Chimeric (allo-auto or even xeno-auto) cultured keratinocyte grafting did not exhibit obvious acute rejection or chronic rejection. Although cultured murine keratinocytes were recognized by allogenic CD8+ T cells, they were not rejected. The precise mechanisms underlying this process were unclear. To analyze how keratinocytes attenuated the immune response, we investigated the effect of culturing on neonatal murine keratinocytes and their immunomodulatory properties.Keratinocytes isolated and purified from BALB/c and C57BL/6 neonatal mice were cultured for 7 days. The expression of B7-1, B7-2, B7-H1 and MHC-I was examined by semi-quantitative RT polymerase chain reaction (PCR), fluorescence microscopy and flow cytometry. Cytotoxicity and mixed lymphocyte response (MLR) assays were performed to determine the effects of keratinocytes on cytotoxic T-lymphocyte (CTL) mediated cell lysis and lymphocyte proliferation.B7-1 was highly expressed in cultured, proliferating murine keratinocytes while no expression of B7-2 and B7-H1 was found. Keratinocytes that expressed B7-1 decreased CTL-mediated cell lysis by an interaction between B7-1 and CTLA-4. In addition, autologous keratinocytes but not allogeneic keratinocytes significantly suppressed auto-specific lymphocyte proliferation in a dose-dependent manner. The modulation was dependent on B7-1 expression and its interaction with CTLA-4.Cultured murine keratinocytes expressed B7-1, but not B7-2 or B7-H1. The keratinocytes attenuated CTL-mediated lysis and suppressed lymphocyte proliferation via an interaction with B7-1 and CTLA-4. Therefore, separate expression of B7-1 induced immunosuppression. Non-professional APCs (antigen presenting cells) which separately express B7-1 may possess an ability to induce immunotolerance and thus act as a regulatory APC.