Mixed cell co-cultures of lymphocytes responding to kidney cells (MLKC), islets of Langerhans (MLIC) and mixed lymphocyte culture (MLC), were used to clarify mechanisms in allogeneic and autoimmune (tissue-associated) antigen presentation. Fresh kidney cortical tubular cells (KC), Madin-Darby canine kidney (MDCK), and dog kidney (6247) (DK) cell lines were used in MLKC reactions, and islets of Langerhans were used in the MLIC as putative antigen presenting cells (APC). The stimulating cells were treated with purified or recombinant dog interferon-gamma (IFN-γ), and the detection of class II MHC molecule expression was assessed by a moneclonal antibody (mAb) (B1F6). Transcription of MHC class II mRNA and IFN-γ mRNA was measured by semiquantitative polymerase chain reaction, and detection of a kidney cell (tissue-associated) antigen molecule was assessed by the mAb I1F6 that recognizes 72 and 150 kDa tubular cell protein(s) (KT1). The MDCK cell line constitutively expressed low levels of MHC class II molecules and KT1. The steady-state level of the MHC class II mRNA transcription was virtually unaltered by treatment with IFN-γ (400 units) for 48 hours; however, the MHC cell surface protein expression was enhanced. The KC and DK cell lines constitutively expressed KT1, but not MHC class II molecules; these cells required a minimum of 4000 units, and a 62-hour incubation with IFN-γ was needed to upregulate both surface MHC class II molecules and the transcription of corresponding specific mRNA. In the MLKC reaction both the MDCK and DK cell lines, as well as fresh KC cells, could serve as lymphocyte activators. This could be amplified by exogenous IFN-γ. The removal of APC from the responding T cell population did not reduce the IFN-γ effect. This indicates that IFN-γ treatment allows for the expression of all the co-stimulating factors and/or adhesion molecules necessary for these cells to serve as (surrogate) APC (direct as opposed to indirect antigen presentation). The requirements for purified IFN-γ to increase this amplification was greater in the MLKC reactions with kidney cells than in the MLC reactions. The mAbs anti-IFN-γ and I1F6 differed in their ability to inhibit lymphocyte proliferation depending on the different cell types involved. The I1F6 inhibited the MDCK and DK cell-driven MLKC (in the absence of exogenous IFN-γ). The anti-IFN-γ mAb did not inhibit MDCK- (allogeneic)-driven MLKC, in which it was seen that the translation or posttranslation of class II MHC molecules could be upregulated by even small amounts of (endogenous) IFN-γ, and neither I1F6 nor anti-IFN-γ had any effect on the purely allogeneic MLC reactions. These findings suggest that the paracrine effects of endogenous IFN-γ levels, at critical concentrations and durations, might play a crucial role in T lymphocyte activation by graft cells acting as surrogate APC in presenting tissue-associated nominal antigens in vivo.
ASJC Scopus subject areas
- Immunology and Allergy