Regenerative medicine · 2014
Heterologous expression of mutated HLA-G1 reduces alloreactivity of human dermal fibroblasts.
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Teklemariam Takele, Zhao Longmei, Hantash Basil M
Аннотация
AIM: To engineer a stable HLA-G molecule and evaluate its immunomodulatory properties in transgenic human dermal fibroblasts (HDFs).
MATERIALS & 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.
CONCLUSION: 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.
MATERIALS & 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.
CONCLUSION: 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.