title: Visualization of intracellular trafficking of Math1 protein in different cell types with a newly-constructed nonviral gene delivery plasmid.
abstract: BACKGROUND
  In recent years, Math1 gene therapy was indicated to be the future therapy for deafness in combination with other growth factors. However, Math1 delivery using adenovirus-mediated gene delivery or electroporation was impractical. The contribution of Math1 in the combined procedure was not clearly elucidated using the existing plasmids. Nonviral gene delivery vectors are expected to be extremely safe and convenient. The present study aimed to construct the pCDNA6.2/C-EmGFP-Math1 plasmid and evaluate its transfection efficiency and intracellular trafficking of Math1 protein corresponding to transcription regulation function.
  
  
  METHODS
  After constructing the pCDNA6.2/C-EmGFP-Math1 expression plasmid, the plasmid was transfected into different cell lines and primary cochlear cells using Lipofectamine 2000. Transfection efficiencies of the plasmid were evaluated. Transfection efficiencies using liposome nanoparticles containing Math1 plasmid were also assessed. Intracellular trafficking of Math1 was monitored using confocal microscopy.
  
  
  RESULTS
  Different cell types can be transfected with high transfection efficiencies by the pcDNA6.2/C-EmGFP-Math1 plasmid using Lipofectamine 2000. Liposome nanoparticles containing the Math1 plasmid expressed the gene with variable efficiencies, depending on the particle size, surface charge and PEGylation status. Unique intracellular trafficking of Math1 was demonstrated in different cell types.
  
  
  CONCLUSIONS
  The newly-constructed plasmid pcDNA6.2/C-EmGFP-Math1 was suitable for nonviral gene delivery of Math1. Unique intracellular trafficking of Math1 with dynamics from the cytoplasm to the nucleus was demonstrated. The modification of mesenchymal stem cells by Math1 gene delivery and by brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor treatments can potentially be applied to cell replacement for the treatment of cochlear spiral ganglion cell loss in deafness.
authors.affiliation: Department of Otolaryngology, University of Tampere, Tampere, Finland.
authors.forename: Weikai
authors.initials: W
authors.lastname: Zhang
authors.forename: Ya
authors.initials: Y
authors.lastname: Zhang
authors.forename: Rohit
authors.initials: R
authors.lastname: Sood
authors.forename: Sanjeev
authors.initials: S
authors.lastname: Ranjan
authors.forename: Elena
authors.initials: E
authors.lastname: Surovtseva
authors.forename: Aqeel
authors.initials: A
authors.lastname: Ahmad
authors.forename: Paavo K J
authors.initials: PK
authors.lastname: Kinnunen
authors.forename: Ilmari
authors.initials: I
authors.lastname: Pyykkö
authors.forename: Jing
authors.initials: J
authors.lastname: Zou
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000378:metabolism*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D042541:Intracellular Space / Q000378:metabolism*; D008055:Lipids; D008081:Liposomes / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D053758:Nanoparticles; D010641:Phenotype; D010865:Pilot Projects; D010957:Plasmids / Q000235:genetics; D021381:Protein Transport; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D008055:Lipids; D008081:Liposomes; D011993:Recombinant Fusion Proteins; C086724:Lipofectamine