mass of polydimethylaminoethylmethacrylate is in the range of 8000-50,000 Da.