| Non-viral cutaneous DNA vaccine delivery using electroporation, liposome formulation (DOTAP:Cholesterol), and sonoporation was investigated for improving transfection and immune responses in vivo in rabbits. The effect of electroporation pulse parameters, liposome formulation, and sonoporation on cutaneous gene delivery was studied using a reporter plasmid coding for beta-galactosidase. Pulse voltage was found to have a more profound effect than the pulse length and pulse number. The expression of beta-galactosidase was transient with maximum levels at 48 h. The expression of beta-galactosidase with 300 V pulses was 129-fold higher compared to passive injection at 48 h. Cationic liposome formulation alone and in combination with sonoporation significantly enhanced (p < 0.05) cutaneous gene delivery compared to passive injection. However, the efficiency of electroporation mediated gene delivery was better than the other methods investigated.; The effect of electroporation, liposome formulation, and sonoporation on cutaneous DNA vaccine delivery was investigated using a DNA vaccine against Hepatitis B virus (HBV). Electroporation pulses of 300 V induced significantly higher (p < 0.05) humoral and cellular immune responses compared to passive injection and liposome formulation alone or in combination with sonoporation. Increasing the dose of DNA vaccine resulted in slightly enhanced immune responses with passive delivery, but not with electroporation. Electroporation mediated DNA vaccine delivery produced a Th1 type immune response. Liposome formulation alone and in combination with sonoporation significantly enhanced (p < 0.05) antibody responses compared to passive, but not cellular immune responses.; The safety of electroporation was studied by investigating the changes in skin barrier function, irritation, viability, and microscopic structure. The electroporation pulses induced skin barrier perturbation and irritation as indicated by elevated transepidermal water loss and erythema/edema, respectively. Microscopic studies revealed inflammatory responses in the epidermis following electroporation. However, these changes were reversible within a week. Skin viability was not affected by the electroporation protocols tested. In conclusion, electroporation mediated non-viral cutaneous gene transfer enhanced the transfection and induced strong humoral/cellular immune responses without initiating any irreversible changes in the skin. |
