A New Generation Nanotherapeutic: pHEMA-Chitosan Nanocomposites in siRNA Delivery

2020 ◽  
Vol 16 ◽  
Author(s):  
Erdal Eroğlu ◽  
Hüseyin Saygın Portakal ◽  
Ayşenur Pamukçu
Keyword(s):  
Ftir Spectroscopy ◽  
Electrostatic Interactions ◽  
Fluorescent Protein ◽  
Sirna Delivery ◽  
Chitosan Nanoparticles ◽  
Average Diameter ◽  
Reticuloendothelial System ◽  
Nucleic Acid Delivery ◽  
Vis Spectroscopy

Background: Despite great hopes for small interfering RNA (siRNA)-based gene therapies, restrictions, including the presence of nucleases, reticuloendothelial system and undesired electrostatic interactions between nucleic acids and the cell membrane, limit the success of these approaches. In the last few decades, non-viral nucleic acid delivery vectors in nano size with high biocompatibility, low toxicity and proton sponge effect have emerged as magic bullets to overcome these drawbacks. Objective: This study aimed to develop poly(2-hydroxyethyl methacrylate) (pHEMA)-chitosan nanoparticles (PCNp), and to transfect green fluorescent protein (GFP)-silencing siRNA (GsiR) in vitro. Method: First, PCNp displaying core-shell structure was synthesized and thereafter GsiR was encapsulated into the core of PCNp. The synthesized PCNp with/without GsiR were characterized using ultraviolet-visible (UV-vis)-spectroscopy, Fourier-transform infrared (FTIR) spectroscopy, thermal decomposition, atomic force microscopy (AFM), scanning electron microscopy (SEM), zeta potential and dynamic light scattering (DLS) measurements. Encapsulation of siRNA into the pHEMA core coated with chitosan shell was demonstrated using fluorescence and FTIR spectroscopy. Results: The surface charge of PCNSs and PCNSs-GsiR were found to be +39.5 and +40.2, respectively. In DLS analysis, an insignificant shift in the Z-average diameter of PCNp was observed from 109 nm to 133 nm using encapsulation of GsiR. In comparison to other studied nanomaterials and a commercial transfection reagent, our findings suggest a promising GFP-silencing effect of 45%. Conclusion: To our knowledge, we have obtained comparable silencing activity with the other studied equivalents despite using the lowest concentration of siRNA in existing literature.

scholarly journals Covalently tethering siRNA to hydrogels for localized, controlled release and gene silencing

2019 ◽  
Vol 5 (8) ◽  
pp. eaax0801 ◽  
Author(s):  
Minh Khanh Nguyen ◽  
Cong Truc Huynh ◽  
Alex Gilewski ◽  
Samantha E. Wilner ◽  
Keith E. Maier ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Therapeutic Potential ◽  
Sirna Delivery ◽  
Hydrolytic Degradation ◽  
Nucleic Acid Delivery ◽  
Covalent Bonds ◽  
Disulfide Linkages ◽  
Green Fluorescent ◽  
Interfering Rna ◽  
Transfection Agent

Small interfering RNA (siRNA) has found many applications in tissue regeneration and disease therapeutics. Effective and localized siRNA delivery remains challenging, reducing its therapeutic potential. Here, we report a strategy to control and prolong siRNA release by directly tethering transfection-capable siRNA to photocrosslinked dextran hydrogels. siRNA release is governed via the hydrolytic degradation of ester and/or disulfide linkages between the siRNA and hydrogels, which is independent of hydrogel degradation rate. The released siRNA is shown to be bioactive by inhibiting protein expression in green fluorescent protein–expressing HeLa cells without the need of a transfection agent. This strategy provides an excellent platform for controlling nucleic acid delivery through covalent bonds with a biomaterial and regulating cellular gene expression, which has promising potential in many biomedical applications.

scholarly journals The Impact of Nylon-3 Copolymer Composition on the Efficiency of siRNA Delivery to Glioblastoma Cells

Nanomaterials ◽  
2019 ◽  
Vol 9 (7) ◽  
pp. 986 ◽  
Author(s):  
Natascha Hartl ◽  
Friederike Adams ◽  
Gabriella Costabile ◽  
Lorenz Isert ◽  
Markus Döblinger ◽  
...  
Keyword(s):  
Particle Size ◽  
Electrostatic Interactions ◽  
Laser Doppler Anemometry ◽  
Sirna Delivery ◽  
Copolymer Composition ◽  
Distribution Analysis ◽  
Glioblastoma Cells ◽  
Promising Therapeutic Approach ◽  
The Impact

Glioblastoma multiforme is a devastating disease that has attracted enormous attention due to poor prognosis and high recurrence. Small interfering RNA (siRNA) in principle offers a promising therapeutic approach by the downregulation of disease-related genes via RNA interference. For efficient siRNA delivery to target sites, cationic polymers are often used in preclinical studies for the protection of siRNA and complex formation based on electrostatic interactions. In an effort to develop biocompatible and efficient nanocarriers with a translational outlook for optimal gene silencing at reduced toxicity, we synthesized two sets of nylon-3 copolymers with variable cationic content (DM or NM monomer) and hydrophobic subunits (CP monomer) and evaluated their suitability for in vitro siRNA delivery into glioblastoma cells. DM0.4/CP0.6 and NM0.4/CP0.6 polymers with similar subunit ratios were synthesized to compare the effect of different cationic subunits. Additionally, we utilized NM0.2/CP0.8 polymers to evaluate the impact of the different hydrophobic content in the polymer chain. The siRNA condensation ability and polymer–siRNA complex stability was evaluated by unmodified and modified SYBR gold assays, respectively. Further physicochemical characteristics, e.g., particle size and surface charge, were evaluated by dynamic light scattering and laser Doppler anemometry, whereas a relatively new method for polyplex size distribution analysis—tunable resistive pulse sensing—was additionally developed and compared to DLS measurements. Transfection efficiencies, the route of cell internalization, and protein knockdown abilities in glioblastoma cells were investigated by flow cytometry. Furthermore, cellular tolerability was evaluated by MTT and LDH assays. All the polymers efficiently condensed siRNA at N/P ratios of three, whereas polymers with NM cationic subunits demonstrated smaller particle size and lower polyplex stability. Furthermore, NM0.2/CP0.8 polyplexes with the highest hydrophobic content displayed significantly higher cellular internalization in comparison to more cationic formulations and successful knockdown capabilities. Detailed investigations of the cellular uptake route demonstrated that these polyplexes mainly follow clathrin-mediated endocytotic uptake mechanisms, implying high interaction capacity with cellular membranes. Taken together with conducive toxicity profiles, highly hydrophobic nylon-3 polymers provide an appropriate siRNA delivery agent for the potential treatment of glioblastoma.

scholarly journals Characterization of Plasmid DNA Location within Chitosan/PLGA/pDNA Nanoparticle Complexes Designed for Gene Delivery

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Hali Bordelon ◽  
Alexandru S. Biris ◽  
Cristina M. Sabliov ◽  
W. Todd Monroe
Keyword(s):  
Nucleic Acids ◽  
Plasmid Dna ◽  
Genetic Manipulation ◽  
Chitosan Nanoparticles ◽  
Complex Surface ◽  
Surface Enhanced Raman Spectroscopy ◽  
Nucleic Acid Delivery ◽  
Label Free

Poly(D,L-lactide-co-glycolide-) (PLGA-)chitosan nanoparticles are becoming an increasingly common choice for the delivery of nucleic acids to cells for various genetic manipulation techniques. These particles are biocompatible, with tunable size and surface properties, possessing an overall positive charge that promotes complex formation with negatively charged nucleic acids. This study examines properties of the PLGA-chitosan nanoparticle/plasmid DNA complex after formation. Specifically, the study aims to determine the optimal ratio of plasmid DNA:nanoparticles for nucleic acid delivery purposes and to elucidate the location of the pDNA within these complexes. Such characterization will be necessary for the adoption of these formulations in a clinical setting. The ability of PLGA-chitosan nanoparticles to form complexes with pDNA was evaluated by using the fluorescent intercalating due OliGreen to label free plasmid DNA. By monitoring the fluorescence at different plasmid: nanoparticle ratios, the ideal plasmid:nanoparticle ration for complete complexation of plasmid was determined to be 1:50. Surface-Enhanced Raman Spectroscopy and gel digest studies suggested that even at these optimal complexation ratios, a portion of the plasmid DNA was located on the outer complex surface. This knowledge will facilitate future investigations into the functionality of the systemin vitroandin vivo.

scholarly journals Tyrosine-Modification of Polypropylenimine (PPI) and Polyethylenimine (PEI) Strongly Improves Efficacy of siRNA-Mediated Gene Knockdown

Nanomaterials ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1809 ◽  
Author(s):  
Sandra Noske ◽  
Michael Karimov ◽  
Achim Aigner ◽  
Alexander Ewe
Keyword(s):  
Sirna Delivery ◽  
Nucleic Acid Delivery ◽  
Cationic Polymers ◽  
Small Interfering Rnas ◽  
Polymeric Systems ◽  
Rna Molecules ◽  
First Time ◽  
Tyrosine Modification

The delivery of small interfering RNAs (siRNA) is an efficient method for gene silencing through the induction of RNA interference (RNAi). It critically relies, however, on efficient vehicles for siRNA formulation, for transfection in vitro as well as for their potential use in vivo. While polyethylenimines (PEIs) are among the most studied cationic polymers for nucleic acid delivery including small RNA molecules, polypropylenimines (PPIs) have been explored to a lesser extent. Previous studies have shown the benefit of the modification of small PEIs by tyrosine grafting which are featured in this paper. Additionally, we have now extended this approach towards PPIs, presenting tyrosine-modified PPIs (named PPI-Y) for the first time. In this study, we describe the marked improvement of PPI upon its tyrosine modification, leading to enhanced siRNA complexation, complex stability, siRNA delivery, knockdown efficacy and biocompatibility. Results of PPI-Y/siRNA complexes are also compared with data based on tyrosine-modified linear or branched PEIs (LPxY or PxY). Taken together, this establishes tyrosine-modified PPIs or PEIs as particularly promising polymeric systems for siRNA formulation and delivery.

scholarly journals Combining Inulin Multifunctional Polycation and Magnetic Nanoparticles: Redox-Responsive siRNA-Loaded Systems for Magnetofection

Polymers ◽  
2019 ◽  
Vol 11 (5) ◽  
pp. 889 ◽  
Author(s):  
Carla Sardo ◽  
Emanuela Fabiola Craparo ◽  
Barbara Porsio ◽  
Gaetano Giammona ◽  
Gennara Cavallaro
Keyword(s):  
Magnetic Field ◽  
Nucleic Acid ◽  
Sirna Delivery ◽  
Superparamagnetic Iron Oxide ◽  
Cell Uptake ◽  
Nucleic Acid Delivery ◽  
Crosslinking Process ◽  
Wide Range ◽  
Theranostic Applications

Superparamagnetic Iron Oxide Nanoparticles (SPIONs) are recognized as one of the most promising agents for theranostic applications. Among methods designed for siRNA delivery, magnetofection, that is, nucleic acid cell uptake under the influence of a magnetic field acting on magnetic nucleic acid vectors, is emerging as a unique approach to combining advantages such as strong improvement of the kinetics of the delivery process and the possibility of localizing nucleic acid delivery to an area where the magnetic field is applied. This paper reports on the preparation of siRNA loaded magnetoplexes—named ICD@SS@SPIONs/siRNA—by controlled crosslinking, in the presence of SPIONs, of the polycation INU-C-DETA, synthesized starting from the polysaccharide inulin by grafting diethylenetriamine and cystamine molecules. The obtained ICD@SS@SPIONs/siRNA have suitable chemical-physical characteristics to be employed for iv administration and are also able to release siRNA in a redox-triggered manner thanks to intracellular glutathione (GSH) mediated reduction of disulphide bridges formed during the crosslinking process. Moreover, ICD@SS@SPIONs/siRNA are able to produce magnetic targeting in vitro on breast cancer cells, without appreciable cyto- and hemo-toxic effects, in a wide range of concentrations. Finally, protein binding to nanoparticles revealed that obtained systems are potentially longer circulating and applicable as a smart multifunctional agents for cancer therapy.

scholarly journals Characteristics, Cryoprotection Evaluation and In Vitro Release of BSA-Loaded Chitosan Nanoparticles

Marine Drugs ◽  
2020 ◽  
Vol 18 (6) ◽  
pp. 315
Author(s):  
Qinying Yan ◽  
Jiaqi Weng ◽  
Xieqi Wu ◽  
Weiwei Wang ◽  
Qingliang Yang ◽  
...  
Keyword(s):  
Electrostatic Interactions ◽  
Release Kinetics ◽  
Chitosan Nanoparticles ◽  
In Vitro Release ◽  
Spectroscopic Techniques ◽  
Release Process ◽  
Dynamic High Pressure Microfluidization ◽  
High Consistency ◽  
And Function

Chitosan nanoparticles (CS-NPs) are under increasing investigation for the delivery of therapeutic proteins, such as vaccines, interferons, and biologics. A large number of studies have been taken on the characteristics of CS-NPs, and very few of these studies have focused on the microstructure of protein-loaded NPs. In this study, we prepared the CS-NPs by an ionic gelation method, and bovine serum albumin (BSA) was used as a model protein. Dynamic high pressure microfluidization (DHPM) was utilized to post-treat the nanoparticles so as to improve the uniformity, repeatability and controllability. The BSA-loaded NPs were then characterized for particle size, Zeta potential, morphology, encapsulation efficiency (EE), loading capacity (LC), and subsequent release kinetics. To improve the long-term stability of NPs, trehalose, glucose, sucrose, and mannitol were selected respectively to investigate the performance as a cryoprotectant. Furthermore, trehalose was used to obtain re-dispersible lyophilized NPs that can significantly reduce the dosage of cryoprotectants. Multiple spectroscopic techniques were used to characterize BSA-loaded NPs, in order to explain the release process of the NPs in vitro. The experimental results indicated that CS and Tripolyphosphate pentasodium (TPP) spontaneously formed the basic skeleton of the NPs through electrostatic interactions. BSA was incorporated in the basic skeleton, adsorbed on the surface of the NPs (some of which were inlaid on the NPs), without any change in structure and function. The release profiles of the NPs showed high consistency with the multispectral results.

scholarly journals Ex Vivo Evaluation of Insulin Nanoparticles Using Chitosan and Arabic Gum

2011 ◽  
Vol 2011 ◽  
pp. 1-6 ◽  
Author(s):  
M. R. Avadi ◽  
A. M. M. Sadeghi ◽  
Naser Mohamadpour Dounighi ◽  
R. Dinarvand ◽  
F. Atyabi ◽  
...  
Keyword(s):  
Size Distribution ◽  
Electrostatic Interactions ◽  
Ex Vivo ◽  
Chitosan Nanoparticles ◽  
Arabic Gum ◽  
Release Studies ◽  
Polymeric Delivery ◽  
In Vitro Insulin Release ◽  
Nanoparticulate Systems

Polymeric delivery systems based on nanoparticles have emerged as a promising approach for peroral insulin delivery. The aim of the present study was to investigate the release of insulin nanoparticulate systems and ex vivo studies. The nanoparticles were prepared by the ion gelation method. Particle size distribution, zeta potential, and polydispersity index of the nanoparticles were determined. It was found that the nanoparticles carried positive charges and showed a size distribution in the range of 170–200 nm. The electrostatic interactions between the positively charged group of chitosan and negatively charged groups of Arabic gum play an important role in the association efficiency of insulin in nanoparticles. In vitro insulin release studies showed an initial burst followed by a slow release of insulin. The mucoadhesion of the nanosystem was evaluated using excised rat jejunum. Ex vivo studies have shown a significant increase in absorption of insulin in the presence of chitosan nanoparticles in comparison with free insulin.

scholarly journals In Vitro Enzymatic Digestibility of Glutaraldehyde-Crosslinked Chitosan Nanoparticles in Lysozyme Solution and Their Applicability in Pulmonary Drug Delivery

Molecules ◽  
2019 ◽  
Vol 24 (7) ◽  
pp. 1271 ◽  
Author(s):  
Nazrul Islam ◽  
Hui Wang ◽  
Faheem Maqbool ◽  
Vito Ferro
Keyword(s):  
Electron Microscopy ◽  
Drug Delivery ◽  
Structural Integrity ◽  
Chitosan Nanoparticles ◽  
Morphological Characterization ◽  
Pulmonary Drug Delivery ◽  
Oil Emulsion ◽  
Vis Spectroscopy ◽  
Before And After

Herein, the degradation of low molecular weight chitosan (CS), with 92% degree of deacetylation (DD), and its nanoparticles (NP) has been investigated in 0.2 mg/mL lysozyme solution at 37 °C. The CS nanoparticles were prepared using glutaraldehyde crosslinking of chitosan in a water-in-oil emulsion system. The morphological characterization of CS particles was carried out using scanning electron microscopy (SEM) and Transmission Electron Microscopy (TEM) techniques. Using attenuated total reflectance Fourier transform infrared (ATR-FTIR) and UV-VIS spectroscopy, the structural integrity of CS and its NPs in lysozyme solution were monitored. The CS powder showed characteristic FTIR bands around 1150 cm−1 associated with the glycosidic bridges (C-O-C bonds) before and after lysozyme treatment for 10 weeks, which indicated no CS degradation. The glutaraldehyde crosslinked CS NPs showed very weak bands associated with the glycosidic bonds in lysozyme solution. Interestingly, the UV-VIS spectroscopic data showed some degradation of CS NPs in lysozyme solution. The results of this study indicate that CS with a high DD and its NPs crosslinked with glutaraldehyde were not degradable in lysozyme solution and thus unsuitable for pulmonary drug delivery. Further studies are warranted to understand the complete degradation of CS and its NPs to ensure their application in pulmonary drug delivery.

Preparation and In Vitro Release Study of Quaternized Chitosan Nanoparticles

2014 ◽  
Vol 1053 ◽  
pp. 466-472
Author(s):  
Yan Wen ◽  
Xiao Yang Zhang ◽  
Li Sheng ◽  
Xi Jun Lian
Keyword(s):  
Electrostatic Interactions ◽  
Fourier Transforms ◽  
Sodium Tripolyphosphate ◽  
Drug Carrier ◽  
Chitosan Nanoparticles ◽  
Buffer Solution ◽  
Solution Ph ◽  
Substitution Degree ◽  
Positively Charged

In order to exploit a novel drug carrier for 5-fluorocrai (5-Fu) with improved entrapment efficiency (EE) and loading capacity (LC) determined by UV/Vis spectrophotometer, N-[(2-hydroxy-3-trimethylammonium) propyl] chitosan chloride (HTCC) nanoparticles were prepared via the ionic gelation based on the electrostatic interactions between positively charged HTCC and negatively charged sodium tripolyphosphate (TPP). The in vitro release behavior of 5-Fu molecules encapsulated within HTCC nanoparticles took a controlled manner in PBS buffer solution (pH 7.4), initiating from a rapid release within 2 hours following a slower sustained release. A higher substitution degree of quaternary ammonium group (DQ) along HTCC chain generated a stronger electrostatic interaction between positively charged HTCC and negatively charged 5-Fu as a result of a higher positively charge density, and contributed to the controlled release of 5-Fu from HTCC nonaparticles. The formation mechanism of HTCC nanoparticles and 5-Fu-loaded HTCC nanoparticles was probed based on the results of fourier transforms infrared spectroscopy (FTIR), transmission electron microscopy (TEM), and size and zeta potential analysis.

scholarly journals Sugar Functionalized Synergistic Dendrimers for Biocompatible Delivery of Nucleic Acid Therapeutics

Polymers ◽  
2018 ◽  
Vol 10 (9) ◽  
pp. 1034 ◽  
Author(s):  
Shuqin Han ◽  
Tsogzolmaa Ganbold ◽  
Qingming Bao ◽  
Takashi Yoshida ◽  
Huricha Baigude
Keyword(s):  
Nucleic Acid ◽  
Sirna Delivery ◽  
Green Fluorescence Protein ◽  
Nucleic Acid Delivery ◽  
Cationic Polymers ◽  
Nucleic Acid Therapeutics ◽  
Fluorescence Protein ◽  
Interfering Rna

Sugars containing cationic polymers are potential carriers for in vitro and in vivo nucleic acid delivery. Monosaccharides such as glucose and galactose have been chemically conjugated to various materials of synergistic poly-lysine dendrimer systems for efficient and biocompatible delivery of short interfering RNA (siRNA). The synergistic dendrimers, which contain lipid conjugated glucose terminalized lysine dendrimers, have significantly lower adverse impact on cells while maintaining efficient cellular entry. Moreover, the synergistic dendrimers complexed to siRNA induced RNA interference (RNAi) in the cells and profoundly knocked down green fluorescence protein (GFP) as well as the endogenously expressing disease related gene Plk1. The new synergic dendrimers may be promising system for biocompatible and efficient siRNA delivery.

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