Efficient Nonviral Transfection of Human Bone Marrow Mesenchymal Stromal Cells Shown Using Placental Growth Factor Overexpression

الملخص EN

Background.

Human mesenchymal stromal/stem cells (hMSCs) hold great therapeutic potential due to their immunomodulatory and tissue regenerative properties.

Enhancement of biological features of hMSCs by transfection has become a focus of investigation for cell- and gene-based therapies.

However, many of the current transient transfection methods result in either low transfection efficiency or high cytotoxicity.

Methods.

In order to find a transfection method that would address the current issues of low transfection efficiency and high cytotoxicity, 6 commercially available cationic lipid and polymer reagents were tested on human bone marrow-derived MSCs (hBM-MSCs) using GFP as a reporter gene.

One transfection method using TransIT-2020 was selected and tested with an emphasis on cell quality (viability, identity, and yield), as well as efficacy with a human placental growth factor (PlGF) plasmid.

Results.

TransIT-2020 yielded the highest fluorescence signal per cell out of the methods that did not decrease cell recovery.

Transfecting GFP to 5 hBM-MSC donors using TransIT-2020 yielded 24–36% GFP-expressing cells with a viability of 85–96%.

hBM-MSC identity was unaffected as CD90, CD105, and CD73 markers were retained (>95%+) after transfection.

When this method was applied to PlGF expression, there was up to a 220-fold increase in secretion.

Both growth and secretion of PlGF in overexpressing hBM-MSC were sustained over 7 days, confirming the sustainability and applicability of the TransIT-2020 transfection system.

Discussion.

We report a simple and efficient method for transient transfection that has not been reported for hBM-MSCs, encompassing high levels of plasmid expression without significant changes to fundamental hBM-MSC characteristics.