炎症性腸疾患における末梢血CD4^+CD25^+T細胞の解析

CD^+でD25^T細胞は調節性T細胞(Treg)と称される.炎症性腸疾患(IBD)モデルマウスヘのTreg移入が腸炎発症を抑制することが報告され,TregがIBDの病因に関与している可能性が示された.そこで我々は病期の異なる同一IBD患者末梢血Treg(CD4^+CD25^+T細胞,CD4^+CD25^T細胞)の割合,Tregに恒常的に発現し抑制機能の鍵とされるGITRおよび細胞内CTLA-4陽性細胞の割合を調べ,ヒトTregがIBDの病態に関与しているか否かを検討した.〔対象および方法〕対象は潰瘍性大腸炎(UC)7人,クローン病(CD)8人,健常人7人とした.末梢血単核球を採取し,CD4,CD25,に対するモノクローナル抗体を用いてフローサイトメトリーにより解析した.CD4^+T細胞中のCD25^+細胞,CD25^細胞の割合(Treg/CD4^+)をCD,UC,健常人において測定した.患者群では同一患者における緩解,活動期にそれぞれ測定を行い,また細胞表面GITR,細胞内CTLA-4も同時に測定し差異を認めるか否かを検討した.〔結果〕CD4^+CD25^+/CD4^+(%)は同一UC患者で,緩解期に比べ活動期で有意に高かった.またCD4^+CD25^/CD4^+(%)は健常人と比較し,活動期CD患者で有意に高かった.一方,活動期CD患者のCD4^+CD25^細胞中CTLA-4陽性細胞の割合(CD4^+CD25^CTLA-4^+/CD4^+CD25^(%))は健常人やCD緩解期に比べて有意に低く,GITR陽性細胞の割合(CD4^+CD25^GITR^+/CD4^+CD25^(%))はCD緩解期に比べてCD活動期で有意に高かった.〔結論〕強い細胞増殖抑制機能を持つCTLA-4はCd活動期CD4^+CD25^で有意に低く,逆にTregの抑制機能を無効とするGITRは有意に高かった.また同様の傾向はUCには認められなかった.同一CD患者の異なる病期でTregの機能に関与するmoleculeの変化を認めた事実はCDの病因にTregが関与している可能性を強く示唆させた.Background and Aim: CD^4+CD25^ T cells act as regulatory T cells. CTLA-4 and GITR are associated with the suppressor function of regulatory T cells. The aim of this study was to assess any differences in the expression patterns of these molecules on regulatory T cells of patients with inflammatory bowel disease (IBD). Methods: The study subjects were 15 patients with IBD (ulcerative colitis=7, Crohn\u27s disease=8) and 7 healthy individuals. Peripheral blood mononuclear cells were obtained by centrifugation of fresh blood over Ficoll-paque and stained with anti-CD4, CD25, and GITR antibodies. For CTLA-4 staining, cells were fixed and permeabilized by saponin prior to incubation with anti-CTLA-4 antibody. Cell samples were analyzed on a FACScaliber. Each patient was investigated both in the active and inactive state. Results: The percentage of CD4^+CD25^+ cells was significantly higher in active than in inactive ulcerative colitis. In contrast, the percentage of CD4^+CD25^ cell was significantly higher in active Crohn\u27s disease than in healthy individuals. CTLA-4^+ cell counts in CD4^+CD25^ T cells were significantly lower in active Crohn\u27s disease than in control and inactive Crohn\u27s disease patients, although no difference was observed between active and inactive ulcerative colitis. In contrast, GITR expression was significantly higher in active than inactive Crohn\u27s disease. Conclusion: Our results suggest that CD4 + CD25high T cells are probably involved in the pathogenesis of Crohn\u27s disease

炎症性腸疾患における末梢血CD4^+CD25^+T細胞の解析

CD^+でD25^<high>T細胞は調節性T細胞(Treg)と称される.炎症性腸疾患(IBD)モデルマウスヘのTreg移入が腸炎発症を抑制することが報告され,TregがIBDの病因に関与している可能性が示された.そこで我々は病期の異なる同一IBD患者末梢血Treg(CD4^+CD25^+T細胞,CD4^+CD25^<high>T細胞)の割合,Tregに恒常的に発現し抑制機能の鍵とされるGITRおよび細胞内CTLA-4陽性細胞の割合を調べ,ヒトTregがIBDの病態に関与しているか否かを検討した.〔対象および方法〕対象は潰瘍性大腸炎(UC)7人,クローン病(CD)8人,健常人7人とした.末梢血単核球を採取し,CD4,CD25,に対するモノクローナル抗体を用いてフローサイトメトリーにより解析した.CD4^+T細胞中のCD25^+細胞,CD25^<high>細胞の割合(Treg/CD4^+)をCD,UC,健常人において測定した.患者群では同一患者における緩解,活動期にそれぞれ測定を行い,また細胞表面GITR,細胞内CTLA-4も同時に測定し差異を認めるか否かを検討した.〔結果〕CD4^+CD25^+/CD4^+(%)は同一UC患者で,緩解期に比べ活動期で有意に高かった.またCD4^+CD25^<high>/CD4^+(%)は健常人と比較し,活動期CD患者で有意に高かった.一方,活動期CD患者のCD4^+CD25^<high>細胞中CTLA-4陽性細胞の割合(CD4^+CD25^<high>CTLA-4^+/CD4^+CD25^<high>(%))は健常人やCD緩解期に比べて有意に低く,GITR陽性細胞の割合(CD4^+CD25^<high>GITR^+/CD4^+CD25^<high>(%))はCD緩解期に比べてCD活動期で有意に高かった.〔結論〕強い細胞増殖抑制機能を持つCTLA-4はCd活動期CD4^+CD25^<high>で有意に低く,逆にTregの抑制機能を無効とするGITRは有意に高かった.また同様の傾向はUCには認められなかった.同一CD患者の異なる病期でTregの機能に関与するmoleculeの変化を認めた事実はCDの病因にTregが関与している可能性を強く示唆させた.Background and Aim: CD^4+CD25^<high> T cells act as regulatory T cells. CTLA-4 and GITR are associated with the suppressor function of regulatory T cells. The aim of this study was to assess any differences in the expression patterns of these molecules on regulatory T cells of patients with inflammatory bowel disease (IBD). Methods: The study subjects were 15 patients with IBD (ulcerative colitis=7, Crohn's disease=8) and 7 healthy individuals. Peripheral blood mononuclear cells were obtained by centrifugation of fresh blood over Ficoll-paque and stained with anti-CD4, CD25, and GITR antibodies. For CTLA-4 staining, cells were fixed and permeabilized by saponin prior to incubation with anti-CTLA-4 antibody. Cell samples were analyzed on a FACScaliber. Each patient was investigated both in the active and inactive state. Results: The percentage of CD4^+CD25^+ cells was significantly higher in active than in inactive ulcerative colitis. In contrast, the percentage of CD4^+CD25^<high> cell was significantly higher in active Crohn's disease than in healthy individuals. CTLA-4^+ cell counts in CD4^+CD25^<high> T cells were significantly lower in active Crohn's disease than in control and inactive Crohn's disease patients, although no difference was observed between active and inactive ulcerative colitis. In contrast, GITR expression was significantly higher in active than inactive Crohn's disease. Conclusion: Our results suggest that CD4 + CD25high T cells are probably involved in the pathogenesis of Crohn's disease