Background: A mouse model of combined UVA/riboflavin irradiation to eliminate stromal cells and other antigen-presenting cells in the cornea provides the basis for a probably low risk of corneal transplantation. Methods: After abrasion of the epithelium, the central corneas of mouse eyes were treated with UVA/riboflavin in vitro. Histological studies of hematoxylin-eosin and immunohistochemical staining with caspase 3 were performed. Dissected mouse corneas were analyzed by Western blot. Results: Apoptotic cells were shown on the central corneal stroma; a cell-free zone was displayed in the cornea. Numbers of dead cells increased according to cultivation time. However, the endothelium survived due to the adjustment of the irradiation dose. Conclusions: A cell-free zone in the stroma of the mouse cornea was produced by UVA/riboflavin irradiation in vitro. The technique makes possible to prevent or reduce immunological reactions and the risk of graft rejection by pretreatment of the donor cornea, ultimately prolonging graft survival. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Background: A mouse model of combined UVA/riboflavin irradiation to eliminate stromal cells and other antigen-presenting cells in the cornea provides the basis for a probably low risk of corneal transplantation. Methods: After abrasion of the epithelium, the central corneas of mouse eyes were treated with UVA/riboflavin in vitro. Histological studies of hematoxylin-eosin and immunohistochemical staining with caspase 3 were performed. Dissected mouse corneas were analyzed by Western blot. Results: Apoptotic cells were shown on the central corneal stroma; a cell-free zone was displayed in the cornea. Numbers of dead cells increased according to cultivation time. However, the endothelium survived due to the adjustment of the irradiation dose. Conclusions: A cell-free zone in the stroma of the mouse cornea was produced by UVA/riboflavin irradiation in vitro. The technique makes possible to prevent or reduce immunological reactions and the risk of graft rejection by pretreatment of the donor cornea, ultimately prolonging graft survival. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

Background Previously, we identified a major myeloid-derived proinflammatory subpopulation of human blood dendritic cells which we termed slanDCs (e.g. Schäkel et al. (2006) Immunity 24, 767–777). The slan epitope is an O-linked sugar modification (6-sulfo LacNAc, slan) of P-selectin glycoprotein ligand-1 (PSGL-1). As slanDCs can induce neoantigen-specific CD4+ T cells and tumor-reactive CD8+ cytotoxic T cells, they appear as promising targets for an in vivo delivery of antigens for vaccination. However, tools for delivery of antigens to slanDCs were not available until now. Moreover, it is unknown whether or not antigens delivered via the slan epitope can be taken up, properly processed and presented by slanDCs to T cells. Methodology/Principal Findings Single chain fragment variables were prepared from presently available decavalent monoclonal anti-slan IgM antibodies but failed to bind to slanDCs. Therefore, a novel multivalent anti-slanDC scaffold was developed which consists of two components: (i) a single chain bispecific recombinant diabody (scBsDb) that is directed on the one hand to the slan epitope and on the other hand to a novel peptide epitope tag, and (ii) modular (antigen-containing) linker peptides that are flanked at both their termini with at least one peptide epitope tag. Delivery of a Tetanus Toxin-derived antigen to slanDCs via such a scBsDb/antigen scaffold allowed us to recall autologous Tetanus-specific memory T cells. Conclusions/Significance In summary our data show that (i) the slan epitope can be used for delivery of antigens to this class of human-specific DCs, and (ii) antigens bound to the slan epitope can be taken up by slanDCs, processed and presented to T cells. Consequently, our novel modular scaffold system may be useful for the development of human vaccines.

Background Previously, we identified a major myeloid-derived proinflammatory subpopulation of human blood dendritic cells which we termed slanDCs (e.g. Schäkel et al. (2006) Immunity 24, 767–777). The slan epitope is an O-linked sugar modification (6-sulfo LacNAc, slan) of P-selectin glycoprotein ligand-1 (PSGL-1). As slanDCs can induce neoantigen-specific CD4+ T cells and tumor-reactive CD8+ cytotoxic T cells, they appear as promising targets for an in vivo delivery of antigens for vaccination. However, tools for delivery of antigens to slanDCs were not available until now. Moreover, it is unknown whether or not antigens delivered via the slan epitope can be taken up, properly processed and presented by slanDCs to T cells. Methodology/Principal Findings Single chain fragment variables were prepared from presently available decavalent monoclonal anti-slan IgM antibodies but failed to bind to slanDCs. Therefore, a novel multivalent anti-slanDC scaffold was developed which consists of two components: (i) a single chain bispecific recombinant diabody (scBsDb) that is directed on the one hand to the slan epitope and on the other hand to a novel peptide epitope tag, and (ii) modular (antigen-containing) linker peptides that are flanked at both their termini with at least one peptide epitope tag. Delivery of a Tetanus Toxin-derived antigen to slanDCs via such a scBsDb/antigen scaffold allowed us to recall autologous Tetanus-specific memory T cells. Conclusions/Significance In summary our data show that (i) the slan epitope can be used for delivery of antigens to this class of human-specific DCs, and (ii) antigens bound to the slan epitope can be taken up by slanDCs, processed and presented to T cells. Consequently, our novel modular scaffold system may be useful for the development of human vaccines.