Dataset Information

Influence of DNA-Microbubble Coupling on Contrast Ultrasound-Mediated Gene Transfection in Muscle and Liver.

ABSTRACT: BACKGROUND:Contrast ultrasound-mediated gene delivery (CUMGD) is a promising approach for enhancing gene therapy that relies on microbubble (MB) cavitation to augment complementary deoxyribonucleic acid (cDNA) transfection. The aims of this study were to determine optimal conditions for charge-coupling cDNA to MBs and to evaluate the advantages of surface loading for gene transfection in muscle and liver. METHODS:Charge coupling of fluorescently labeled cDNA to either neutral MBs (MBN) or cationic MBs (MB+) in low- to high-ionic conditions (0.3%-1.8% NaCl) was assessed by flow cytometry. MB aggregation from cDNA coupling was determined by electrozone sensing. Tissue transfection of luciferase in murine hindlimb skeletal muscle and liver was made by CUMGD with MBN or MB+ combined with subsaturated, saturated, or supersaturated cDNA concentrations (2.5, 50, and 200 ?g/10(8) MBs). RESULTS:Charge-coupling of cDNA was detected for MB+ but not MBN. Coupling occurred over almost the entire range of ionic conditions, with a peak at 1.2% NaCl, although electrostatic interference occurred at >1.5% NaCl. DNA-mediated aggregation of MB+ was observed at ?0.6% NaCl but did not reduce the ability to produce inertial cavitation. Transfection with CUMGD in muscle and liver was low for both MBs at subsaturation concentrations. In muscle, higher cDNA concentrations produced a 10-fold higher degree of transfection with MB+, which was approximately fivefold higher (P < .05) than that for MBN. There was no effect of DNA supersaturation. The same pattern was seen for liver except that supersaturation further increased transfection with MBN equal to that of MB+. CONCLUSIONS:Efficient charge-coupling of cDNA to MB+ but not MBN occurs over a relatively wide range of ionic conditions without aggregation. Transfection with CUMGD is much more efficient with charge-coupling of cDNA to MBs and is not affected by supersaturation except in the liver, which is specialized for macromolecular and cDNA uptake.

SUBMITTER: Xie A

PROVIDER: S-EPMC6454880 | biostudies-literature | 2016 Aug

REPOSITORIES: biostudies-literature

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Influence of DNA-Microbubble Coupling on Contrast Ultrasound-Mediated Gene Transfection in Muscle and Liver.

Xie Aris A Wu Melinda D MD Cigarroa Gabriella G Belcik J Todd JT Ammi Azzdine A Moccetti Federico F Lindner Jonathan R JR

Journal of the American Society of Echocardiography : official publication of the American Society of Echocardiography 20160604 8

<h4>Background</h4>Contrast ultrasound-mediated gene delivery (CUMGD) is a promising approach for enhancing gene therapy that relies on microbubble (MB) cavitation to augment complementary deoxyribonucleic acid (cDNA) transfection. The aims of this study were to determine optimal conditions for charge-coupling cDNA to MBs and to evaluate the advantages of surface loading for gene transfection in muscle and liver.<h4>Methods</h4>Charge coupling of fluorescently labeled cDNA to either neutral MBs ...[more]

PMID: 27267307

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Project description:BackgroundThe present study aimed to determine whether ultrasound-targeted microbubble destruction (UTMD)-mediated angiopoietin 1 (Ang1) gene transfection can improve angiogenesis and potentially reverse left ventricular (LV) structural and sympathetic nerve remodeling in canines with myocardial infarction (MI).MethodsThirty dogs were randomly divided into groups (n=10/group) as follows: the MI group (MI dogs without UTMD treatment), the UTMD group (MI dogs with UTMD-mediated negative control plasmid treatment), and the UTMD-Ang1 group (MI dogs with UTMD-mediated Ang1 plasmid treatment). LV dimensions, systolic function, and synchrony were used to reflect the structural remodeling. The density of tyrosine hydroxylase (TH)- and growth-associated protein 43 (GAP43)-positive nerve fibers were calculated to assess the sympathetic nerve remodeling.ResultsOne month after treatment, the UTMD-Ang1 group showed lower LV end-diastolic dimension (LVEDD: 31.2±2.3 mm) and higher LV ejection fraction (LVEF: 44.6%±4.3%) than the MI group (LVEDD: 34.5±2.2 mm, t=2.282, P=0.014; LVEF: 37.3%±3.1%, t=3.718, P=0.003) and the UTMD group (LVEDD: 34.1±2.8 mm, t=2.264, P=0.040; LVEF: 39.3%±4.5%, t=2.408, P=0.030). LV synchrony was higher in the UTMD-Ang1 group compared with the MI group by 2-dimensional speckle-tracking echocardiography. Angiogenic density was higher in the UTMD group than the MI group but was highest in the UTMD-Ang1 group according to immunohistochemistry of CD31 and α-smooth muscle actin staining. The density of TH- and GAP43-positive nerve fibers were decreased in the UTMD-Ang1 group (TH: 1,928.2±376.6 μm2/mm2; GAP43: 2,090.8±329.2 μm2/mm2) compared with the MI group (TH: 2916.5±558.4 μm2/mm2, t=4.069, P=0.001; GAP43: 3,275.4±548.6 μm2/mm2, t=5.153, P=0.000) and the UTMD group (TH: 2,552.7±408.1 μm2/mm2, t=3.181, P=0.007; GAP43: 2,630.5±419.3 μm2/mm2, t=2.863, P=0.013). The relative Ang1 and sarcoplasmic reticulum Ca2+-ATPase 2a protein levels were significantly higher in the UTMD-Ang1 group than the UTMD and MI groups by Western blot, while the phospholamban levels exhibited the opposite trend. Plasma norepinephrine and N-terminal pro-B-type-natriuretic peptide were significantly reduced in the UTMD-Ang1 group from day 1 to 1 month after MI.ConclusionsUTMD-mediated Ang1 transfection can promote angiogenesis, reverse LV structural and sympathetic nerve remodeling, and improve LV synchrony after MI.

Dataset Information

Influence of DNA-Microbubble Coupling on Contrast Ultrasound-Mediated Gene Transfection in Muscle and Liver.

Publications

Influence of DNA-Microbubble Coupling on Contrast Ultrasound-Mediated Gene Transfection in Muscle and Liver.

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