In vitroexpansion or induction of regulatory T cells from olcerative colitis patients

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CD4^+CD25^+ regulatory T cells (Treg) possess broad range of immunoregulatory activity which regulates autoimmune disorders including inflammatory bowel diseases in animal models. Aiming the clinical application of Treg for the treatment of ulcerative colitis (UC), we investigated whether Treg can be expanded or induced in vitro. We stimulated Tregs isolated from the leukapheresis products from UC patients with anti-CD3/anti-CD28-bearing beads and IL-2. Treg could be increased 18 times in 10 days culture. Cultured Treg maintained the expression of a Treg-specific marker, FOXP3 and significantly suppressed the CD4^+ T cell-proliferation upon coculture. Thus, Treg can be expanded in vitro.Next, we tested whether Treg can be induced in vitro CD4^+CD255non-Treg were stimulated and cultured in the presence of TGF-β1. Cultured cells expressed FOXP3 and suppressed the CD4^+ T cell-proliferation upon coculture. Therefore, Treg can be even induced from non-Treg in vitro.Then, we investigate whether induced Treg are able to suppress intestinal inflammation. We induced Treg from CD4^+ T cells stimulating with anti-CD3/anti-CD28 and IL-2 in the presence of rapamycin (RAPA). When CD4^+ T cells were stimulated in the presence of RAPA, 17% was CD25^+FoxP3^+ at the end of 21-day culture. In contrast, few cells stimulated without RAPA expressed CD25 and FoxP3. CB-17 Scid mice were transferred with CD4^+CD62L^+CD25^-T cells, which developed chronic persistent colitis. Co-transfer of CD4^+ T cells stimulated in the presence of RAPA suppressed colitis as naturally-occurring Treg, while CD4^+ T cells cultured in the absence of RAPA failed to ameliorate intestinal inflammation. Therefore, RAPA induced Treg, which are capable of suppressing colitis.These results indicate that Treg can be expanded or induced in vitro and transfer of induced Treg may be efficacious for the treatment of inflammatory bowel diseases.