Stabilizing Coacervate by Microfluidic Engulfment Induced by Controlled Interfacial Energy.

Low interfacial energy, an intrinsic property of complex coacervate, enables the complex coacervate to easily encapsulate desired cargo substances, making it widely used in encapsulation applications. Despite this advantage, the low interfacial energy of the complex coacervate makes it unstable against mechanical mixing, and changes in pH and salt concentration. Hence, a chemical cross-linker is usually added to enhance the stability of the complex coacervate at the expense of sacrificing all intrinsic properties of the coacervate, including phase transition of the coacervate from liquid to solid. In this study, we observed an abrupt increase in the interfacial energy of the coacervate phase in mineral oil. By controlling the interfacial energy of the coacervate phase using a microfluidic device, we successfully created double engulfed PEG-diacrylate (PEGDA) coacervate microparticles, named DEPOT, in which the coacervate is engulfed in a cross-linked PEGDA shell. The engulfed coacervate remained as a liquid phase, retained its original low interfacial energy property to encapsulate the desired cargo substances, and infiltrated into the target site by a simple solvent exchange from oil to water.

pH-triggered surface charge-reversal nanoparticles alleviate experimental murine colitis via selective accumulation in inflamed colon regions.

In this study, we developed pH-triggered surface charge-reversal lipid nanoparticles (LNPs), loaded with budesonide, which could precisely deliver the drug to inflamed colon segments for the treatment of ulcerative colitis. Polyethyleneimine (PEI) was used to render LNPs cationic (PEI-LNPs), and Eudragit® S100 (ES) was coated on PEI-LNPs to obtain pH-triggered charge-reversal LNPs (ES-PEI-LNPs). ES coating avoided a burst drug release under acidic conditions mimicking the stomach and early small intestine environments and showed a sustained release in the colon. The surface charge of ES-PEI-LNPs switched from negative to positive under colonic conditions owing to pH-triggered removal of the ES coating. Bioimaging of the mouse gastrointestinal tract and confocal analysis of colon tissues revealed that ES-PEI-LNPs selectively accumulated in an inflamed colon. Furthermore, ES-PEI-LNPs mitigated experimental colitis in mice. These results suggest that the pH-triggered charge-reversal LNPs could be a promising drug carrier for ulcerative colitis therapy and other colon-targeted treatments.

Bacteria-Adhesive Nitric Oxide-Releasing Graphene Oxide Nanoparticles for MRPA-Infected Wound Healing Therapy.

A bacteria-infected wound can lead to being life-threatening and raises a great economic burden on the patient. Here, we developed polyethylenimine 1.8k (PEI1.8k) surface modified NO-releasing polyethylenimine 25k (PEI25k)-functionalized graphene oxide (GO) nanoparticles (GO-PEI25k/NO-PEI1.8k NPs) for enhanced antibacterial activity and infected wound healing via binding to the bacterial surface. In vitro antibacterial activity and in vivo wound healing efficacy in an infected wound model were evaluated compared with NO-releasing NPs (GO-PEI25k/NO NPs). Surface modification with PEI1.8k can enhance the ability of nanoparticles to adhere to bacteria. GO-PEI25k/NO-PEI1.8k NPs released NO in a sustained manner for 48 h and exhibited the highest bactericidal activity (99.99% killing) against methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Pseudomonas aeruginosa (MRPA) without cytotoxicity to L929 mouse fibroblast cells at 0.1 mg/mL. In the MRPA-infected wound model, GO-PEI25k/NO-PEI1.8k NPs showed 87% wound size reduction while GO-PEI25k/NO NPs showed 23% wound size reduction at 9 days postinjury. Masson trichrome and hematoxylin and eosin staining revealed that GO-PEI25k/NO-PEI1.8k NPs enhanced re-epithelialization and collagen deposition, which are comparable to healthy mouse skin tissue. GO-PEI25k/NO-PEI1.8k NPs hold promise as effective antibacterial and wound healing agents.

Tumor-Penetrable Nitric Oxide-Releasing Nanoparticles Potentiate Local Antimelanoma Therapy.

Although nitric oxide (NO) has been emerging as a novel local anticancer agent because of its potent cytotoxic effects and lack of off-target side effects, its clinical applications remain a challenge because of the short effective diffusion distance of NO that limits its anticancer activity. In this study, we synthesized albumin-coated poly(lactic-co-glycolic acid) (PLGA)-conjugated linear polyethylenimine diazeniumdiolate (LP/NO) nanoparticles (Alb-PLP/NO NPs) that possess tumor-penetrating and NO-releasing properties for an effective local treatment of melanoma. Sufficient NO-loading and prolonged NO-releasing characteristics of Alb-PLP/NO NPs were acquired through PLGA-conjugated LP/NO copolymer (PLP/NO) synthesis, followed by nanoparticle fabrication. In addition, tumor penetration ability was rendered by the electrostatic adsorption of the albumin on the surface of the nanoparticles. The Alb-PLP/NO NPs showed enhanced intracellular NO delivery efficiency and cytotoxicity to B16F10 murine melanoma cells. In B16F10-tumor-bearing mice, the Alb-PLP/NO NPs showed improved extracellular matrix penetration and spatial distribution in the tumor tissue after intratumoral injection, resulting in enhanced antitumor activity. Taken together, the results suggest that Alb-PLP/NO NPs represent a promising new modality for the local treatment of melanoma.

PEI/NONOates-doped PLGA nanoparticles for eradicating methicillin-resistant Staphylococcus aureus biofilm in diabetic wounds via binding to the biofilm matrix.

Wounds infected with methicillin-resistant Staphylococcus aureus (MRSA) biofilm represent a high risk in patients with diabetes. Nitric oxide (NO) has shown promise in dispersing biofilm and wound healing. For an effective treatment of MRSA biofilm-infected wounds, however, NO needs to be supplied to the biofilm matrix in a sustainable manner due to a short half-life and limited diffusion distance of NO. In this study, polyethylenimine/diazeniumdiolate (PEI/NONOate)-doped PLGA nanoparticles (PLGA-PEI/NO NPs) with an ability to bind to the biofilm matrix are developed to facilitate the NO delivery to MRSA biofilm-infected wound. In simulated wound fluid, PLGA-PEI/NO NPs show an extended NO release over 4 days. PLGA-PEI/NO NPs firmly bind to the MRSA biofilm matrix, resulting in a greatly enhanced anti-biofilm activity. Moreover, PLGA-PEI/NO NPs accelerate healing of MRSA biofilm-infected wounds in diabetic mice along with complete biofilm dispersal and reduced bacterial burden. These results suggest that the biofilm-binding NO-releasing NPs represent a promising NO delivery system for the treatments of biofilm-infected chronic wounds.

S-Nitrosoglutathione loaded poly(lactic-co-glycolic acid) microparticles for prolonged nitric oxide release and enhanced healing of methicillin-resistant Staphylococcus aureus-infected wounds.

Methicillin-resistant Staphylococcus aureus (MRSA)-infected wounds have become a significant clinical issue worldwide. Recently, nitric oxide (NO) has emerged as a potent antibacterial agent against MRSA infections and a wound-healing enhancer. Nevertheless, clinical applications of NO have been largely restricted by its gaseous state and short half-life. In this study, our aim was to develop S-nitrosoglutathione (GSNO, an endogenous NO donor)-loaded poly(lactic-co-glycolic acid) [PLGA] microparticles (GSNO-MPs) that release NO over a prolonged period, to accelerate the healing of MRSA-infected wounds with less frequent dosing. GSNO was successfully encapsulated into PLGA microparticles by a solid-in-oil-in-water emulsion solvent evaporation method. Scanning electron microscopy and X-ray diffraction analyses confirmed the successful fabrication of GSNO-MPs. The latter released NO in a prolonged manner over 7 days and exerted a remarkable antibacterial activity against MRSA in a concentration- and time-dependent manner. Moreover, GSNO-MPs had good antibacterial efficacy and were found to accelerate wound healing in a mouse model of MRSA-infected wounds. Therefore, NO-releasing MPs devised in this study may be a promising option for the treatment of cutaneous wounds infected by drug-resistant bacteria such as MRSA.

A three-dimensional assemblage of gingiva-derived mesenchymal stem cells and NO-releasing microspheres for improved differentiation.

Stem cell therapy is an attractive approach to bone tissue regeneration. Nitric oxide (NO) has been reported to facilitate osteogenic differentiation of stem cells. To enhance osteogenic differentiation of gingiva-derived mesenchymal stem cells (GMSCs), we designed a method for in situ delivery of exogenous NO to these cells. A NO donor, polyethylenimine/NONOate, was incorporated into poly(lactic-co-glycolic acid) microspheres to deliver NO to the cells for an extended period of time under in vitro culture conditions. A hybrid aggregate of GMSCs and NO-releasing microspheres was prepared by the hanging drop technique. Confocal microscopy revealed homogeneous arrangement of the stem cells and microspheres in heterospheroids. Western blot analysis and live-dead imaging showed no significant change in cell viability. Importantly, the in situ delivery of NO within the heterospheroids enhanced osteogenic differentiation indicated by a 1.2-fold increase in alkaline phosphatase activity and an approximately 10% increase in alizarin red staining. In addition, a low dose of NO promoted proliferation of the GMSCs in this 3D system. Thus, delivery of the NO-releasing microsphers to induce differentiation of stem cells within this three dimensional system may be one of possible strategies to direct differentiation of a stem cell-based therapeutic agent toward a specific lineage.

Nitric oxide-releasing poly(lactic-co-glycolic acid)-polyethylenimine nanoparticles for prolonged nitric oxide release, antibacterial efficacy, and in vivo wound healing activity.

Nitric oxide (NO)-releasing nanoparticles (NPs) have emerged as a wound healing enhancer and a novel antibacterial agent that can circumvent antibiotic resistance. However, the NO release from NPs over extended periods of time is still inadequate for clinical application. In this study, we developed NO-releasing poly(lactic-co-glycolic acid)-polyethylenimine (PEI) NPs (NO/PPNPs) composed of poly(lactic-co-glycolic acid) and PEI/diazeniumdiolate (PEI/NONOate) for prolonged NO release, antibacterial efficacy, and wound healing activity. Successful preparation of PEI/NONOate was confirmed by proton nuclear magnetic resonance, Fourier transform infrared spectroscopy, and ultraviolet/visible spectrophotometry. NO/PPNPs were characterized by particle size, surface charge, and NO loading. The NO/PPNPs showed a prolonged NO release profile over 6 days without any burst release. The NO/PPNPs exhibited potent bactericidal efficacy against methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa concentration-dependently and showed the ability to bind on the surface of the bacteria. We also found that the NO released from the NO/PPNPs mediates bactericidal efficacy and is not toxic to healthy fibroblast cells. Furthermore, NO/PPNPs accelerated wound healing and epithelialization in a mouse model of a MRSA-infected wound. Therefore, our results suggest that the NO/PPNPs presented in this study could be a suitable approach for treating wounds and various skin infections.

Intraocular Pharmacokinetics of Povidone-Iodine and Its Effects on Experimental Staphylococcus epidermidis Endophthalmitis.

PURPOSE: The purpose of this study was to investigate the pharmacokinetics and safety of intravitreal povidone-iodine (PVI) and its efficacy against experimental Staphylococcus epidermidis endophthalmitis. METHODS: In phase I, forty New Zealand white rabbits were divided into groups I and II and received intravitreal 0.1% and 0.3% PVI, respectively. Electroretinography (ERG) and histologic examinations were conducted at baseline, 6, and 12 hours. Half-life was determined using high-performance liquid chromatography. In phase II, after the induction of S. epidermidis endophthalmitis, 0.1% and 0.3% PVI were injected intravitreally once in groups A and B and three times every second day in groups C and D (n = 10 in each group). Electroretinographs, histologic examinations, and vitreous cultures were conducted on day 14. RESULTS: Electroretinography and histologic examinations did not reveal any notable retinal damage in phase I in either group. Half-lives were 3.27 and 3.58 hours in groups I and II, respectively. In phase II, all groups demonstrated marked improvement, compared to controls. Bacterial growth was found in four eyes in group A (20, 60, 60, and 70 colony forming units [CFU]) and in three eyes in group B (20, 40, and 60 CFU) but not in those belonging to groups C and D at day 14. Retinal damage with lymphocyte infiltration in the inner retinal layers was more common in groups A and B than in groups C and D. CONCLUSIONS: Half-life of PVI was approximately 3 hours in the vitreous. Repeated injection of intraocular PVI, even at low concentrations, is most likely to be effective for the treatment of bacterial endophthalmitis.

Colon-targeted delivery of budesonide using dual pH- and time-dependent polymeric nanoparticles for colitis therapy.

Single pH-dependent drug delivery systems have been widely used for colon-targeted delivery, but their efficiency is often hampered by the variation in gut pH. To overcome the limitation of single pH-dependent delivery systems, in this study, we developed and evaluated the therapeutic potential of budesonide-loaded dual pH/time-dependent nanoparticles (NPs) for the treatment of colitis. Eudragit FS30D was used as a pH-dependent polymer, and Eudragit RS100 as a time-dependent controlled release polymer. Single pH-dependent NPs (pH_NPs), single time-dependent NPs (Time_NPs), and dual pH/time-dependent NPs (pH/Time_NPs) were prepared using the oil-in-water emulsion method. The physicochemical properties and drug release profiles of these NPs in gastrointestinal (GI) tract conditions were investigated. The therapeutic potential and in vivo distribution of the NPs were evaluated in a dextran sulfate sodium (DSS)-induced colitis mice model. The pH/Time_NPs prevented a burst drug release in acidic pH conditions and showed sustained release at a colonic pH. The in vivo distribution study in the mice GI tract demonstrated that pH/Time_NPs were more efficiently delivered to the inflamed colon than pH_NPs were. Compared to the single pH_NPs-treated group, the pH/Time_NPs-treated group showed increased body weight and colon length and markedly decreased disease activity index, colon weight/length ratios, histological damage, and inflammatory cell infiltration in colon tissue. Our results demonstrate that the dual pH/time-dependent NPs are an effective oral colon-targeted delivery system for colitis therapy.

Enzyme/pH dual sensitive polymeric nanoparticles for targeted drug delivery to the inflamed colon.

Novel nanoparticles whose drug release profiles are controlled by both enzyme and pH were prepared for the colon-specific drug delivery using a polymeric mixture of enzyme-sensitive azo-polyurethane and pH-sensitive Eudragit S100 (ES-Azo.pu). The enzyme/pH dual sensitive nanoparticles were designed to release a drug based on a two-fold approach which specifically aimed to target drug delivery to the inflamed colon while preventing the burst release of drugs in the stomach and small intestine. Single pH-sensitive (ES) and dual sensitive (ES-Azo.pu) nanoparticles were prepared using an oil-in-water emulsion solvent evaporation method and coumarin-6 (C-6) was used as a model drug. The successful formation of ES and ES-azo.pu nanoparticles that have 214 nm and 244 nm in mean particle size, respectively, was confirmed by scanning electron microscopy and qNano. ES nanoparticles showed almost 100% of burst drug release at pH 7.4, whereas ES-Azo.pu nanoparticles prevented the burst drug release at pH 7.4, followed by a sustained release phase thereafter. Furthermore, ES-Azo.pu nanoparticles exhibited enzyme-triggered drug release in the presence of rat cecal contents obtained from a rat model of colitis. An in vivo localization study in rat gastrointestinal tract demonstrated that ES-Azo.pu nanoparticles were selectively distributed in the inflamed colon, showing 5.5-fold higher C-6 than ES nanoparticles. In conclusion, the enzyme/pH dual sensitive nanoparticles presented in this study can serve as a promising strategy for colon-specific drug delivery against inflammatory bowel disease and other colon disorders.

Size-controlled biodegradable nanoparticles: preparation and size-dependent cellular uptake and tumor cell growth inhibition.

Biodegradable nanoparticles with diameters below 1000nm are of great interest in the contexts of targeted delivery and imaging. In this study, we prepared PLGA nanoparticles with well-defined sizes of ∼70nm (NP70), ∼100nm (NP100), ∼200nm (NP200), ∼400nm (NP400), ∼600nm (NP600) and ∼1000nm (NP1000) using facile fabrication methods based on a nanoprecipitation and solvent evaporation techniques. The nanoparticles showed a narrow size distribution with high yield. Then the size-controlled biodegradable nanoparticles were used to investigate how particle size at nanoscale affects interactions with tumor cells and macrophages. Interestingly, an opposite size-dependent interaction was observed in the two cells. As particle size gets smaller, cellular uptake increased in tumor cells and decreased in macrophages. We also found that paclitaxel (PTX)-loaded nanoparticles showed a size-dependent inhibition of tumor cell growth and the size-dependency was influenced by cellular uptake and PTX release. The size-controlled biodegradable nanoparticles described in this study would provide a useful means to further elucidate roles of particle size on various biomedical applications.