Optimized Pretreatment Method for Long-Term Transduction Efficiency of AAV Into Rat Bladder

Abstract Objective We sought the optimal preprocessing method to improve the long-term transfer efficiency of adeno-associated viral (AAV) vectors of rat bladder urothelium. Methods Rat bladders were pretreated by eight different chemical reagents including PBS, trypsin, poly-L-lysine (PLL), protamine sulfate (PS), sodium dodecyl sulfate (SDS), n-dodecyl-β-d-maltoside (DDM), HCl or NaOH followed by transurethral instillation with AAV2/9-CMV-zsgreen. The transfection efficiency was evaluated by the percentage of urothelium with green fluorescence after 4 weeks. Results We employed eight different chemical components and eleven pretreatment methods. The rat bladder was harvested and evaluated without any mortality at 4 weeks post-infection. Compared with other treatment groups, the rate of urothelium transfection is comparatively high in the groups of rats treated with HCl or NaOH. In addition, rats treated with 0.1N NaOH 4 min showed significantly highest transfection efficiency (about 3%). The histopathologic results showed regular morphology of the bladder treated with HCl or NAOH and no significant damage was observed. Conclusion Pretreatment with 0.1N NaOH 4 min could improve the transfection efficiency of AAV in bladder urothelium and maintain the transfection efficiency for a long time without pathological damage. Importance We found that pretreatment of the bladder with HCl and NaOH can improve the transfection efficiency of AAV for a long time compared to control group. In addition, rats treated with 0.1N NaOH 4 min could have the most significant effect among these strategies, which was discovered for the first time. This could be valuable for future bladder gene therapy.