Delivery of antisense oligodeoxyribonucleotide lipopolyplex nanoparticles assembled by microfluidic hydrodynamic focusing.
A multi-inlet microfluidic hydrodynamic focusing (MF) system to prepare lipopolyplex (LP) containing Bcl-2 antisense deoxyoligonucleotide (ODN) was developed and evaluated. The lipopolyplex nanoparticles consist of ODN:protamine:lipids (1:0.3:12.5wt/wt ratio) and the lipids included DC-Chol:egg PC:PEG-DSPE (40:58:2mol/mol%). Using K562 human erythroleukemia cells, which contain an abundance of Bcl-2 and overexpression of transferrin receptors (TfR), and G3139 (oblimerson sodium or Genasense(TM)) as a model cell line and drug, respectively, the Bcl-2 down-regulation at the mRNA and protein levels as well as cellular uptake and apoptosis was compared between the conventional bulk mixing (BM) method and the MF method. The lipopolyplex size and surface charge were characterized by dynamic light scattering (DLS) and zeta potential (zeta) measurement, respectively, while the ODN encapsulation efficiency was determined by gel electrophoresis. Cryogenic transmission electron microscopy (Cryo-TEM) was used to determine the morphology of LPs. Our results demonstrated that MF produced LP nanoparticles had similar structures but smaller size and size distribution compared to BM LP nanoparticles. MF LP nanoparticles had higher level of Bcl-2 antisense uptake and showed more efficient down-regulation of Bcl-2 protein level than BM LP nanoparticles.
Koh
Chee Guan
CG
Department of Chemical and Biomolecular Engineering, The Ohio State University, Columbus, OH 43210, USA.
Zhang
Xulang
X
Liu
Shujun
S
Golan
Sharon
S
Yu
Bo
B
Yang
Xiaojuan
X
Guan
Jingjiao
J
Jin
Yan
Y
Talmon
Yeshayahu
Y
Muthusamy
Natarajan
N
Chan
Kenneth K
KK
Byrd
John C
JC
Lee
Robert J
RJ
Marcucci
Guido
G
Lee
L James
LJ
D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D020285:Cryoelectron Microscopy; D015536:Down-Regulation; D004337:Drug Carriers / Q000737:chemistry*; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D020014:K562 Cells; D008081:Liposomes; D044085:Microfluidics / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry*; D020319:Oligodeoxyribonucleotides, Antisense / Q000008:administration & dosage* / Q000737:chemistry / Q000235:genetics; D010316:Particle Size; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D011990:Receptors, Transferrin / Q000096:biosynthesis; D013499:Surface Properties
D004337:Drug Carriers; D008081:Liposomes; D020319:Oligodeoxyribonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D011990:Receptors, Transferrin; D016650:Fluorescein-5-isothiocyanate