title: Localized Delivery of shRNA against PHD2 Protects the Heart from Acute Myocardial Infarction through Ultrasound-Targeted Cationic Microbubble Destruction.
abstract: Hypoxia-inducible factor 1α (HIF-1α) plays a critical protective role in ischemic heart disease. Under normoxic conditions, HIF-1α was degraded by oxygen-dependent prolyl hydroxylase-2 (PHD2). Gene therapy has become a promising strategy to inhibit the degradation of HIF-1α and to improve cardiac function after ischemic injury. However, conventional gene delivery systems are difficult to achieve a targeted and localized gene delivery into the ischemic myocardia. Here, we report the localized myocardial delivery of shRNA against PHD2 through ultrasound-targeted microbubble destruction (UTMD) for protection the heart from acute myocardial infarction. In this study, a novel cationic microbubble was fabricated by using of the thin-film hydration and sonication method. The resulting microbubbles had a 28.2 ± 2.21 mV surface zeta potential and could greatly improve DNA binding performance, achieving 17.81 ± 1.46 μg of DNA loading capacity per 5 × 108microbubbles. Combined with these cationic microbubbles, UTMD-mediated gene delivery was evaluated and the gene transfection efficiency was optimized in the H9C2 cardiac cells. Knockdown of PHD2 gene was successfully realized by UTMD-mediated shPHD2 transfection, resulting in HIF-1α-dependent protective effects on H9C2 cells through increasing the expression of HIF-1α, VEGF and bFGF. We further employed UTMD-mediated shPHD2 transfection into the localized ischemic myocardia in a rat ischemia model, demonstrating significantly reduced infarct size and greatly improved the heart function. The silencing of PHD2 and the up-regulation of its downstream genes in the treated myocardia were confirmed. Histological analysis further revealed numbers of HIF-1α- and VEGF-, and CD31-positive cells/mm2in the shPHD2-treated group were significantly greater than those in the sham or control vector groups (P < 0.05). In conclusion, our study provides a promising strategy to realize ultrasound-mediated localized myocardial shRNA delivery to protect the heart from acute myocardial infarction via cationic microbubbles.
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhang
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Zhenxing
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Pingping
authors.initials: P
authors.lastname: Ren
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Manjie
authors.initials: M
authors.lastname: You
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Lingyun
authors.initials: L
authors.lastname: Fang
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Yihan
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Paul C. Lauterbur Research Center for Biomedical Imaging, Institute of biomedical and Health Engineering, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Yan
authors.affiliation: Paul C. Lauterbur Research Center for Biomedical Imaging, Institute of biomedical and Health Engineering, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China.
authors.forename: Hairong
authors.initials: H
authors.lastname: Zheng
authors.affiliation: Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Hubei Province Key Laboratory of Molecular Imaging, Wuhan, China.
authors.forename: Mingxing
authors.initials: M
authors.lastname: Xie
mesh_terms: D000818:Animals; D002460:Cell Line; D004195:Disease Models, Animal; D055785:Gene Knockdown Techniques; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000032:analysis; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D045423:Microbubbles; D058990:Molecular Targeted Therapy / Q000379:methods*; D009203:Myocardial Infarction / Q000517:prevention & control*; D011393:Procollagen-Proline Dioxygenase / Q000037:antagonists & inhibitors*; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D051381:Rats; D013010:Sonication / Q000379:methods*; D014162:Transfection / Q000379:methods*; D016896:Treatment Outcome
chemical_list: C497444:Hif1a protein, rat; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D034741:RNA, Small Interfering; D011393:Procollagen-Proline Dioxygenase; C580731:Egln1 protein, rat; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases
keywords: Cationic microbubbles.; Gene delivery; Ischemic myocardial disease; Prolyl hydroxylase-2; Ultrasound