Transfection of Difficult-to-Transfect Rat Primary Cortical Neurons with Magnetic Nanoparticles

Jin, Wu-Yang; Lin, Donghai; Nguyen, Anh H.; Abdelrasoul, Gaser N.; Chen, Jian; Mar, Arthur; Qian, Feng; Fu, Qiang; Kovalchuk, Igor; Wang, Yutian; Chen, Jie

doi:10.1166/jbn.2018.2604

Cited by 13 publications

(12 citation statements)

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“…In this way, our study is unique as we utilize primary cells. There has been a previous study by Jin et al [33] using a magnetic nanoparticle delivery system to transfect rat primary cortical neurons with a CRISPR plasmid, but delivery to these primary cells were found to be at a much lesser degree relative to studies done in immortalized HEK cells. Therefore, delivery in primary cell systems is an needed area of investigation in this field.…”

Section: Discussionmentioning

confidence: 91%

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Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmid

Ringel‐Scaia

McDaniel

et al. 2020

J Nanobiotechnol

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Background: The clustered regularly interspaced short palindromic repeats (CRISPR) and Cas9 protein system is a revolutionary tool for gene therapy. Despite promising reports of the utility of CRISPR-Cas9 for in vivo gene editing, a principal problem in implementing this new process is delivery of high molecular weight DNA into cells. Results: Using poly(lactic-co-glycolic acid) (PLGA), a nanoparticle carrier was designed to deliver a model CRISPR-Cas9 plasmid into primary bone marrow derived macrophages. The engineered PLGA-based carriers were approximately 160 nm and fluorescently labeled by encapsulation of the fluorophore 6,13-bis(triisopropylsilylethynyl) pentacene (TIPS pentacene). An amine-end capped PLGA encapsulated 1.6 wt% DNA, with an encapsulation efficiency of 80%. Release studies revealed that most of the DNA was released within the first 24 h and corresponded to ~ 2-3 plasmid copies released per nanoparticle. In vitro experiments conducted with murine bone marrow derived macrophages demonstrated that after 24 h of treatment with the PLGA-encapsulated CRISPR plasmids, the majority of cells were positive for TIPS pentacene and the protein Cas9 was detectable within the cells. Conclusions: In this work, plasmids for the CRISPR-Cas9 system were encapsulated in nanoparticles comprised of PLGA and were shown to induce expression of bacterial Cas9 in murine bone marrow derived macrophages in vitro. These results suggest that this nanoparticle-based plasmid delivery method can be effective for future in vivo applications of the CRISPR-Cas9 system.

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Section: Discussionmentioning

confidence: 91%

“…Several studies that have successfully employed nanoparticle delivery of CRISPR-Cas9 plasmids have done so using immortalized cell lines [31][32][33]. In this way, our study is unique as we utilize primary cells.…”

Section: Discussionmentioning

confidence: 99%

Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmid

Ringel‐Scaia

McDaniel

et al. 2020

J Nanobiotechnol

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Section: Methodsmentioning

confidence: 99%

“…Furthermore, the complex of the negatively charged plasmid with FN-Glu-PEI25K MNPs mitigates the positive charge of the FN-Glu-PEI25K MNPs. We previously utilized FN-Glu-PEI25K MNPs as gene nano-carriers, and the GFP plasmid/ FN-Glu-PEI25K MNPs showed lower toxicity than the positive control (lipofectamine 2000) [16,41]. The size of nanoparticles was estimated using Hitachi HF-3300 Transmission Electronic Microscope (300 kVTEM), where we measured the size of 150 nanoparticles using ImageJ software, and the size of the nanoparticles was calculated based on the histogram of the size distribution.…”

Section: Plos Onementioning

confidence: 99%

Ultrasound-assisted magnetic nanoparticle-based gene delivery

et al. 2020

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Targeted gene delivery is important in biomedical research and applications. In this paper, we synergistically combine non-viral chemical materials, magnetic nanoparticles (MNPs), and a physical technique, low-intensity pulsed ultrasound (LIPUS), to achieve efficient and targeted gene delivery. The MNPs are iron oxide super-paramagnetic nanoparticles, coated with polyethyleneimine (PEI), which makes a high positive surface charge and is favorable for the binding of genetic materials. Due to the paramagnetic properties of the MNPs, the application of an external magnetic field increases transfection efficiency while LIPUS stimulation enhances cell viability and permeability. We found that stimulation at the intensity of 30 mW/cm 2 for 10 minutes yields optimal results with a minimal adverse effect on the cells. By combining the effect of the external magnetic field and LIPUS, the genetic material (GFP or Cherry Red plasmid) can enter the cells. The flow cytometry results showed that by using just a magnetic field to direct the genetic material, the transfection efficiency on HEK 293 cells that were treated by our MNPs was 56.1%. Coupled with LIPUS stimulation, it increased to 61.5% or 19% higher than the positive control (Lipofectamine 2000). Besides, compared with the positive control, our method showed less toxicity. Cell viability after transfection was 63.61%, which is 19% higher than the standard transfection technique. In conclusion, we designed a new gene-delivery method that is affordable, targeted, shows lowtoxicity, yet high transfection efficiency, compared to other conventional approaches.

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“…Because LIPUS was previously shown to promote cell growth and enhance cell membrane permeability, in this study, we considered combining the LIPUS device application to enhance genetic material delivery into the cells using the PEI coated MNPs [31], [32]. Performing the cell transfection with LIPUS and MNPs under the external magnetic field, we achieved high transfection efficiency with low cytotoxicity.…”

Section: Introductionmentioning

confidence: 99%

Ultrasound-Assisted Magnetic Nanoparticle-Based Gene Delivery

Chen

Zhang

Abdelrasoul

et al. 2020

Preprint

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Low-intensity pulsed ultrasound (LIPUS), a special type of ultrasonic stimulation, is attracting a lot of attention for both clinical and scientific research. In this paper, we report a concept of a new method using magnetic nanoparticles (MNPs) for LIPUSassisted gene delivery. The MNPs are iron oxide superparamagnetic nanoparticles, coated with polyethyleneimine (PEI), which introduces a high positive surface charge, favorable for the binding of genetic material. Due to the paramagnetic properties of the MNPs, the application of an external magnetic field increases transfection efficiency; meanwhile, LIPUS stimulation enhances cell permeability. We found out that stimulation at the intensity of 30 mW/cm 2 for 10 minutes yields optimal results with a minimal adverse effect on the cells. Combining the effect of the external magnetic field and LIPUS, the genetic material (GFP or Cherry Red plasmid in our case) can enter the cells. The flow cytometry results showed that by using just a magnetic field to direct the genetic material, the transfection efficiency of HEK 293 cells that were treated with our MNPs was 56.1%. Coupled with LIPUS stimulation, it increased to 61.5% or 19% higher than the positive control (Lipofectamine 2000). In addition, compared with the positive control, our method showed less toxicity. Cell viability after transfection was 63.61%, 19% higher than with the standard transfection technique. In conclusion, we designed a new gene-delivery technique that is affordable, targeted, shows low-toxicity, yet high transfection efficiency, compared to other conventional approaches.

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Transfection of Difficult-to-Transfect Rat Primary Cortical Neurons with Magnetic Nanoparticles

Cited by 13 publications

References 0 publications

Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmid

Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmid

Ultrasound-assisted magnetic nanoparticle-based gene delivery

Ultrasound-Assisted Magnetic Nanoparticle-Based Gene Delivery

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