title: A new strategy for intracellular delivery of enzyme using mesoporous silica nanoparticles: superoxide dismutase.
abstract: We developed mesoporous silica nanoparticle (MSN) as a multifunctional vehicle for enzyme delivery. Enhanced transmembrane delivery of a superoxide dismutase (SOD) enzyme embedded in MSN was demonstrated. Conjugation of the cell-penetrating peptide derived from the human immunodeficiency virus 1 (HIV) transactivator protein (TAT) to mesoporous silica nanoparticle is shown to be an effective way to enhance transmembrane delivery of nanoparticles for intracellular and molecular therapy. Cu,Zn-superoxide dismutase (SOD) is a key antioxidant enzyme that detoxifies intracellular reactive oxygen species, ROS, thereby protecting cells from oxidative damage. In this study, we fused a human Cu,Zn-SOD gene with TAT in a bacterial expression vector to produce a genetic in-frame His-tagged TAT-SOD fusion protein. The His-tagged TAT-SOD fusion protein was expressed in  E. coli  using IPTG induction and purified using FMSN-Ni-NTA. The purified TAT-SOD was conjugated to FITC-MSN forming FMSN-TAT-SOD. The effectiveness of FMSN-TAT-SOD as an agent against ROS was investigated, which included the level of ROS and apoptosis after free radicals induction and functional recovery after ROS damage. Confocal microscopy on live unfixed cells and flow cytometry analysis showed characteristic nonendosomal distribution of FMSN-TAT-SOD. Results suggested that FMSN-TAT-SOD may provide a strategy for the therapeutic delivery of antioxidant enzymes that protect cells from ROS damage.
authors.affiliation: Department of Chemistry, National Taiwan University, Taipei 106, Taiwan.
authors.forename: Yi-Ping
authors.initials: YP
authors.lastname: Chen
authors.forename: Chien-Tsu
authors.initials: CT
authors.lastname: Chen
authors.forename: Yann
authors.initials: Y
authors.lastname: Hung
authors.forename: Chih-Ming
authors.initials: CM
authors.lastname: Chou
authors.forename: Tsang-Pai
authors.initials: TP
authors.lastname: Liu
authors.forename: Ming-Ren
authors.initials: MR
authors.lastname: Liang
authors.forename: Chao-Tsen
authors.initials: CT
authors.lastname: Chen
authors.forename: Chung-Yuan
authors.initials: CY
authors.lastname: Mou
mesh_terms: D017209:Apoptosis; D002462:Cell Membrane / Q000737:chemistry* / Q000378:metabolism; D016503:Drug Delivery Systems; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D018613:Microscopy, Confocal; D053758:Nanoparticles / Q000737:chemistry*; D010316:Particle Size; D016062:Porosity; D017382:Reactive Oxygen Species / Q000378:metabolism; D012822:Silicon Dioxide / Q000737:chemistry* / Q000378:metabolism; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013499:Surface Properties; D054321:rev Gene Products, Human Immunodeficiency Virus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D017382:Reactive Oxygen Species; D054321:rev Gene Products, Human Immunodeficiency Virus; D012822:Silicon Dioxide; D013482:Superoxide Dismutase