Back signaling of HLA class I molecules and T/NK cell receptor ligands in epithelial cells reflects the rejection-specific microenvironment in renal allograft biopsies.

The role of endothelial cells in the pathophysiology of antibody-mediated rejection after renal transplantation has been widely investigated. We expand this scenario to the impact of epithelial cells on the microenvironment during rejection. Primary proximal tubular epithelial cells were stimulated via HLA class I, CD155 and CD166 based on their potential signal-transducing capacity to mediate back signaling after encounter with either T/NK cells or donor-specific antibodies. Upon crosslinking of these ligands with mAbs, PTEC secreted IL-6, CXCL1,8,10, CCL2, and sICAM-1. These proteins were also released by PTEC as consequence of a direct interaction with T/NK cells. Downmodulation of the receptor CD226 on effector cells confirmed the involvement of this receptor/ligand pair in back signaling. In vivo, CD155 and CD166 expression was detectable in proximal and distal tubuli of renal transplant biopsies, respectively. The composition of the protein microenvironment in these biopsies showed a substantial overlap with the PTEC response. Cluster and principal component analyses of the microenvironment separated unsuspicious from rejection biopsies and, furthermore, ABMR, TCMR, and borderline rejection. In conclusion, our results provide evidence that epithelial cells may contribute to the rejection process and pave the way to a better understanding of the pathomechanisms of kidney allograft rejection.

Down-Regulation of MHC Class I Expression in Human Keratinocytes Using Viral Vectors Containing US11 Gene of Human Cytomegalovirus and Cultivation on Bovine Collagen-Elastin Matrix (Matriderm®): Potential Approach for an Immune-Privileged Skin Substitute.

Skin transplantation, especially in burn patients, is still challenging because surgeons are faced with limited disposability of autologous donor side material. The in vitro culture of keratinocytes has become an important reconstructive option. However, only non-immunogenic allogenic keratinocytes offer the opportunity to develop a skin graft that can overcome rejection. The purpose of the study was to develop targeted gene modification of keratinocytes in order to reduce immunogenicity for the use as allogenic transplantable skin graft by decreasing the expression of MHC class I. To reduce MHC class I expression, viral vectors containing the US11 gene of human cytomegalovirus were generated and tested on their functionality using Western blotting, indirect immunofluorescence staining, and flow cytometry. Transfected keratinocytes were seeded on commercially available bovine collagen-elastin matrices and further cultured for histological and cell survival assays. Results showed transient down-regulation of MHC class I after 24 h post-transfection, with recovery of MHC class I expression after 48 h. Histological assessments showed long-term cell survival as well as histological patterns comparable to epidermal layers of healthy human skin. The data postulates the potential application of US11 transfected keratinocytes as an approach towards an immune-privileged skin substitute. Nevertheless, further studies and data are needed.