The Effect of Oxidative Stress and Memantine-Incorporated Reactive Oxygen Species-Sensitive Nanoparticles on the Expression of N-Methyl-d-aspartate Receptor Subunit 1 in Brain Cancer Cells for Alzheimer's Disease Application.

The aim of this study is to fabricate reactive oxygen species (ROS)-sensitive nanoparticles composed of succinyl ß-cyclodextrin (bCDsu), memantine and thioketal linkages for application in Alzheimer's disease, and to investigate the suppression of N-methyl-d-aspartate (NMDA) receptor 1 (NMDAR1) in cells. Thioketal diamine was attached to the carboxyl group of bCDsu to produce thioketal-decorated bCDsu conjugates (bCDsu-thioketal conjugates) and memantine was conjugated with thioketal dicarboxylic acid (memantine-thioketal carboxylic acid conjugates). Memantine-thioketal carboxylic acid conjugates were attached to bCDsu-thioketal conjugates to produce bCDsu-thioketal-memantine (bCDsuMema) conjugates. SH-SY5Y neuroblastoma cells and U87MG cells were used for NMDAR1 protein expression and cellular oxidative stress. Nanoparticles of bCDsuMema conjugates were prepared by means of a dialysis procedure. Nanoparticles of bCDsuMema conjugates had small particle sizes less than 100 nm and their morphology was found to be spherical in transmission electron microscopy observations (TEM). Nanoparticles of bCDsuMema conjugates responded to H2O2 and disintegrated or swelled in aqueous solution. Then, the nanoparticles rapidly released memantine according to the concentration of H2O2. In an in vivo animal imaging study, thioketal-decorated nanoparticles labelled with fluorescent dye such as chlorin e6 (Ce6) showed that the fluorescence intensity was stronger in the brain than in other organs, indicating that bCDsuMema nanoparticles can efficiently target the brain. When cells were exposed to H2O2, the viability of cells was time-dependently decreased. Memantine or bCDsuMema nanoparticles did not practically affect the viability of the cells. Furthermore, a western blot assay showed that the oxidative stress produced in cells using H2O2 increased the expression of NMDAR1 protein in both SH-SY5Y and U87MG cells. Memantine or bCDsuMema nanoparticles efficiently suppressed the NMDAR1 protein, which is deeply associated with Alzheimer's disease. Fluorescence microscopy also showed that H2O2 treatment induced green fluorescence intensity, which represents intracellular ROS levels. Furthermore, H2O2 treatment increased the red fluorescence intensity, which represents the NMDAR1 protein, i.e., oxidative stress increases the expression of NMDAR1 protein level in both SH-SY5Y and U87MG cells. When memantine or bCDsuMema nanoparticles were treated in cells, the oxidative stress-mediated expression of NMDAR1 protein in cells was significantly decreased, indicating that bCDsuMema nanoparticles have the capacity to suppress NMDAR1 expression in brain cells, which has relevance in terms of applications in Alzheimer's disease.

Dynamic Nanocomposite Microgel Assembly with Microporosity, Injectability, Tissue-Adhesion, and Sustained Drug Release Promotes Articular Cartilage Repair and Regeneration.

Owing to the lack of blood vessels, nerves, and lymph, articular cartilage defect is difficult to self-repair. Although several cartilage tissue engineering products have been authorized for clinical use, there are still some problems such as large surgical wounds, weak adhesion with the host tissue, and the limited source of autologous chondrocytes. In this paper, a novel dynamic nanocomposite microgel assembly with excellent microporosity, injectability, tissue-adhesion, and sustained kartogenin (KGN) release is reported. Specifically, KGN-loaded cyclodextrin nanoparticles are synthesized through nanoemulsification and incorporated into bone marrow mesenchymal stem cell (BMSCs)-laden microgels via droplet-based microfluidics and photo-crosslinking, which are then bottom-up assembled via dynamic crosslinking between dopamine-modified hyaluronic acid and phenylboronic acid groups on microgel surface. Results reveal that the microgel assembly can avoid the cell endocytosis of nanoparticles, ensure the high BMSC viability during the regular cell culture, cryopreservation and injection process, promote the chondrogenic differentiation of BMSCs. In addition, animal expriment proves the newborn cartilages present the typical characteristics of articular cartilage. In brief, this microgel assembly not only offers convenience for clinical use (injectability, tissue adhesion) but also provides good microenvironments for chondrogenesis (controlled drug release, interconnected micropores), indicative of its promising application for cartilage repair and regeneration.

Inhibition of IL-1ß release from macrophages targeted with necrosulfonamide-loaded porous nanoparticles.

Inflammation is required for protective responses against pathogens and is thus essential for survival, but sustained inflammation can lead to diseases, such as atherosclerosis and cancer. Two important mediators of inflammation are the cytokines IL-1ß and IL-18, which are produced by myeloid cells of the immune system, including macrophages. These cytokines are released into the extracellular space through pores formed in the plasma membrane by the oligomerized protein gasdermin D (GSDMD). Necrosulfonamide (NSA) was recently identified as an effective GSDMD inhibitor and represents a promising therapeutic agent in GSDMD-dependent inflammatory diseases. Here, we targeted NSA to both mouse and human macrophages by using three different types of porous nanoparticles (NP), i.e. mesoporous silica (MSN), porous crosslinked cyclodextrin carriers (CD-NP), and a mesoporous magnesium-phosphate carrier (MPC-NP), all displaying high loading capacities for this hydrophobic drug. Cellular uptake and intracellular NSA delivery were tracked in time-lapse experiments by live-cell, high-throughput fluorescence microscopy, demonstrating rapid nanoparticle uptake and effective targeted delivery of NSA to phagocytic cells. Notably, a strong cytostatic effect was observed when a macrophage cell line was exposed to free NSA. In contrast, cell growth was much less affected when NSA was delivered via the nanoparticle carriers. Utilizing NSA-loaded nanoparticles, a successful concentration-dependent suppression of IL-1ß secretion from freshly differentiated primary murine and human macrophages was observed. Functional assays showed the strongest suppressive effect on human macrophages when using CD-NP for NSA delivery, followed by MSN-NP. In contrast, MPC-NP completely blocked the metabolic activity in macrophages when loaded with NSA. This study demonstrates the potential of porous nanoparticles for the effective delivery of hydrophobic drugs to macrophages in order to suppress inflammatory responses.

Nanoparticulate ß-Cyclodextrin with Gallium Tetraphenylporphyrin Demonstrates in Vitro and in Vivo Antimicrobial Efficacy against Mycobacteroides abscessus and Mycobacterium avium.

The emergence of drug-resistant pathogens causes the greatest challenge for drug development research. Recently, gallium(III)-based compounds have received great attention as novel antimicrobial agents against drug-resistant pathogens. Here, we synthesized a new ß-cyclodextrin Ga nanoparticle (CDGaTP) using Ga tetraphenylporphyrin (GaTP, a hemin analogue) and ß-cyclodextrin. The newly synthesized nanoparticle was nontoxic and efficient at a single dose, showing sustained drug release for 15 days in vitro. CDGaTP's activity with transferrin or lactoferrin was tested, and synergism in activity was observed against nontuberculosis mycobacteria (NTM), Mycobacterium avium (M. avium), and Mycobacteroides abscessus. Human serum albumin (HSA) decreased the efficacy of both GaTP and CDGaTP in a concentration-dependent manner. The NTMs incubated with GaTP or CDGaTP significantly produced reactive oxygen species (ROS), indicating potential inhibition of antioxidant enzymes, such as catalase. The single-dose CDGaTP displayed a prolonged intracellular inhibitory activity in an in vitro macrophage infection model against both NTMs. In addition, CDGaTP, not GaTP, was effective in a murine lung M. avium infection model when delivered via intranasal administration. These results suggest that CDGaTP provides new opportunities for the development of gallium-porphyrin based antibiotics.

Curcumin Encapsulated in Crosslinked Cyclodextrin Nanoparticles Enables Immediate Inhibition of Cell Growth and Efficient Killing of Cancer Cells.

The efficiency of anti-cancer drugs is commonly determined by endpoint assays after extended incubation times, often after days. Here we demonstrate that curcumin encapsulated in crosslinked cyclodextrin nanoparticles (CD-NP) acts extremely rapidly on cell metabolism resulting in an immediate and complete inhibition of cell growth and in efficient cancer-cell killing only few hours after incubation. This early onset of anti-cancer action was discovered by live-cell high-throughput fluorescence microscopy using an environmental stage. To date, only very few examples of covalently crosslinked nanoscale CD-based (CD-NP) drug carriers exist. Crosslinking cyclodextrins enables the adsorption of unusually high payloads of hydrophobic curcumin (762 µg CC/mg CD-NP) reflecting a molar ratio of 2.3:1 curcumin to cyclodextrin. We have investigated the effect of CD-NP encapsulated curcumin (CD-CC-NP) in comparison to free, DMSO-derived curcumin nanoparticles (CC-NP) on 4 different cell lines. Very short incubations times as low as 1 h were applied and cell responses after medium change were subsequently followed over two days. We show that cell proliferation is inhibited nearly immediately in all cell lines and that a cell- and concentration dependent cancer-cell killing occurs. Anti-cancer effects were similar with free and encapsulated curcumin, however, encapsulation in CD-NP drastically extends the long-term photostability and anti-cancer activity of curcumin. Curcumin-sensitivity is highest in HeLa cells reaching up to 90% cell death under these conditions. Sensitivity decreased from HeLa to T24 to MDA MB-231 cells. Strikingly, the immortalized non-cancerous cell line MCF-10A was robust against curcumin concentrations that were highly toxic to the other cell lines. Our results underline the potential of curcumin as gentle and yet effective natural anti-cancer agent when delivered solvent-free in stabilizing and biocompatible drug carriers such as CD-NP that enable efficient cellular delivery.

Oral insulin delivery by carboxymethyl-ß-cyclodextrin-grafted chitosan nanoparticles for improving diabetic treatment.

In this paper, a new oral insulin formulation, insulin-loaded carboxymethyl-ß-cyclodextrin-grafted chitosan nanoparticles (insulin/CMCD-g-CS NPs), was fabricated by ionic crosslinking technique. The therapeutic efficacy of new formulation was investigated in detail. Firstly, the CMCD-g-CS was synthesized by EDC-mediated esterification reaction. The prepared CMCD-g-CS exhibited favourable loading capacity and encapsulation efficiency of drug. The release experiment in vitro showed that the nanocarrier could efficiently protect encapsulated insulin at simulated gastric environment and release drug in the simulated colonic fluid. The insulin/CMCD-g-CS NPs effectively promoted drug internalization into Caco-2 cells and could reversibly open the tight junction between cells. The oral administration of insulin/CMCD-g-CS NPs could lastingly decrease blood sugar level in diabetic mice. The liver function study verified that the insulin/CMCD-g-CS NPs had not obvious toxicity to experimental mice. Therefore, the CMCD-g-CS could be an effective and safe oral insulin delivery carrier for future clinical application. A new biocompatible polysaccharide nanoparticle was fabricated as oral insulin delivery carrier for improving diabetic treatment.

Kinetics of γ-cyclodextrin nanoparticle suspension eye drops in tear fluid.

PURPOSE: We have developed nanoparticle γ-cyclodextrin dexamethasone (DexNP) and dorzolamide (DorzNP) eye drops that provide sustained high drug concentrations on the eye surface. To test these characteristics, we measured dexamethasone and dorzolamide levels in tear fluid in humans following eye drop administration. METHODS: Concentration of dexamethasone was measured by mass spectrometry. One drop of DexNP was instilled into one eye. Tear fluid was sampled with microcapillary pipettes at seven time-points after drop instillation. Control eyes received Maxidex(®) (dexamethasone). The same procedure was performed for dorzolamide with DorzNP and Trusopt(®) . RESULTS: Six subjects were included in each group. The peak concentration (µg/ml ± standard deviation) of dexamethasone for DexNP eye drops (636.6 ± 399.1) was up to 19-fold higher than with Maxidex(®) (39.3 ± 18.9) (p < 0.001). At 4 hr, DexNP was still 10 times higher than Maxidex(®) . In addition, DexNP resulted in about 30-fold higher concentration of dissolved dexamethasone in the tear fluid of extended time period allowing more drug to partition into the eye tissue. The overall concentration of dorzolamide was about 50% higher for DorzNP (59.5 ± 76.9) than Trusopt(®) (40.0 ± 76.7) (p < 0.05). CONCLUSION: The results indicate high and extended concentration of dissolved dexamethasone with DexNP, which can explain the greater and longer lasting effect of dexamethasone in the cyclodextrin nanoparticle drug delivery platform. Dexamethasone seems to fit the cyclodextrin nanoparticle suspension drug delivery platform with longer duration and higher concentrations in tear fluid than available commercial drops, while dorzolamide is less suitable.

A vision for cyclodextrin nanoparticles in drug delivery systems and pharmaceutical applications.

Cyclodextrins (CDs) have brought a revolution in the pharmaceutical field over the last decade. Natural and modified CDs (α-CD and ß-CD) have been studied and some have gained US FDA approval or achieved 'Generally Regarded as Safe' (GRAS) status. Another characteristic of CDs is the ease with which they can be induced to form supramolecular structures for its use in drug delivery. CDs, grafted or crosslinked with polymers, are now being developed into 'smart' systems for efficient targeted drug delivery, especially for hydrophobic drugs. Amphiphilic CDs have the ability to form nanospheres or nanocapsules via a simple nanoprecipitation technique. This review deals with different types of CDs, and their efficacy, physicochemical properties and transformation into nanoparticles with interesting in vitro and in vivo applications.

New formulation of an old drug in hypertension treatment: the sustained release of captopril from cyclodextrin nanoparticles.

Captopril (CAP) was the first angiotensin I-converting enzyme (ACE) inhibitor to be developed and is widely used in hypertension treatment. On the other hand, cyclodextrins (CDs) are cyclic oligosaccharides whose cone-shaped cavity allows formation of noncovalent inclusion complexes with appropriately sized guest molecules, thus modifying guest physical, chemical, and biological properties. Herein, the physicochemical characterization and in vivo ACE inhibition evaluation of seven CAP/CD complexes are reported. The inclusion complexes were prepared by spray-drying, freeze-drying, kneading, or lyophilization methods and characterized by nuclear magnetic resonance, Fourier-transformed infrared spectroscopy, X-ray diffraction, differential scanning calorimetry, and scanning electron microscopy techniques. In vivo assays compared CAP and CAP/CD complex administration (0.5 mg kg(-1) or 0.09 mg kg(-1), n = 4-7) to evaluate the ACE inhibition by continuous infusion of angiotensin I (30 ng 50 µL(-1) min(-1)) in conscious Wistar rats. The physicochemical analysis demonstrated complete amorphization and complexation between CAP and CDs, indicating the substitution of water molecules inside the CD cavity with CAP. During the infusion of angiotensin I, the administration of all CAP/CD complexes induced a reduction in mean arterial pressure similar to that observed upon CAP administration. The nanoparticles obtained by the kneading method (CAP/α-CD:KM) showed a potent and long-lasting inhibitory activity (∼22 hours) on the angiotensin I pressor effect. The results suggest that the inclusion complex of CAP and α-CD can function as a novel antihypertensive formulation that may improve therapeutic use of CAP by reducing its oral dose administration to once per day, thus providing better quality of life for almost 25% of the world's population who suffer from hypertension.