In vitro generation of regulatory CD8+ T cells similar to those found in mice with anterior chamber-associated immune deviation

PURPOSE. When injected intravenously into naive mice, peritoneal exudate cells (PECs) incubated with ovalbumin (OVA) in the presence of transforming growth factor (TGF)-beta 2 induce immune deviation similar to that evoked by injection of OVA into the anterior chamber of the eye. Intraocular antigen injection elicits two distinct populations of regulatory T cells that impair delayed hypersensitivity (DH) by two different mechanisms: a CD4(+) T cell that suppresses the induction of DH (afferent) and a CD8(+) T cell that inhibits DH expression. In an effort to understand the origin and mechanism of action of these regulatory cells, CD8(+) T cells from OVA-specific T cell receptor (Tcr) transgenic mice (OT-1) were used. METHODS. CD8(+) T cells were harvested from Tcr transgenic OT-I mice whose Tcr recognize an OVA peptide in the context of the class I major histocompatibility complex molecule K-b. These cells were stimulated in vitro with OVA-pulsed PECs exposed (or not) to TGF-beta 2, then analyzed for their capacity to proliferate, to secrete various cytokines, to lyse OVA-expressing target cells, and to regulate bystander T cells in vitro and in vivo. RESULTS. When OVA-pulsed PECs were used in vitro as stimulators, responding OT-I T cells proliferated and preferentially secreted interferon (IFN)-gamma, interleukin (IL)-2, and tumor necrosis factor (TNF)-alpha, rather than IL-4 and IL-10. When the stimulator PECs were pretreated with TGF-beta 2 and then pulsed with OVA, responding OT-I T cells proliferated even more swiftly, but they secreted significantly less IFN-gamma, IL-2, and TNF-alpha, and no IL-4 or IL-10. OT-I cells, which constitutively display cytotoxicity toward OVA-expressing target cells, lost this activity when stimulated with OVA-pulsed, TGF-beta 2-pretreated PECs. Moreover, OT-I T cells stimulated in this manner displayed the capacity to inhibit proliferation of OVA-primed T cells exposed to OVA in vitro and to suppress in vivo the expression of OVA-triggered DH. CONCLUSIONS. OVA-pulsed PECs, pretreated with TGF-beta 2, coerce naive OVA-specific CD8(+) T cells to become efferent regulators of DH similar to the regulatory T cells evoked by intraocular injection of OVA.