AJP - Lung Cellular and Molecular Physiology, Vol 269, Issue 1 45-L51, Copyright © 1995 by American Physiological Society
Optimization of cationic lipid-mediated gene transfer to airway epithelia
A. J. Fasbender, J. Zabner and M. J. Welsh
Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City 52242, USA.
The use of cationic lipids for gene transfer to airway epithelia has shown
promise in in vitro and in vivo studies. However, previous studies have
used a wide variety of different lipid preparations and different
formulations. Few studies have been designed to optimize the variables
involved in transfection, and none have been focused on airway epithelia.
Therefore we examined variables that affect cationic lipid-mediated
transfection of HeLa cells and of airway epithelial cells grown on
permeable filter supports at the air-liquid interface. To quantitate
expression of cDNA, we assayed expression of luciferase. We found that the
ratio of DNA to lipid was an important variable that determined
transfection efficiency. In both HeLa and airway epithelial cells, the
optimum charge ratio of cationic lipid to anionic DNA was approximately
1.25, consistent with the notion that a positively charged complex
facilitates interaction with the negatively charged cell membrane. After
testing a series of readily available cationic lipids, we found that
1,2-dimyristyloxypropyl-N,N-dimethyl-hydroxyethyl ammonium bromide
(DMRIE)/dioleoyl phosphatidylethandamine (DOPE) appeared to show good
efficacy. The concentration of DNA and cationic lipid also played an
important role: in HeLa cells the optimum concentration of cationic lipid
was approximately 5 microM and in airway epithelial cells was approximately
15 microM.(ABSTRACT TRUNCATED AT 250 WORDS)
