title: Heterologous expression of mutated HLA-G1 reduces alloreactivity of human dermal fibroblasts.
abstract: OBJECTIVE
  To engineer a stable HLA-G molecule and evaluate its immunomodulatory properties in transgenic human dermal fibroblasts (HDFs).
  
  
  METHODS
  A mutated HLA-G1 (mHLA-G1) molecule was generated by modifying the endoplasmic reticulum retrieval motif and 3'-untranslated region miRNA-binding sites of HLA-G1. Immunomodulatory properties of transgenic HDF-mHLA-G1 were evaluated in vitro.
  
  
  RESULTS
  Stable mHLA-G1 expressing HDF cells were successfully generated and flow cytometry analysis revealed that mHLA-G1 efficiently localized to the cell surface. Natural killer cell-mediated cytolysis of HDF-mHLA-G1/green fluorescent protein (GFP) was reduced by 73% compared with HDF-GDP. HDF-mHLA-G1/GFP decreased phytohemagglutinin-activated peripheral blood mononuclear cell proliferation by 30% versus HDF-GFP.
  
  
  CONCLUSIONS
  We are the first to successfully create a human fibroblast source with reduced alloreactivity using a novel mHLA-G1 construct. This approach may be extended to other cell types including human embryonic stem cells for use in allogeneic transplantation for cell-based regenerative medicine applications.
authors.affiliation: Escape Therapeutics, Inc., San Jose, CA, USA.
authors.forename: Takele
authors.initials: T
authors.lastname: Teklemariam
authors.forename: Longmei
authors.initials: L
authors.lastname: Zhao
authors.forename: Basil M
authors.initials: BM
authors.lastname: Hantash
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D000078404:Epidermal Cells; D004817:Epidermis / Q000276:immunology* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000276:immunology* / Q000378:metabolism; D059951:HLA-G Antigens / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D051379:Mice; D009154:Mutation / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D059951:HLA-G Antigens; D012333:RNA, Messenger
keywords: HLA-G; PBMC; PiggyBac; alloreactivity; cytotoxicity; immune tolerance; nucleofection