The effect of mucolytic agents on gene transfer across a CF sputum barrier in vitro. Academic Article uri icon

Overview

MeSH

  • Adenoviridae
  • Cell Line
  • Genetic Vectors
  • Humans
  • Liposomes

MeSH Major

  • Cystic Fibrosis
  • Deoxyribonucleases
  • Gene Transfer Techniques
  • Genetic Therapy
  • Sputum

abstract

  • Trials of gene transfer for cystic fibrosis (CF) are currently underway. However, direct application to the airways may be impeded by the presence of airway secretions. We have therefore assessed the effect of CF sputum on the expression of the reporter gene beta-galactosidase complexed with the cationic liposome DC-Chol/DOPE in a number of cell lines in vitro. Transfection was markedly inhibited in the presence of sputum; the effect was concentration dependent and was only partially ameliorated by removal of sputum with phosphate-buffered saline (PBS) washing before gene transfer. However, treatment of the sputum-covered cells with recombinant human DNase (rhDNase, 50 micrograms/ml) but not with N-acetylcysteine, Nacystelyn, lysine (all 20 mM) or recombinant alginase (0.5 U/ml) significantly (P < 0.005) improved gene transfer. Adenovirus-mediated gene transfer efficiency in the presence of sputum was similarly inhibited, and again, treatment with rhDNase before transfection significantly improved gene transfer (P < 0.005). Transfection of Cos 7 cells in the presence of exogenous genomic DNA alone demonstrated similar inhibition to that observed with sputum and was also ameliorated by pre-treatment of DNA-covered cells with rhDNase. In a separate series of experiments performed in the absence of added sputum or genomic DNA, increasing concentrations of rhDNase resulted in a concentration-related decline in transfection efficiency. However, even at the highest concentration (500 micrograms/ml of rhDNase), transfection efficiency remained more than 50% of control. Thus, pre-treatment of CF airways with rhDNase may be appropriate before liposome or adenovirus-mediated gene therapy.

publication date

  • January 1998

has subject area

  • Adenoviridae
  • Cell Line
  • Cystic Fibrosis
  • Deoxyribonucleases
  • Gene Transfer Techniques
  • Genetic Therapy
  • Genetic Vectors
  • Humans
  • Liposomes
  • Sputum

Research

keywords

  • Journal Article

Identity

Language

  • eng

Digital Object Identifier (DOI)

  • 10.1038/sj.gt.3300556

PubMed ID

  • 9536269

Additional Document Info

start page

  • 91

end page

  • 98

volume

  • 5

number

  • 1