scholarly journals Hyaluronidase and Collagenase Increase the Transfection Efficiency of Gene Electrotransfer in Various Murine Tumors

2012 ◽  
Vol 23 (1) ◽  
pp. 128-137 ◽  
Author(s):  
Maja Cemazar ◽  
Muriel Golzio ◽  
Gregor Sersa ◽  
Jean-Michel Escoffre ◽  
Andrej Coer ◽  
...  
Keyword(s):  
Transfection Efficiency ◽  
Gene Electrotransfer ◽  
Murine Tumors

scholarly journals Analysis of DNA mobility and gene electrotransfer in 3D in vitro collagen gel model

2020 ◽  
Author(s):  
Saša Haberl Meglič ◽  
Mojca Pavlin
Keyword(s):  
3D Model ◽  
Transfection Efficiency ◽  
Collagen Gel ◽  
Gene Electrotransfer ◽  
Electric Pulses ◽  
Impaired Mobility ◽  
Efficient Gene ◽  
Dna Mobility

Abstract Background: Gene electrotransfer is an established method that enables transfer of DNA into cells with electric pulses. Several studies analyzed different parameters; however the question of the mechanisms involved in gene electrotransfer remains open. One of main obstacles toward efficient gene electrotransfer in vivo is relatively poor DNA mobility in tissues.Objective and method: In order to analyze the effect of impaired mobility on gene electrotransfer efficiency, we applied electric pulses with different durations on plated cells, cells grown on collagen layer and cells embedded in collagen gel (3D model) and compared gene electrotransfer efficiency and viability of cells.Results: We obtained the highest transfection of plated cells, while transfection efficiency of embedded cells in 3D model was lowest and similar as in in vivo. To further analyze poor DNA mobility in 3D model, we applied DNA in top of or injected it into 3D model and showed that former way increases gene electrotransfer efficiency as was shown in in vivo studies.Conclusion: We reported empirically and theoretically evidence that DNA has impaired mobility and diffusion in collagen environment. In addition our method provides resembling in vivo situation, where gene electrotransfer mechanisms can be studied.

scholarly journals Investigation of Plasmid DNA Delivery and Cell Viability Dynamics for Optimal Cell Electrotransfection In Vitro

2020 ◽  
Vol 10 (17) ◽  
pp. 6070
Author(s):  
Sonam Chopra ◽  
Paulius Ruzgys ◽  
Martynas Maciulevičius ◽  
Milda Jakutavičiūtė ◽  
Saulius Šatkauskas
Keyword(s):  
Cell Membrane ◽  
Cell Viability ◽  
High Voltage ◽  
Plasmid Dna ◽  
Transfection Efficiency ◽  
Dna Delivery ◽  
Output Parameter ◽  
Gene Electrotransfer ◽  
Electric Pulses

Electroporation is an effective method for delivering plasmid DNA molecules into cells. The efficiency of gene electrotransfer depends on several factors. To achieve high transfection efficiency while maintaining cell viability is a tedious task in electroporation. Here, we present a combined study in which the dynamics of both evaluation types of transfection efficiency and the cell viability were evaluated in dependence of plasmid concentration as well as at the different number of high voltage (HV) electric pulses. The results of this study reveal a quantitative sigmoidal (R2 > 0.95) dependence of the transfection efficiency and cell viability on the distance between the cell membrane and the nearest plasmid. We propose this distance value as a new, more accurate output parameter that could be used in further optimization studies as a predictor and a measure of electrotransfection efficiency.

scholarly journals Different Influences of Lipofection and Electrotransfection on In Vitro Gene Delivery to Primary Cultured Cortex Neurons

2016 ◽  
Vol 3;19 (3;3) ◽  
pp. 189-196
Author(s):  
Lan Xiao
Keyword(s):  
Gene Transfer ◽  
Cultured Cells ◽  
Transfection Efficiency ◽  
Dendritic Arborization ◽  
Therapeutic Modality ◽  
Gene Electrotransfer ◽  
Cns Diseases ◽  
Neuronal Viability ◽  
Optimized Protocol

Background: Many pain states are linked to central nervous system (CNS) diseases involving the dysfunction of dendritic arborization, making restoration a promising therapeutic strategy. Transfection of primary cortex neurons offers the possibility to study mechanisms which are important for the restoration of proper arborization. Its progress is, however, limited at present due to the lack of suitable gene transfer techniques. Objective: To obtain better insight into the transfection potential of currently used techniques, 2 non-viral transfection methods, lipofection and gene electrotransfer (GET), were compared. Study Design: This is a comparison study performed on cultured cells. Methods: The transfection efficiency and neuronal viability, as well as the neuronal dendritic arborization after lipofection or GET, were compared. Primary cultured cortex neurons were transfected with the pEGFP-N1 plasmid, either using Lipofectamine 2000 (2, 3, or 4μL) or with electroporation, with our previously optimized protocol (200V/25 ms). Results: Transfection efficiency and cell viability were inversely proportional for lipofection. The appropriate ratio of Lipofectamine and plasmid DNA provides optimal conditions for lipofection. Although GET offered higher transfection efficiency, it could not induce complex dendritic arborization, which made it unsuitable for in vitro gene transfer into cortex neurons. Limitations: Limitations include species variability and translational applicability for CNS diseases and pain states related to potential toxicity. Conclusions: Based on these findings, lipofection might be advantageous for in vitro application to primary cultured cortex neurons. Pain states, stress mediated pathogenesis, and certain CNS diseases might potentially utilize this important technique in the future as a therapeutic modality. Key words: Lipofection, gene electrotransfer, CNS diseases, pain states, dendritic arborization, transfection of primary cortex neurons

Stepwise Development of Biomimetic Chimeric Peptides for Gene Delivery

2020 ◽  
Vol 27 (8) ◽  
pp. 698-710
Author(s):  
Roya Cheraghi ◽  
Mahboobeh Nazari ◽  
Mohsen Alipour ◽  
Saman Hosseinkhani
Keyword(s):  
Gene Delivery ◽  
Targeted Delivery ◽  
Viral Vectors ◽  
Large Scale ◽  
Transfection Efficiency ◽  
Cationic Lipids ◽  
Target Cells ◽  
Scale Production ◽  
Stepwise Development ◽  
Chimeric Peptides

Gene-based therapy largely relies on the vector type that allows a selective and efficient transfection into the target cells with maximum efficacy and minimal toxicity. Although, genes delivered utilizing modified viruses transfect efficiently and precisely, these vectors can cause severe immunological responses and are potentially carcinogenic. A promising method of overcoming this limitation is the use of non-viral vectors, including cationic lipids, polymers, dendrimers, and peptides, which offer potential routes for compacting DNA for targeted delivery. Although non-viral vectors exhibit reduced transfection efficiency compared to their viral counterpart, their superior biocompatibility, non-immunogenicity and potential for large-scale production make them increasingly attractive for modern therapy. There has been a great deal of interest in the development of biomimetic chimeric peptides. Biomimetic chimeric peptides contain different motifs for gene translocation into the nucleus of the desired cells. They have motifs for gene targeting into the desired cell, condense DNA into nanosize particles, translocate the gene into the nucleus and enhance the release of the particle into the cytoplasm. These carriers were developed in recent years. This review highlights the stepwise development of the biomimetic chimeric peptides currently being used in gene delivery.

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