title: Ultrasound-mediated vascular gene transfection by cavitation of endothelial-targeted cationic microbubbles.
abstract: OBJECTIVE
  Ultrasound-mediated gene delivery can be amplified by acoustic disruption of microbubble carriers that undergo cavitation. We hypothesized that endothelial targeting of microbubbles bearing cDNA is feasible and, through optimizing proximity to the vessel wall, increases the efficacy of gene transfection.
  
  
  BACKGROUND
  Contrast ultrasound-mediated gene delivery is a promising approach for site-specific gene therapy, although there are concerns with the reproducibility of this technique and the safety when using high-power ultrasound.
  
  
  METHODS
  Cationic lipid-shelled decafluorobutane microbubbles bearing a targeting moiety were prepared and compared with nontargeted microbubbles. Microbubble targeting efficiency to endothelial adhesion molecules (P-selectin or intercellular adhesion molecule [ICAM]-1) was tested using in vitro flow chamber studies, intravital microscopy of tumor necrosis factor-alpha (TNF-α)-stimulated murine cremaster muscle, and targeted contrast ultrasound imaging of P-selectin in a model of murine limb ischemia. Ultrasound-mediated transfection of luciferase reporter plasmid charge coupled to microbubbles in the post-ischemic hindlimb muscle was assessed by in vivo optical imaging.
  
  
  RESULTS
  Charge coupling of cDNA to the microbubble surface was not influenced by the presence of targeting ligand, and did not alter the cavitation properties of cationic microbubbles. In flow chamber studies, surface conjugation of cDNA did not affect attachment of targeted microbubbles at microvascular shear stresses (0.6 and 1.5 dyne/cm(2)). Attachment in vivo was also not affected by cDNA according to intravital microscopy observations of venular adhesion of ICAM-1-targeted microbubbles and by ultrasound molecular imaging of P-selectin-targeted microbubbles in the post-ischemic hindlimb in mice. Transfection at the site of high acoustic pressures (1.0 and 1.8 MPa) was similar for control and P-selectin-targeted microbubbles but was associated with vascular rupture and hemorrhage. At 0.6 MPa, there were no adverse bioeffects, and transfection was 5-fold greater with P-selectin-targeted microbubbles.
  
  
  CONCLUSIONS
  We conclude that ultrasound-mediated transfection at safe acoustic pressures can be markedly augmented by endothelial juxtaposition.
authors.affiliation: Division of Cardiovascular Medicine, Oregon Health & Science University, Portland, Oregon 97239, USA.
authors.forename: Aris
authors.initials: A
authors.lastname: Xie
authors.forename: Todd
authors.initials: T
authors.lastname: Belcik
authors.forename: Yue
authors.initials: Y
authors.lastname: Qi
authors.forename: Terry K
authors.initials: TK
authors.lastname: Morgan
authors.forename: Shivam A
authors.initials: SA
authors.lastname: Champaneri
authors.forename: Sarah
authors.initials: S
authors.lastname: Taylor
authors.forename: Brian P
authors.initials: BP
authors.lastname: Davidson
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhao
authors.forename: Alexander L
authors.initials: AL
authors.lastname: Klibanov
authors.forename: Michael A
authors.initials: MA
authors.lastname: Kuliszewski
authors.forename: Howard
authors.initials: H
authors.lastname: Leong-Poi
authors.forename: Azzdine
authors.initials: A
authors.lastname: Ammi
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Lindner
mesh_terms: D000818:Animals; D003287:Contrast Media; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000000981:diagnostic imaging* / Q000378:metabolism*; D015316:Genetic Therapy / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D045423:Microbubbles; D015203:Reproducibility of Results; D014162:Transfection / Q000379:methods*; D014465:Ultrasonics; D014463:Ultrasonography
chemical_list: D003287:Contrast Media