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J.–L. Bourges, F. Behar–Cohen; Tranfection Of Dendritic Cells By Electroporation . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1285.
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To assess the potential of electroporation to transfect a GFP plasmid in dendritic cells and observe the kinetics of these transfected cells during the acute phase of the corneal allograft rejection.
Material and Methods: :
The Plasmid GFP was tranfected by electroporation into cornea of Brown–Norway (BN) rats. These corneas were used later as corneal buttons (3mm) for transplant into Lewis rat recipients. The early clinical stages of acute graft rejection are observed by day 5 (D5) and the end stages of rejection were observed by D13 after penetrating keratoplasty (PKP). Animals were sacrificed on (D3), (D6) and (D14) after transplantation and the eyes processed for the preparation of flat–mount of the corneas including the recipient bed and the transplanted graft. Draining regional lymph nodes were also collected and analysed. Transfected but not grafted corneas of BN rats and fellow corneas of the grafted lewis rats were used as controls.
At day 3 after PKP, the GFP positive dendritic cell density dramatically increased in the graft and across the sutures. On the GFP transfected BN corneas, some GFP positive dendritic cells were identified in the mid periphery and across the limbus area also on D6. These cells were also observed in the ciliary body. No GFP positive dendritic cells were detected in the host cornea. At D14, the dendritic cell density markedly decreased in the graft area whether some positive cells were present in the recipient cornea. Lymph nodes did not express GFP at any time point.
Corneal dendritic cells can efficiently be tranfected by electroporation technique. This approach could be applied for the potential modulation of the local immune response after corneal graft transplantation.
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