Efficient and sustained transgene expression in human corneal cells mediated by a lentiviral vector

Area of Science:

• Ophthalmology
• Gene Therapy
• Molecular Biology

Background:

• Efficient gene transfer to corneal tissues is crucial for developing novel therapies.
• Existing vectors often fail to transduce all corneal cell types or provide sustained transgene expression.

Purpose of the Study:

• To investigate the efficacy of a recombinant lentiviral vector for gene transfer into human corneal tissue.
• To assess the efficiency and duration of transgene expression in corneal cells using lentiviral vectors.

Main Methods:

• Utilized a lentiviral vector encoding enhanced green fluorescent protein (eGFP).
• Transduced human primary keratocytes in vitro and human corneal tissue ex vivo.
• Assessed transduction efficiency using fluorescent-activated cell sorting (FACS) and microscopy.
• Evaluated transgene expression stability over 60 days in culture.

Main Results:

• Lentiviral vectors efficiently transduced human keratocytes in vitro, with 92% efficiency at a multiplicity of infection (MOI) of 1000.
• Stable eGFP expression was observed in transduced keratocytes for 60 days.
• Efficient in situ transduction of corneal tissue was achieved, with eGFP expression in endothelial, epithelial, and stromal cells.
• Transgene expression in corneal tissue persisted for 60 days in organ culture.

Conclusions:

• Recombinant lentiviral vectors are effective tools for efficient gene transfer into human corneal cells.
• These vectors facilitate temporally stable transgene expression in corneal tissues.
• Lentiviral vector-mediated gene transfer holds potential for clinical applications in ophthalmology.