Nonviral Transfection of Suspension Cells in Ultrasound Standing Wave Fields

Lee, Yu Hsiang; Peng, Ching An

doi:10.1016/j.ultrasmedbio.2006.10.015

Cited by 14 publications

(16 citation statements)

References 38 publications

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“…An aqueous stock solution of PEI was prepared by diluting 1 mg of the commercial solution in 1000 mL DI water, neutralized with HCl and filtered at 0.2 μm (Millipore, USA). The PEI/DNA complexes were performed by mixing PEI and plasmid DNA at 6:1 of N/P ratio (PEI nitrogen: DNA phosphate ratio) in the DI water [7] . The complexes were incubated for 20-30 min at room temperature and stored in 4°C.…”

Section: Preparation Of Pei/dna Complexesmentioning

confidence: 99%

“…Recent studies have shown that ultrasound-targeted microbubble destruction (UTMD) is a new, non-viral, promising method for the gene delivery to the heart [1][2][3][4][5] . The combination of ultrasound (US) irradiation with polyethylenimine (PEI) can not only improve the transfection efficiency markedly [6,7] , but also achieve specific targeting [8] . Researches showed that UTMD combined with PEI could enhance the transfecZhiyi CHEN, male, born in 1975, Doctorial Candidate E-mai: winchen@smail.hust.edu.cn # Corresponding author * The project was supported by a grant from the National Natural Sciences Foundation of China (No.…”

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confidence: 99%

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Efficient gene delivery to myocardium with ultrasound targeted microbubble destruction and polyethylenimine

Chen

Xie

Wang

et al. 2008

J. Huazhong Univ. Sci. Technol. [Med. Sci.]

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The aim of present study was to evaluate the feasibility and efficiency of enhanced green fluorescent protein (EGFP) gene delivery to myocardium in vivo by ultrasound targeted microbubble destruction (UTMD) and polyethylenimine (PEI). SonoVue/DNA and PEI/DNA/SonoVue complexes were prepared. Gel electrophoresis analysis was performed to determine the structural integrity of plasmid DNA or PEI/DNA after UTMD. Solutions of plasmid DNA, SonoVue/DNA, PEI/DNA complexes or PEI/DNA/SonoVue complexes were respectively transduced into BALB/c mice hearts by means of transthoracic ultrasound irradiation. Mice undergoing PBS injection, plasmid injection or PEI/DNA complexes injection without ultrasound irradiation served as controls. Gene expression in myocardium was detected 4 days after treatment. Cryosections and histological examinations were conducted. Electrophoresis gel assay showed no damage to DNA or PEI/DNA complexes after UTMD. When the heart was not exposed to ultrasound, the expression of EGFP was observed in the subendocardial myocardium obviously. The strongest expression was detected in the anterior wall of the left ventricle when the heart was exposed to ultrasound alone. Injection of PEI/DNA complexes and UTMD resulted in the highest transfection efficiency and the distributional difference of EGFP was not obvious. No tissue damage was seen histologically. In conclusion, a combination of UTMD and PEI was highly effective in transfecting mice hearts without causing any apparently adverse effect. It provides an alternative to current clinical gene therapy and opens a new concept of non-viral gene delivery for the treatment of cardiac disease.

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Section: Preparation Of Pei/dna Complexesmentioning

confidence: 99%

mentioning

confidence: 99%

Efficient gene delivery to myocardium with ultrasound targeted microbubble destruction and polyethylenimine

Chen

Xie

Wang

et al. 2008

J. Huazhong Univ. Sci. Technol. [Med. Sci.]

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“…Effect of NLS peptide and PEI on DNA integrity using a deoxyribonuclease (DNase) protection assay. To determine whether the PEI/NLS complexes protect the pEGFP-N 3 plasmid from DNase degradation reference earlier works (24) in the present study, the pEGFP-N 3 plasmid was incubated with the PEI at N:P ratios of 6:1 (19) with or without 120 µg NLS (24) for 1 h at room temperature. DNase endonuclease (0.25 U; Thermo Fisher Scientific, Inc.) was then added to pEGFP-N3 plasmid, PEI/DNA and PEI/DNA/NLS solutions for 5, 10 or 15 min at 37˚C.…”

Section: Methodsmentioning

confidence: 99%

“…A number of studies have demonstrated that UTMD significantly increases the permeability of a cell membrane, thereby improving the efficiency of gene transfection (15)(16)(17). In addition, the combination of ultrasound (US) irradiation with PEI markedly improves transfection efficiency (18,19) and specific targeting (20). Yoo and Jeong (21) demonstrated the plasmid DNA/PEI complexes containing NLS attached to psoralen, a nucleic acid-intercalating agent, increased transfection efficiencies in COS-1 cells, indicating that this complex may be used as a potential DNA carrier for therapeutic applications.…”

Section: Introductionmentioning

confidence: 99%

Efficient gene therapy with a combination of ultrasound-targeted microbubble destruction and PEI/DNA/NLS complexes

Wang

Zhou

Cao

et al. 2017

Molecular Medicine Reports

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Current strategies of gene transfection are not efficient at achieving a notable therapeutic effect. The aim of the present study was to combine ultrasound‑targeted microbubble destruction (UTMD) with a polyethylenimine/pEGFP‑N3 plasmid/nuclear localization sequence (PEI/DNA/NLS) complex gene delivery system, and evaluate the transfection efficiency of enhanced green fluorescent protein (EGFP) gene delivery to 293T cells using this system. The formation of PEI/DNA/NLS complexes and the protective effects of PEI/NLS were verified by gel electrophoresis. Solutions consisting of the plasmid alone, PEI/DNA complexes, PEI/DNA/NLS complexes, UTMD+DNA, UTMD+PEI/DNA complexes, and UTMD+PEI/DNA/NLS complexes were transduced into 293T cells via ultrasound irradiation. The expression of GFP was observed using an inverted microscope and transfection efficiency was detected by flow cytometry following 24 h incubation in vitro. Cell activity was detected using a Cell Counting kit (CCK)‑8 assay. Gel electrophoresis confirmed the formation of PEI/DNA/NLS complexes and demonstrated that PEI/NLS exhibited protective effects on plasmid integrity for a limited time. Inverted microscope observations revealed that a greater GFP signal was observed with the combined action of PEI/DNA/NLS complexes with UTMD, and flow cytometry analysis demonstrated the highest level of transfection efficiency in this group. In addition, the viability of the cells detected by CCK‑8 and treated with PEI/DNA/NLS complexes with UTMD was>80%. In conclusion, the combination of UTMD and PEI/DNA/NLS complexes was highly effective for the efficient transfection of 293T cells without causing excessive cell damage. This method may provide a novel and effective gene transduction system to be applied in clinical treatments.

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“…Some of these studies use a standing wave field, specifically with planar resonators. Although the mechanisms are unclear, there is evidence to suggest that these systems have the potential to 15 produce high transfection rates and improve viability over that reported using inertial cavitation [75][76][77] . Whilst the potential for poration in planar resonators is promising 78 there still remains little evidence of the cell physiology, cell dynamics, acoustic environment and poration mechanism in such systems.…”

Section: Cell-interaction Studies and Sonoporationmentioning

confidence: 99%

Acoustofluidics 9: Modelling and applications of planar resonant devices for acoustic particle manipulation

2012

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5This article introduces the design, construction and applications of planar resonant devices for particle and cell manipulation. These systems rely on the pistonic action of a piezoelectric layer to generate a one dimensional axial variation in acoustic pressure through a system of acoustically tuned layers. The resulting acoustic standing wave is dominated by planar variations in pressure causing particles to migrate to planar pressure nodes (or antinodes depending on particle and fluid properties). The consequences of 10 lateral variations in the fields are discussed, and rules for designing resonators with high energy density within the appropriate layer for a given drive voltage presented.  IntroductionThere is a need to manipulate micron-scale particles and cells in many areas of physics, analytical chemistry and the biosciences. Techniques such as filtration, centrifugation and sedimentation are well-established in macro-scale applications, but in microfluidic 15 systems the use of other approaches including optical, magnetic, dielectrophoretic and acoustic forces are of interest. Acoustic radiation forces, typically at ultrasonic frequencies in the hundreds of kHz to tens of MHz region, have wavelengths that are well matched to microfluidic channel scales, yet are capable of generating potential wells with significantly larger length scales. The technology is also relatively straightforward to integrate within microfluidic systems. Acoustic radiation forces 20Acoustic radiation forces can be generated on particles by both travelling and standing acoustic fields. Those in standing wave fields (of primary interest in the applications considered here) are generated by the nonlinear interaction between the acoustic field scattered by the particle and the standing wave field itself. The time averaged radiation force () F r on a small (in comparison with a wavelength) spherical particle of volume V located at r within a stationary acoustic field was shown by Gor'kov 1 to relate to the gradients of the time averaged kinetic and potential energy densities ( kin E and pot E respectively) within the field:The kinetic energy density gradient (a function of the acoustic velocity field within the standing wave) is weighted by a function of the densities ( on them that tends to move them to the acoustic pressure node, and the acoustic velocity antinode. In a planar resonator these are colocated. Generating the required acoustic fieldMany approaches to generating the required acoustic wave field have been reported in the literature. These include the use of near-field effects 2 or focussed ultrasound 3, 4 to trap particles and cells, the excitation of lateral standing waves in which the predominant energy 35 gradients run parallel to the face of the excitation transducer 5,6 , and the generation of cylindrical resonances to focus 7 or arrange

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Nonviral Transfection of Suspension Cells in Ultrasound Standing Wave Fields

Cited by 14 publications

References 38 publications

Efficient gene delivery to myocardium with ultrasound targeted microbubble destruction and polyethylenimine

Efficient gene delivery to myocardium with ultrasound targeted microbubble destruction and polyethylenimine

Efficient gene therapy with a combination of ultrasound-targeted microbubble destruction and PEI/DNA/NLS complexes

Acoustofluidics 9: Modelling and applications of planar resonant devices for acoustic particle manipulation

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