Electrotransfection and Lipofection Show Comparable Efficiency for In Vitro Gene Delivery of Primary Human Myoblasts

Abstract

Transfection of primary human myoblasts offers the possibility to study mechanisms that are important for muscle regeneration and gene therapy of muscle disease. Cultured human myoblasts were selected here because muscle cells still proliferate at this developmental stage, which might have several advantages in gene therapy. Gene therapy is one of the most sought-after tools in modern medicine. Its progress is, however, limited due to the lack of suitable gene transfer techniques. To obtain better insight into the transfection potential of the presently used techniques, two non-viral transfection methods--lipofection and electroporation--were compared. The parameters that can influence transfection efficiency and cell viability were systematically approached and compared. Cultured myoblasts were transfected with the pEGFP-N1 plasmid either using Lipofectamine 2000 or with electroporation. Various combinations for the preparation of the lipoplexes and the electroporation media, and for the pulsing protocols, were tested and compared. Transfection efficiency and cell viability were inversely proportional for both approaches. The appropriate ratio of Lipofectamine and plasmid DNA provides optimal conditions for lipofection, while for electroporation, RPMI medium and a pulsing protocol using eight pulses of 2 ms at E = 0.8 kV/cm proved to be the optimal combination. The transfection efficiencies for the optimal lipofection and optimal electrotransfection protocols were similar (32 vs. 32.5%, respectively). Both of these methods are effective for transfection of primary human myoblasts; however, electroporation might be advantageous for in vivo application to skeletal muscle.

Multicomponent mannose-containing liposomes efficiently deliver RNA in murine immature dendritic cells and provide productive anti-tumour response in murine melanoma model

“…Our data show that transfection efficiency of Lipofectamine 2000 which approved oneself in NA delivery in different cell lines [54][55][56][57] was low in the case of murine bone-marrow derived DC progenitors and their immature offsets. Nevertheless, the efficiency of Lipofectamine 2000 was demonstrated for RNA delivery into monocyte-derived human CD14 + DCs [58].…”

Global liposome research in the period of 1995–2014: a bibliometric analysis

“…We calculated the proportion of publications that was attributed to high income, upper middle-income, lower middle-income, and low-income countries, as categorized by the World Bank (Mars et al 2015). This categorization is according to 2013 Gross National Income per capita (GPC).…”

Comparative Histological Study on the Effect of Stem Cells, and Gene Modified Stem Cells in Experimentally-Induced Diabetes Type 1 Cardiomyopathy

“…A part of cardiac muscle tissue was homogenized and processed for mRNA extraction followed by using reverse transcriptase (RT) for complementary deoxyribonucleic acid (cDNA) sequencing. Lipofectamine 3000 transfection [15] of cultured and labeled AMSCs by SERC2a [16] was performed.…”