Stable gene expression by self-complementary adeno-associated viruses in human MSCs.

Area of Science:

• Biotechnology
• Gene Therapy
• Stem Cell Biology

Background:

• Mesenchymal stem cells (MSCs) are crucial for treating human diseases.
• Genetic modification of MSCs requires safe and efficient gene transfer vectors.
• Self-complementary adeno-associated viruses (scAAVs) are emerging as potential gene delivery tools.

Purpose of the Study:

• To evaluate the efficacy and safety of scAAVs for genetic modification of human bone marrow and umbilical cord blood MSCs.
• To assess the long-term transgene expression and differentiation potential of genetically modified MSCs.
• To determine the feasibility of using distinct scAAV serotypes for co-expression of multiple transgenes.

Main Methods:

• Human bone marrow and umbilical cord blood MSCs were transduced with various scAAV serotypes.
• Transduction efficiency was quantified using flow cytometry.
• Osteogenic differentiation potential was assessed using standard assays.
• Long-term gene expression was evaluated in vivo in rat brains.

Main Results:

• scAAV2 and scAAV5 demonstrated effective and safe transgene expression in both MSC types.
• High transduction efficiencies (around 67%) were achieved with scAAV2 at a multiplicity of infection of 1000.
• Co-infection with scAAV2 and scAAV5 allowed for the expression of different transgenes in the same MSC.
• Genetically modified MSCs maintained long-term gene expression for three months in vivo and did not show impaired osteogenic differentiation.

Conclusions:

• scAAVs, particularly scAAV2 and scAAV5, represent a promising platform for the genetic modification of human MSCs.
• This approach facilitates safe, long-term transgene expression without compromising MSC differentiation capacity.
• scAAV-mediated gene transfer holds significant potential for developing novel cell-based therapies.