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Antigen‐Specific apoptosis in immortalized T cells by soluble MHC class II‐peptide complexes
Author(s) -
ARIMILLI SUBHASHINI,
MUMM JOHN B,
NAG BISHWAJIT
Publication year - 1996
Publication title -
immunology and cell biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.999
H-Index - 104
eISSN - 1440-1711
pISSN - 0818-9641
DOI - 10.1038/icb.1996.13
Subject(s) - epitope , microbiology and biotechnology , t cell receptor , t cell , biology , major histocompatibility complex , mhc restriction , antigen , cytotoxic t cell , antigen presenting cell , mhc class ii , apoptosis , human leukocyte antigen , mhc class i , immunology , immune system , biochemistry , in vitro
Summary The recognition of T cell receptors (TCR) by purified major histocompatibility complex (MHC) class II‐peptide complexes in the absence of costimulatory signals leads to the induction of T cell nonresponsiveness or anergy. In a recent study using human T cell clones, it was observed that prolonged incubation of resting T cells with soluble MHC II‐peptide complexes appears to result in T cell apoptosis. The present study shows that the engagement of TCR by soluble MHC II‐peptide complexes also results in antigen‐specific apoptosis in immortalized T cells. Apoptosis was demonstrated in a herpes saimiri virus (HSV) transformed human T cell clone (SS8T) restricted for HLA‐DR2 in association with an epitope from the myelin basic protein [MBP(84–102)]. A dose‐ and time‐dependent T cell death was observed upon incubation of SS8T cloned T cells with purified complexes of native human HLA‐DR2 and MBP(83–102)Y 83 peptide. The specificity of T cell apoptosis was demonstrated by exposing SS8T cells with DR2 alone and DR2 bound to another high affinity epitope [MBP(124–143)] from the same MBP. Recently, we have shown that the complexes of HLA‐DR2 and [MBP(83–102)Y 83 ] can be reconstituted by refolding Escherichia coli expressed individual DR2 a and β (B5*010l) polypeptide chains lacking the transmembrane region. When SS8T cloned T cells were exposed to purified reconstituted rDR2.MBP(83–102)Y 83 complexes, similar apoptosis of T cells was observed. Agarose gel analysis of T cells incubated with complexes showed a degradation of celluar deoxyribonucleic acid (DNA) to oligonucleosomal bands, a characteristic of apoptosis. The quantitative detection of DNA strand breaks was performed by pulsing T cells with 5‐bromo‐2′‐deoxyuridine(BrdU) followed by the detection of BrdU‐labelled DNA fragments using an antibody sandwich enzyme‐linked immuno assay (ELISA). The fragmentation of DNA was also measured by double fluorescence flow cytometry by 3′ end labelling of fragmented DNA with biotinylated‐deoxyuridine triphosphate (dUTP) in the presence of terminal deoxynucleotide transferase (TdT) enzyme. The expression of the bel ‐2 protein in SS8T cells following TCR engagement by soluble MHC II‐peptide complexes was monitored by chemiluminescence blot analysis using anti‐ bel ‐2 monoclonal antibody. Finally, the nucleosomal condensation of T cells following complex treatment, characteristics of typical apoptosis, was demonstrated by transmission electron microscopy. These results suggest that the binding of soluble MHC class II‐peptide complexes to TCR induces antigen‐specific apoptosis in transformed CD4 positive T cells in vitro. Such induction of apoptosis by soluble MHC II‐peptide complexes may provide a novel therapeutic strategy to delete autoreactive T cells in various autoimmune diseases.