In vitro model to study different nanoformulations in human nasal epithelial cell line (RPMI2650)

Parkinson's disease (PD) is a progressive neurological disorder that affects movements and is characterized by symptoms such as tremors, rigidity, and slowness of movement and it is caused by the degeneration of dopaminergic neurons in the brain. The idea behind this study is to use the nasal route as access to the Central Nervous System to deliver the neurotransmitter dopamine (DA) in order to bypass the blood-brain barrier. In this perspective, the first barrier that a drug has to cross is the nasal epithelium; therefore, in this study, we decided to use the RPMI2650 human cell line, which is often chosen as a model of the nasal epithelial barrier and for this reason has been shown to be suitable for use in drug permeation studies. On this cell line, we conducted in vitro viability tests using drug delivery systems composed of solid lipid nanoparticles (SLNs) loaded with DA and grape seed extract (GSE)1 with antioxidant function, synthesized with the aim of reducing oxidative stress and finding a remedy for neurotransmitter deficiency in dopaminergic neurons of PD patients. Cell viability assays were conducted using two different methods, Resazurin and MTT. Resazurin is a blue dye commonly used in cell biology experiments to monitor the growth or viability of cells. MTT, or 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, is another commonly used dye in biological assays; it is a yellow dye that is reduced by living cells to form a purple formazan product and is often used as a measure of cell viability, metabolic activity, or proliferation. Some different formulations of SLNs containing DA and GSE have been synthesized and used: DA-SLNs co-encapsulating (CO) GSE, DA-SLNs adsorbing (ADS) GSE, DA-SLNs CO and ADS GSE in gel Poloxamer/Carbopol and AlgOx/Hydroxypropyl Methylcellulose, DA-SLNs CO and ADS GSE in cryoprotectant sucrose and methyl beta cyclodextrin, DA-FITC-SLNs CO and ADS GSE, DA-SLNs without GSE. After treatment for 6, 12, 24 hours with various nanoformulations, no significant decrease in viable cells number was observed as compared to control condition

Dopamine and Antioxidant Grape Seed Extract loaded chitosan nanoparticles: A preliminary in vitro characterization

Neuronal cell model line SHSY-5Y is extensively adopted when in vitro investigations are related to Parkinson disease (PD) application. Herein, chitosan nanoparticles (CS NPs) were formulated for the co-administration of dopamine (DA) and Grape Seed Extract (GSE) with the aim to gain insight into the interactions occurring between SHSY-5Y and NPs. Following the ionic gelation technique, the mean particle size of the NPs resulted in the range 310–330 nm and the zeta measurements were in the range +16.4 – +35.5 mV. The presence of CS chains on the surface suggested by positive zeta values was also confirmed by FT-IR analysis, whereas storage stability studies upon different temperatures evidenced that, although aggregation occurred, DA autoxidation was prevented because no black suspensions were detected over the time, irrespectively of the temperature assayed. From a biological viewpoint, release studies of CS NPs loaded with DA and GSE showed that in SHSY-5Y cell lines DA accumulation was time-dependent, irrespectively of the presence of GSE. Furthermore, ROS levels and carbonylated proteins both decreased in SHSY-5Y cell line once NPs administering both DA and GSE were incubated, suggesting a significative reduction of oxidative stress which plays a significative role for PD development

Slightly viscous dispersions of mucoadhesive polymers as vehicles for nasal administration of dopamine and grape seed extract-loaded solid lipid nanoparticles

With the aim to find an alternative vehicle to the most used thermosensitive hydrogels for efficient nanotechnology-based nose-to-brain delivery approach for Parkinson’s disease (PD) treatment, in this work we evaluated the Dopamine (DA) and the antioxidant grape seed-derived pro-anthocyanidins (Grape Seed Extract, GSE) co-loaded solid lipid nanoparticles (SLNs) put in slight viscous dispersions (SVDs). These SVDs were prepared by dispersion in water at low concentrations of mucoadhesive polymers to which SLN pellets were added. For the purpose, we investigated two polymeric blends, namely Poloxamer/Carbopol (PF-127/Carb) and oxidized alginate/Hydroxypropylmethyl cellulose (AlgOX/HPMC). Rheological studies showed that the two fluids possess Newtonian behaviour with a viscosity slightly higher that water. The pH values of the SVDs were mainly within the normal range of nasal fluid as well as almost no osmotic effect was associated to both SVDs. All the SVDs were capable to provide DA permeation through nasal porcine mucosa. Moreover, it was found that PF- 127/Carb blend possesses penetration enhancer capability better than the Alg OX/HPMC combination. Flow cytometry studies demonstrated the uptake of viscous liquids incorporating fluorescent SLNs by human nasal RPMI 2650 cell in time-dependent manner. In conclusion, the SVD formulations may be considered promising alternatives to thermosensitive hydrogels strategy. Moreover, in a broader perspective, such SVD formulations may be also hopeful for treating various neurological diseases beyond PD treatment

Solid Lipid Nanoparticles Containing Dopamine and Grape Seed Extract: Freeze-Drying with Cryoprotection as a Formulation Strategy to Achieve Nasal Powders

(1) Background: DA-Gelucire® 50/13-based solid lipid nanoparticles (SLNs) administering the neurotransmitter dopamine (DA) and the antioxidant grape-seed-derived proanthocyanidins (grape seed extract, GSE) have been prepared by us in view of a possible application for Parkinson’s disease (PD) treatment. To develop powders constituted by such SLNs for nasal administration, herein, two different agents, namely sucrose and methyl-β-cyclodextrin (Me-β-CD), were evaluated as cryoprotectants. (2) Methods: SLNs were prepared following the melt homogenization method, and their physicochemical features were investigated by Raman spectroscopy, Scanning Electron Microscopy (SEM), atomic force microscopy (AFM) and X-ray Photoelectron Spectroscopy (XPS). (3) Results: SLN size and zeta potential values changed according to the type of cryoprotectant and the morphological features investigated by SEM showed that the SLN samples after lyophilization appear as folded sheets with rough surfaces. On the other hand, the AFM visualization of the SLNs showed that their morphology consists of round-shaped particles before and after freeze-drying. XPS showed that when sucrose or Me-β-CD were not detected on the surface (because they were not allocated on the surface or completely absent in the formulation), then a DA surfacing was observed. In vitro release studies in Simulated Nasal Fluid evidenced that DA release, but not the GSE one, occurred from all the cryoprotected formulations. Finally, sucrose increased the physical stability of SLNs better than Me-β-CD, whereas RPMI 2650 cell viability was unaffected by SLN-sucrose and slightly reduced by SLN-Me-β-CD. (4) Conclusions: Sucrose can be considered a promising excipient, eliciting cryoprotection of the investigated SLNs, leading to a powder nasal pharmaceutical dosage form suitable to be handled by PD patients

Dopamine - and Grape-Seed-Extract-Loaded Solid Lipid Nanoparticles: Interaction Studies between Particles and Differentiated SH-SY5Y Neuronal Cell Model of Parkinson’s Disease

Parkinson’s disease (PD) is a prevalent neurodegenerative disorder, primarily associated with do-paminergic neuron depletion in the Substantia Nigra. Current treatment focuses on compensating for dopamine (DA) deficiency, but the blood–brain barrier (BBB) poses challenges for effective drug delivery. Using differentiated SH-SY5Y cells, we investigated the co-administration of DA and the antioxidant Grape Seed Extract (GSE) to study the cytobiocompability, the cytoprotection against the neurotoxin Rotenone, and their antioxidant effects. For this purpose, two solid lipid nanoparticle (SLN) formulations, DA-co-GSE-SLNs and GSE-ads-DA-SLNs, were synthesized. Such SLNs showed mean particle sizes in the range of 187–297 nm, zeta potential values in the range of −4.1–−9.7 mV, and DA association efficiencies ranging from 35 to 82%, according to the formulation examined. The results showed that DA/GSE-SLNs did not alter cell viability and had a cytoprotective effect against Rotenone-induced toxicity and oxidative stress. In addition, this study also focused on the evaluation of Alpha-synuclein (aS) levels; SLNs showed the potential to modulate the Rotenone-mediated in-crease in aS levels. In conclusion, our study investigated the potential of SLNs as a delivery system for addressing PD, also representing a promising approach for enhanced delivery of pharmaceutical and antioxidant molecules across the BBB