title: The synergy between structural stability and DNA-binding controls the antibody production in EPC/DOTAP/DOPE liposomes and DOTAP/DOPE lipoplexes.
abstract: We present a comparative study of the physico-chemical properties, in vitro cytotoxicity and in vivo antibody production of surface-complexed DNA in EPC/DOTAP/DOPE (50/25/25% molar) liposomes and DOTAP/DOPE (50/50% molar) lipoplexes. The study aims to correlate the biological behavior and structural properties of the lipid carriers. We used DNA-hsp65, whose naked action as a gene vaccine against tuberculosis has already been demonstrated. Additionally, surface-complexed DNA-hsp65 in EPC/DOTAP/DOPE (50/25/25% molar) liposomes was effective as a single-dose tuberculosis vaccine. The results obtained showed that the EPC inclusion stabilized the DOTAP/DOPE structure, producing higher melting temperature and lower zeta potential despite a close mean hydrodynamic diameter. Resemblances in morphologies were identified in both structures, although a higher fraction of loaded DNA was not electrostatically bound in EPC/DOTAP/DOPE. EPC also induced a striking reduction in cytotoxicity, similar to naked DNA-hsp65. The proper immune response lead to a polarized antibody production of the IgG2a isotype, even for the cytotoxic DOTAP/DOPE. However, the antibody production was detected at 15 and 30 days for DOTAP/DOPE and EPC/DOTAP/DOPE, respectively. Therefore, the in vivo antibody production neither correlates with the in vitro cytotoxicity, nor with the structural stability alone. The synergistic effect of the structural stability and DNA electrostatic binding upon the surface of structures account for the immunological effects. By adjusting the composition to generate proper packing and cationic lipid/DNA interaction, we allow for the optimization of liposome formulations for required immunization or gene therapy. In a specific manner, our results contribute to studies on the tuberculosis therapy and vaccination.
authors.affiliation: Departamento de Processos Biotecnológicos, Faculdade de Engenharia Química, Universidade Estadual de Campinas, UNICAMP, 13083-970 Campinas, SP, Brazil.
authors.forename: Lucimara Gaziola
authors.initials: LG
authors.lastname: de la Torre
authors.forename: Rogério Silva
authors.initials: RS
authors.lastname: Rosada
authors.forename: Ana Paula Fávaro
authors.initials: AP
authors.lastname: Trombone
authors.forename: Fabiani Gai
authors.initials: FG
authors.lastname: Frantz
authors.forename: Arlete A M
authors.initials: AA
authors.lastname: Coelho-Castelo
authors.forename: Celio Lopes
authors.initials: CL
authors.lastname: Silva
authors.forename: Maria Helena Andrade
authors.initials: MH
authors.lastname: Santana
mesh_terms: D000818:Animals; D000917:Antibody Formation; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology; D002412:Cations; D016923:Cell Death; D018834:Chaperonin 60 / Q000235:genetics / Q000276:immunology; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D015342:DNA Probes / Q000737:chemistry; D004337:Drug Carriers; D024202:Electrophoretic Mobility Shift Assay; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry*; D005453:Fluorescence; D008081:Liposomes / Q000737:chemistry*; D051379:Mice; D046529:Microscopy, Electron, Transmission; D010316:Particle Size; D044367:Phase Transition; D010713:Phosphatidylcholines / Q000737:chemistry*; D010714:Phosphatidylethanolamines / Q000737:chemistry*; D010957:Plasmids / Q000235:genetics; D000644:Quaternary Ammonium Compounds / Q000737:chemistry*; D044366:Transition Temperature; D032581:Tuberculosis Vaccines / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D019444:Vaccines, DNA / Q000737:chemistry / Q000235:genetics / Q000276:immunology
chemical_list: C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D001426:Bacterial Proteins; D002412:Cations; D018834:Chaperonin 60; D015342:DNA Probes; D004337:Drug Carriers; D005229:Fatty Acids, Monounsaturated; D008081:Liposomes; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D032581:Tuberculosis Vaccines; D019444:Vaccines, DNA; C074119:heat-shock protein 65, Mycobacterium; D004247:DNA; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane