A combinational treatment for improving the transduction efficiency of lentiviruses in the retinal pigment epithelium cells: Polybrene and Protamine sulfate
Sajad Najafi1 , Javad Ranjbari1 , Azam Rahimpour2 , Hamid Ahmadieh3 , Maryam Maleki Tehrani3 , Fatemeh Suri3 *
- Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
- Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
- Ophthalmic Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Abstract: Viral vectors including lentivirus (LV), adenovirus (AV), and adeno-associated virus (AAV) have been used as common vehicles for gene transfer in gene therapy studies of various human diseases. The efficacy of gene transfer; however, remains unsatisfying; thus, many biological and chemical substances are used to enhance the transduction efficiency (TE). In this article, we aim to evaluate the impact of the combinational use of polybrene (Pb) and protamine sulfate (PS) on the TE of LV particles in the retinal pigment epithelium (RPE) cells.
Methods: LVs were produced using second-generation vectors and used for the transduction of RPE cells extracted from human donors. Different combinations of two enhancers, Pb and PS, were added to the viral mixture. The TE of LVs in human primary RPE cells was evaluated using flow cytometry and calculating the mean fluorescence intensity (MFI) and the percentage of GFP-positive cells.
Results: Pb at all concentrations significantly improved the TE compared to the control virus with the most impact of a 15-fold increase at 10 ?g/ml concentration. The percentage of GFP-positive cells was also most enhanced at that concentration (p-value: 0.006). Also, PS significantly improved the TE in RPE cells compared to control LV. At a combinational concentration of 10 ?g/ml of Pb and 2 ?g/ml of PS, the highest level of TE was detected (MFI: 801, GFP+: 65.4%). However, the value change was insignificant compared to alone enhancers or other combinations.
Conclusion: Pb enhanced the TE of LVs in RPE cells and in combination with PS achieved the highest level of MFI and GFP-positive percentage. This study may suggest a potential combination of enhancers for applications in gene therapy of retinal diseases.
