BACKGROUND: Recently, several ligand interactions have been examined in detail as potential mediators of costimulatory signaling. The CD154/CD40 and CD28/B7 interactions have been highlighted as being among the more-significant contributors to proper activation of unprimed T lymphocytes. Human keratinocytes (HK) and human dermal fibroblasts (HF) are capable of expressing Class II HLA and CD40 antigens after interferon-gamma exposure, yet neither express significant levels of B7. HK and HF have been characterized as "nonprofessional" antigen presenting cells (APC) and their poor APC function has been partially attributed to deficient costimulatory activity. METHODS: In this study, we examined whether substituting for costimulatory signaling events through the addition of cross-linked monoclonal antibodies against the T-cell ligand/s (CD28 and/or CD154) could restore allostimulation. Mixed lymphocyte reactions were performed combining enriched human peripheral blood T cells and allogeneic HK or HF with or without stimulatory anti-CD28 and/or anti-CD154 antibodies. RESULTS: The results show that the addition of anti-CD28 alone permitted HF but not HK to present alloantigen effectively. In contrast, addition of both anti-CD154 and anti-CD28 was required to generate even a moderate proliferative response to allogeneic HK. Further, adding a monomorphic anti-HLA-DR antibody substantially inhibited these responses. Additional experiments suggest that signaling through CD40/CD154 directs HK to produce TGF-beta, which would adversely affect T-cell activation. CONCLUSIONS: The data presented highlight significant differences in signaling capacities for HK versus HF and provide evidence for a partial mechanism by which allogeneic human skin equivalents might be immunologically null upon engraftment.
BACKGROUND: Recently, several ligand interactions have been examined in detail as potential mediators of costimulatory signaling. The CD154/CD40 and CD28/B7 interactions have been highlighted as being among the more-significant contributors to proper activation of unprimed T lymphocytes. Human keratinocytes (HK) and human dermal fibroblasts (HF) are capable of expressing Class II HLA and CD40 antigens after interferon-gamma exposure, yet neither express significant levels of B7. HK and HF have been characterized as "nonprofessional" antigen presenting cells (APC) and their poor APC function has been partially attributed to deficient costimulatory activity. METHODS: In this study, we examined whether substituting for costimulatory signaling events through the addition of cross-linked monoclonal antibodies against the T-cell ligand/s (CD28 and/or CD154) could restore allostimulation. Mixed lymphocyte reactions were performed combining enriched human peripheral blood T cells and allogeneic HK or HF with or without stimulatory anti-CD28 and/or anti-CD154 antibodies. RESULTS: The results show that the addition of anti-CD28 alone permitted HF but not HK to present alloantigen effectively. In contrast, addition of both anti-CD154 and anti-CD28 was required to generate even a moderate proliferative response to allogeneic HK. Further, adding a monomorphic anti-HLA-DR antibody substantially inhibited these responses. Additional experiments suggest that signaling through CD40/CD154 directs HK to produce TGF-beta, which would adversely affect T-cell activation. CONCLUSIONS: The data presented highlight significant differences in signaling capacities for HK versus HF and provide evidence for a partial mechanism by which allogeneic human skin equivalents might be immunologically null upon engraftment.
Authors: John M Centanni; Joely A Straseski; April Wicks; Jacquelyn A Hank; Cathy A Rasmussen; Mary A Lokuta; Michael J Schurr; Kevin N Foster; Lee D Faucher; Daniel M Caruso; Allen R Comer; B Lynn Allen-Hoffmann Journal: Ann Surg Date: 2011-04 Impact factor: 12.969