Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters

Yaw Chong Tong, Shwu Fen Chang, Chia Yang Liu, Winston W Y Kao, Chong Heng Huang, Jiahorng Liaw

Research output: Contribution to journalArticle

46 Citations (Scopus)

Abstract

Background: This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide) -poly(propylene oxide) -poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Methods: Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for β-galactosidase (β-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-β-D-galactoside (X-Gal) color staining, 1,2-dioxetane β-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. Results: The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, β-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Conclusions: Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may involve endocytosis and particle size dependent paracellular transport.

Original languageEnglish
Pages (from-to)956-966
Number of pages11
JournalJournal of Gene Medicine
Volume9
Issue number11
DOIs
Publication statusPublished - Nov 2007

Fingerprint

Ophthalmic Solutions
Micelles
Cornea
Genes
Keratin-12
Plasmids
Rabbits
Lac Operon
Edetic Acid
Galactosidases
Gene Expression
Galactosides
Atomic Force Microscopy
Deoxyribonuclease I
Endocytosis
Cytomegalovirus
Transgenes
Particle Size
Aspartic Acid
Glycine

Keywords

  • Eye drops
  • Particle size
  • Polymeric micelles
  • Specific gene expression

ASJC Scopus subject areas

  • Genetics

Cite this

Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters. / Tong, Yaw Chong; Chang, Shwu Fen; Liu, Chia Yang; Kao, Winston W Y; Huang, Chong Heng; Liaw, Jiahorng.

In: Journal of Gene Medicine, Vol. 9, No. 11, 11.2007, p. 956-966.

Research output: Contribution to journalArticle

Tong, Yaw Chong ; Chang, Shwu Fen ; Liu, Chia Yang ; Kao, Winston W Y ; Huang, Chong Heng ; Liaw, Jiahorng. / Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters. In: Journal of Gene Medicine. 2007 ; Vol. 9, No. 11. pp. 956-966.
@article{ccf87afd745f4cb5a52289456de72f7e,
title = "Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters",
abstract = "Background: This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide) -poly(propylene oxide) -poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Methods: Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for β-galactosidase (β-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-β-D-galactoside (X-Gal) color staining, 1,2-dioxetane β-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. Results: The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, β-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Conclusions: Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may involve endocytosis and particle size dependent paracellular transport.",
keywords = "Eye drops, Particle size, Polymeric micelles, Specific gene expression",
author = "Tong, {Yaw Chong} and Chang, {Shwu Fen} and Liu, {Chia Yang} and Kao, {Winston W Y} and Huang, {Chong Heng} and Jiahorng Liaw",
year = "2007",
month = "11",
doi = "10.1002/jgm.1093",
language = "English",
volume = "9",
pages = "956--966",
journal = "Journal of Gene Medicine",
issn = "1099-498X",
publisher = "John Wiley and Sons Ltd",
number = "11",

}

TY - JOUR

T1 - Eye drop delivery of nano-polymeric micelle formulated genes with cornea-specific promoters

AU - Tong, Yaw Chong

AU - Chang, Shwu Fen

AU - Liu, Chia Yang

AU - Kao, Winston W Y

AU - Huang, Chong Heng

AU - Liaw, Jiahorng

PY - 2007/11

Y1 - 2007/11

N2 - Background: This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide) -poly(propylene oxide) -poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Methods: Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for β-galactosidase (β-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-β-D-galactoside (X-Gal) color staining, 1,2-dioxetane β-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. Results: The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, β-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Conclusions: Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may involve endocytosis and particle size dependent paracellular transport.

AB - Background: This study evaluates the eye drop delivery of genes with cornea-specific promoters, i.e., keratin 12 (K12) and keratocan (Kera3.2) promoters, by non-ionic poly(ethylene oxide) -poly(propylene oxide) -poly(ethylene oxide) (PEO-PPO-PEO) polymeric micelles (PM) to mouse and rabbit eyes, and investigates the underlying mechanisms. Methods: Three PM-formulated plasmids (pCMV-Lac Z, pK12-Lac Z and pKera3.2-Lac Z) containing the Lac Z gene for β-galactosidase (β-Gal) whose expression was driven by the promoter of either the cytomegalovirus early gene, the keratin 12 gene or the keratocan gene, were characterized by critical micelle concentration (CMC), dynamic light scattering (DLS), and atomic force microscopy (AFM). Transgene expression in ocular tissue after gene delivery was analyzed by 5-bromo-4-chloro-3-indolyl-β-D-galactoside (X-Gal) color staining, 1,2-dioxetane β-Gal enzymatic activity measurement, and real-time polymerase chain reaction (PCR) analysis. The delivery mechanisms of plasmid-PM on mouse and rabbit corneas were evaluated by EDTA and RGD (arginine-glycine-aspartic acid) peptide. Results: The sizes of the three plasmid-PM complexes were around 150-200 nm with unimodal distribution. Enhanced stability was found for three plasmid-PM formulations after DNase I treatment. After six doses of eye drop delivery of pK12-Lac Z-PM three times a day, β-Gal activity was significantly increased in both mouse and rabbit corneas. Stroma-specific Lac Z expression was only found in pKera3.2-Lac Z-PM-treated animals with pretreatment by 5 mM EDTA, an opener of junctions. Lac Z gene expression in both pK12-Lac Z-PM and pKera3.2-Lac Z-PM delivery groups was decreased by RGD peptide pretreatment. Conclusions: Cornea epithelium- and stroma-specific gene expression could be achieved using cornea-specific promoters of keratin 12 and keratocan genes, and the gene was delivered with PM formulation through non-invasive, eye drop in mice and rabbits. The transfection mechanism of plasmid-PM may involve endocytosis and particle size dependent paracellular transport.

KW - Eye drops

KW - Particle size

KW - Polymeric micelles

KW - Specific gene expression

UR - http://www.scopus.com/inward/record.url?scp=36148962376&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=36148962376&partnerID=8YFLogxK

U2 - 10.1002/jgm.1093

DO - 10.1002/jgm.1093

M3 - Article

VL - 9

SP - 956

EP - 966

JO - Journal of Gene Medicine

JF - Journal of Gene Medicine

SN - 1099-498X

IS - 11

ER -