Pulmonary gene silencing in transgenic EGFP mice using aerosolised chitosan/siRNA nanoparticles.
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.
Nielsen
Ebbe J B
EJ
Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus, Denmark.
Nielsen
Jan M
JM
Becker
Daniel
D
Karlas
Alexander
A
Prakash
Hridayesh
H
Glud
Sys Z
SZ
Merrison
Jonathan
J
Besenbacher
Flemming
F
Meyer
Thomas F
TF
Kjems
Jørgen
J
Howard
Kenneth A
KA
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
D000336:Aerosols; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D048271:Chitosan
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