We established cytotoxic T-lymphocyte (CTL) lines against colonic epithelial cell line from LPLs of patients with ulcerative colitis by continuous stimulation with human lymphocyte antigen A (HLA-A) matched colonic epithelial cell lines and clones from LPLs using polyclonal stimulation with phytohemagglutinin. The established CTL lines and clones showed cytotoxicity against HLA-A-matched colonic epithelial cell line but not against HLA-mismatched colonic epithelial cell lines, and HLA-A-matched lung and esophagus cell lines. The CTL response was HLA class I-restricted and mediated by CD8-positive T-lymphocytes. Moreover, the CTL line showed cytotoxicity against autologous B-LCLs pulsed with peptides extracted from HLA-A-matched colonic epithelial cell line but not against other organ-derived peptides pulsed and unpulsed autologous B-LCLs. CTL lines and clones established from LPLs of patients with ulcerative colitis showed colon-specific and HLA class I-restricted killing of human colonic epithelial cell line, suggesting that these CTLs may play a role in colonic epithelial cell damage in some patients with ulcerative colitis.