Ultrasound-targeted microbubble destruction (UTMD) assisted delivery of shRNA against PHD2 into H9C2 cells.
Gene therapy has great potential for human diseases. Development of efficient delivery systems is critical to its clinical translation. Recent studies have shown that microbubbles in combination with ultrasound (US) can be used to facilitate gene delivery. An aim of this study is to investigate whether the combination of US-targeted microbubble destruction (UTMD) and polyethylenimine (PEI) (UTMD/PEI) can mediate even greater gene transfection efficiency than UTMD alone and to optimize ultrasonic irradiation parameters. Another aim of this study is to investigate the biological effects of PHD2-shRNA after its transfection into H9C2 cells. pEGFP-N1 or eukaryotic shPHD2-EGFP plasmid was mixed with albumin-coated microbubbles and PEI to form complexes for transfection. After these were added into H9C2 cells, the cells were exposed to US with various sets of parameters. The cells were then harvested and analyzed for gene expression. UTMD/PEI was shown to be highly efficient in gene transfection. An US intensity of 1.5 W/cm2, a microbubble concentration of 300μl/ml, an exposure time of 45s, and a plasmid concentration of 15μg/ml were found to be optimal for transfection. UTMD/PEI-mediated PHD2-shRNA transfection in H9C2 cells significantly down regulated the expression of PHD2 and increased expression of HIF-1α and downstream angiogenesis factors VEGF, TGF-β and bFGF. UTMD/PEI, combined with albumin-coated microbubbles, warrants further investigation for therapeutic gene delivery.
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Li
L
Zhang
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Zhenxing
Z
Sun
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Pingping
P
Ren
Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, Ohio, 43210, United States of America.
Robert J
RJ
Lee
School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, PR China.
Guangya
G
Xiang
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Qing
Q
Lv
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Wei
W
Han
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Jing
J
Wang
The Heart Center, St. Christopher's Hospital for Children/Drexel University College of Medicine, Philadelphia, Pennsylvania, United States of America.
Shuping
S
Ge
Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, 430022, PR China.
Mingxing
M
Xie
D002460:Cell Line; D015870:Gene Expression / Q000235:genetics; D018014:Gene Transfer Techniques; D015316:Genetic Therapy; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics*; D045423:Microbubbles; D010957:Plasmids / Q000235:genetics; D011094:Polyethyleneimine / Q000008:administration & dosage / Q000737:chemistry; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000235:genetics; D014465:Ultrasonics
D034741:RNA, Small Interfering; D016212:Transforming Growth Factor beta; D049452:Green Fluorescent Proteins; D011094:Polyethyleneimine; C490190:EGLN1 protein, human; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases