title: Pulmonary gene silencing in transgenic EGFP mice using aerosolised chitosan/siRNA nanoparticles.
abstract: OBJECTIVE
  This work describes the production and application of an aerosolised formulation of chitosan nanoparticles for improved pulmonary siRNA delivery and gene silencing in mice.
  
  
  METHODS
  Aerosolised chitosan/siRNA nanoparticles were pneumatically formed using a nebulising catheter and sized by laser diffraction. In vitro silencing of aerosolised and non-aerosolised formulations was evaluated in an EGFP endogenous-expressing H1299 cell line by flow cytometry. Non-invasive intratracheal insertion of the catheter was used to study nanoparticle deposition by histological detection of Cy3-labeled siRNA and gene silencing in transgenic EGFP mouse lungs using a flow cytometric method.
  
  
  RESULTS
  Flow cytometric analysis demonstrated minimal alteration in gene silencing efficiency before (68%) and after (62%) aerosolisation in EGFP-expressing H1299 cells. Intratracheal catheter administration in mice resulted in nanoparticle deposition throughout the entire lung in both alveoli and bronchiolar regions using low amounts of siRNA. Transgenic EGFP mice dosed with the aerosolised nanoparticle formulation showed significant EGFP gene silencing (68% reduction compared to mismatch group).
  
  
  CONCLUSIONS
  This work provides a technology platform for effective pulmonary delivery and gene silencing of RNAi therapeutics with potential use in preclinical studies of respiratory disease treatment.
authors.affiliation: Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark.
authors.forename: Ebbe J B
authors.initials: EJ
authors.lastname: Nielsen
authors.forename: Jan M
authors.initials: JM
authors.lastname: Nielsen
authors.forename: Daniel
authors.initials: D
authors.lastname: Becker
authors.forename: Alexander
authors.initials: A
authors.lastname: Karlas
authors.forename: Hridayesh
authors.initials: H
authors.lastname: Prakash
authors.forename: Sys Z
authors.initials: SZ
authors.lastname: Glud
authors.forename: Jonathan
authors.initials: J
authors.lastname: Merrison
authors.forename: Flemming
authors.initials: F
authors.lastname: Besenbacher
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Meyer
authors.forename: Jørgen
authors.initials: J
authors.lastname: Kjems
authors.forename: Kenneth A
authors.initials: KA
authors.lastname: Howard
mesh_terms: D000336:Aerosols; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D048271:Chitosan; D005434:Flow Cytometry; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D049452:Green Fluorescent Proteins / Q000235:genetics*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D034622:RNA Interference
chemical_list: D000336:Aerosols; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D048271:Chitosan
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