title: A fusogenic segment of glycoprotein H from herpes simplex virus enhances transfection efficiency of cationic liposomes.
abstract: BACKGROUND
  Low cellular uptake and endosomal escape are regarded as major limitations for nonviral gene delivery. In the present study, we use cationic liposomes incorporating fusogenic peptides from glycoprotein H of herpes simplex virus to improve cellular internalization and endosomal release of cationic liposome-DNA complexes.
  
  
  METHODS
  A synthetic analogue of a fusogenic peptide domain in glycoprotein H from herpes simplex virus was evaluated for in vitro gene delivery. The fusogenicity of the peptide was evaluated by a lipid mixing assay in neutral and acidic environments. The influence of the peptide on cellular internalization and subcellular distribution of Lipofectamine-pGL3 complexes were evaluated by flow cytometry, fluorescence microscopy, and confocal laser scanning microscopy. The effect of the peptide on transfection efficiency of cationic liposomes was also evaluated by luciferase assay in human cell lines.
  
  
  RESULTS
  A pH-sensitive fusogenicity of the wild-type peptide was observed. In the lipid mixing assay, the peptide mediated 80% membrane fusion at pH 7.4 at a 0.05 : 1 (peptide : lipid) mole ratio. At pH 4.5, 100% of membrane fusion was observed at a 0.005 : 1 ratio. Adding the peptide to Lipofectamine-pGL3 complexes significantly increased cellular uptake and, thus, increased transgene expression up to 30-fold in human cell lines. Nuclear localization of the DNA complex by the fusogenic peptide was observed by confocal microscopy.
  
  
  CONCLUSIONS
  A synthetic analogue of a fusogenic peptide domain from herpes simplex virus has been proven to improve cationic lipid-mediated transfection in vitro. The application of this fusogenic peptide will lead to improved strategies for transfection and successful gene therapy.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN 38163, USA.
authors.forename: Ying
authors.initials: Y
authors.lastname: Tu
authors.forename: Ji-seon
authors.initials: JS
authors.lastname: Kim
mesh_terms: D002460:Cell Line; D005434:Flow Cytometry; D015316:Genetic Therapy / Q000379:methods*; D006028:Glycosphingolipids; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008055:Lipids; D008081:Liposomes; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D017434:Protein Structure, Tertiary / Q000235:genetics; D014162:Transfection / Q000379:methods*; D014759:Viral Envelope Proteins / Q000235:genetics*
chemical_list: D006028:Glycosphingolipids; D008055:Lipids; C086724:Lipofectamine; D008081:Liposomes; C097562:PGL3a glycosphingolipid; D014759:Viral Envelope Proteins; C044200:glycoprotein H, herpes simplex virus type 1