Nebulized anionic guanidinylated O-carboxymethyl chitosan/N-2-hydroxypropyltimehyl ammonium chloride chitosan nanoparticles for siRNA pulmonary delivery: preparation, characterization and in vitro evaluation

Abstract

This study developed a pH-sensitive anionic system composed of guanidinylated O-carboxymethyl chitosan (GOCMCS) and N-2-hydroxypropyltimehyl ammonium chloride chitosan (N-2-HACC) for efficient siRNA delivery to the lungs following nebulization. About 16.8% of guanidine groups were incorporated into O-carboxymethyl chitosan (OCMCS) with the aid of O-methylisourea. Gel electrophoresis images demonstrated that siRNA was successfully encapsulated in nanoparticles ranging from 150 to 180 nm with zeta potential of about -17 mV. The nanoparticles containing GOCMCS existed superior transfection performance compared with their amino-based analogs. The evaluation in vitro revealed that nanoparticles were internalized into A549 cells by energy-dependent endocytosis, then achieved endosomal escape by direct transmembrane penetration of guanidine moieties as well as swelling behavior of nanoparticles due to the pH sensitivity of GOCMCS. The mRNA level of survivin gene was down-regulated to 6.9% using GOCMCS/N-2-HACC/siSurvivin NPs. The survivin siRNA mediated by nanoparticles caused 30% of cell growth inhibition and induced 19.45% of cell apoptosis, which was comparable to Lipofectamin2000. After nebulization of siRNA-loaded nanoparticles, the stability of siRNA was maintained and fine particle fractions were detected by two-stage impinger that accounted for more than 60%. These results suggested that GOCMCS/N-2-HACC nanoparticles possessed potential as safe and efficient carrier for siRNA pulmonary delivery.

Efficient siRNA delivery and gene silencing using a lipopolypeptide hybrid vector mediated by a caveolae-mediated and temperature-dependent endocytic pathway

“…As almost all endocytic pathways are energy-dependent processes, they can be inhibited by low temperature incubation or ATP depletion [56, 57]. To analyze the energy-dependent endocytosis pathway, we used 4 °C for the treatment because cells consume less ATP and block the active transport at this temperature.…”

Novel Ellipsoid Chitosan-Phthalate Lecithin Nanoparticles for siRNA Delivery

“…This assumption is supported by reports indicating that amphoteric polymers have better transfection efficiency compared with their cationic analogs because of their enhanced pH buffering properties. It is hypothesized that polymeric buffers consume endosomal protons and, therefore, derive endosomal proton-chloride co-transporter to keep shuttling protons and chloride ions into endosomes leading to increased osmotic pressure, endosomal swelling, and eventually endosomal rupture and escape of siRNA cargo into the cellular cytoplasm (Oskuee et al, 2009 ; Richard et al, 2013 ; Ni et al, 2017 ; Ryu et al, 2018 ).…”

Co‐delivery of hypoxia inducible factor‐1α small interfering RNA and 5‐fluorouracil to overcome drug resistance in gastric cancer SGC‐7901 cells

“…At the beginning of the study, we selected CS nanoparticles as a delivery vector for 5-FU and siRNA, which has been widely used as gene carrier. 35,36 Following this, an evaluation of the properties of the nanoparticles was conducted, including entrapment efficiency, particle size and zeta potential. The binding efficiency of CS nanoparticles and siRNA was evaluated by agarose gel electrophoresis and the results obtained showed that CS nanoparticles can effectively bind siRNA.…”

“…At the beginning of the study, we selected CS nanoparticles as a delivery vector for 5-FU and siRNA, which has been widely used as gene carrier 35,36. (A) The accumulation of Rho-123 in cells.…”