Ultrasound-targeted gene delivery induces angiogenesis after a myocardial infarction in mice.
OBJECTIVE
This study evaluated the capacity of ultrasound-targeted microbubble destruction (UTMD) to deliver angiogenic genes, improve perfusion, and recruit progenitor cells after a myocardial infarction (MI) in mice.


BACKGROUND
Angiogenic gene therapy after an MI may become a clinically relevant approach to improve the engraftment of implanted cells if targeted delivery can be accomplished noninvasively. The UTMD technique uses myocardial contrast echocardiography to target plasmid gene delivery to the myocardium and features low toxicity, limited immunogenicity, and the potential for repeated application.


METHODS
Empty plasmids (control group) or those containing genes for vascular endothelial growth factor (VEGF), stem cell factor (SCF), or green fluorescent protein (to visualize gene delivery) were incubated with perflutren lipid microbubbles. The microbubble-deoxyribonucleic acid mixture was injected intravenously into C57BL/6 mice at 7 days after coronary artery ligation (MI). The UTMD technique facilitated transgene release into the myocardium. Twenty-one days after MI, myocardial perfusion and function were assessed by contrast echocardiography. Protein expression was quantified by Western blot and enzyme-linked immunosorbent assay. Flow cytometry quantified progenitor cell recruitment to the heart. Blood vessel density was evaluated immunohistochemically.


RESULTS
Green fluorescent protein expression in the infarcted myocardium demonstrated gene delivery. Myocardial VEGF and SCF levels increased significantly in the respective groups (p < 0.05). The physiologic impact of VEGF and SCF gene delivery was confirmed by increased myocardial recruitment of VEGF receptor 2- and SCF receptor (c-kit)-expressing cells, respectively (p < 0.05). Consequently, capillary and arteriolar density (Factor VIII and alpha-smooth muscle actin staining), myocardial perfusion, and cardiac function were all enhanced (p < 0.01 relative to control group) in recipients of VEGF or SCF.


CONCLUSIONS
Noninvasive UTMD successfully delivered VEGF and SCF genes into the infarcted heart, increased vascular density, and improved myocardial perfusion and ventricular function. The UTMD technique may be an ideal method for noninvasive, repeated gene delivery after an MI.
Fujii
Hiroko
H
Division of Cardiovascular Surgery, Toronto General Research Institute, Toronto General Hospital, Toronto, Ontario, Canada.
Sun
Zhuo
Z
Li
Shu-Hong
SH
Wu
Jun
J
Fazel
Shafie
S
Weisel
Richard D
RD
Rakowski
Harry
H
Lindner
Jonathan
J
Li
Ren-Ke
RK
D000818:Animals; D003287:Contrast Media / Q000008:administration & dosage; D003326:Coronary Circulation; D004195:Disease Models, Animal; D004452:Echocardiography; D005260:Female; D005466:Fluorocarbons / Q000008:administration & dosage; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D007262:Infusions, Intravenous; D051379:Mice; D008810:Mice, Inbred C57BL; D045423:Microbubbles; D009203:Myocardial Infarction / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology / Q000628:therapy*; D009206:Myocardium / Q000378:metabolism* / Q000473:pathology; D018919:Neovascularization, Physiologic; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism; D020127:Recovery of Function; D019089:Stem Cell Factor / Q000096:biosynthesis* / Q000235:genetics; D013234:Stem Cells / Q000378:metabolism; D013997:Time Factors; D014465:Ultrasonics; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000378:metabolism; D016277:Ventricular Function, Left
D003287:Contrast Media; D005466:Fluorocarbons; D019089:Stem Cell Factor; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D049452:Green Fluorescent Proteins; C042852:perflutren; D019009:Proto-Oncogene Proteins c-kit; D040301:Vascular Endothelial Growth Factor Receptor-2