Induction of HLA-G-specific human CD8+ T cell lines by stimulation across a polymorphism of HLA-G

Abstract

THE NATURE of alloantigens seen by T lymphocytes and the role of peptide ligands in allorecognition are major factors in understanding the mechanisms of allograft acceptance. T cells of the maternal decidua have the capacity to develop immune responses against paternal HLA-G antigens. It has been suggested that paternal HLA-G antigens may elicit an immune response by T cells of the maternal decidua. Evidence for this comes from structural characteristics of HLA-G, polymorphisms in the gene, and thymic selection events restricted by HLA-G. Strong immune responses mediated by alloreactive and HLA-G-specific T cells under special pathologic circumstances may contribute to fetal loss. Our aim was to determine if human peripheral T cells can recognize HLA-G molecules as alloantigens in vitro. To address this question, the development of human cytotoxic T lymphocytes from 20 healthy individuals was examined by stimulation with myelogeneous leukemia K562 cells transfected with the HLA-G*01011 allele. HLA-G genotypes were determined by DNA-sequence analysis of the above individuals by PCR amplification of genomic DNA using HLA-G-specific primers, followed by fluorescent DNA sequencing using dye primer chemistry to increase our ability to detect heterozygotes. No CTL activity was obtained from individuals who were homozygous or heterozygous for the HLA-G*01011, HLA-G*01012, and HLAG*01013 alleles. Strong CTL responses were found in individuals with nucleotide variations in exon 3 that result in amino acid substitutions at positions 110 (Leu to Ile), 130 (Leu to Phe), and 178 (Met to Ile). Cytotoxic activity was blocked when target cells were preincubated with W6/32 mAb. HLA-G-specific CTL lines were established to examine whether alloreactive CTLs would recognize HLA-G on the surface of the antigen-processing defective cell line T2. These CTLs were able to kill T2 HLA-G transfectants with a specific lysis of 25% to 37%, indicating that some alloreactive HLA-G specific T cells are peptide independent. Our results demonstrate that amino acid substitutions in the peptide binding groove of HLA-G molecules can elicit HLA-G-specific T cell response.