Chitosan enhances transfection efficiency of cationic polypeptides/DNA complexes

Area of Science:

• Biomaterials Science
• Gene Delivery Systems
• Nanotechnology

Background:

• Developing effective non-viral gene delivery vectors is crucial for gene therapy.
• Cationic polypeptides and chitosan (CS) are explored for their potential in DNA complexation and delivery.
• Human cervical carcinoma (HeLa) cells serve as a model for evaluating transfection efficiency and cytotoxicity.

Purpose of the Study:

• To investigate the impact of mixing cationic polypeptides with chitosan (CS) on in vitro gene transfection efficiency.
• To assess the cytotoxicity of these novel polypeptide/CS/DNA complexes in HeLa cells.
• To optimize formulation parameters including polypeptide type, molecular weight, weight ratios, mixing order, pH, and serum presence.

Main Methods:

• Formulation and characterization of polypeptide/DNA and CS/polypeptide/DNA complexes using gel electrophoresis.
• Evaluation of particle size and surface charge of the complexes.
• In vitro transfection efficiency and cytotoxicity assays (MTT assay) in HeLa cells.

Main Results:

• Poly-L-arginine (PLA) with molecular weight >70 kDa demonstrated the highest transfection efficiency.
• The order of mixing CS, PLA, and DNA significantly influenced transfection outcomes.
• Ternary complexes of PLA/DNA/CS (2:1:4) achieved transfection efficiency comparable to polyethyleneimine (PEI)/DNA complexes.
• Cell viability remained above 80% in MTT assays, indicating low cytotoxicity.

Conclusions:

• Chitosan addition to poly-L-arginine/DNA complexes improves DNA delivery potential in vitro.
• The developed CS-based ternary complexes are easy to prepare and exhibit favorable safety profiles.
• These findings highlight the potential of CS/polypeptide complexes as effective and safe non-viral gene delivery vectors.