Immune cell proliferation is suppressed by the interferon‐γ‐induced indoleamine 2,3‐dioxygenase expression of fibroblasts populated in collagen gel (FPCG)

Abstract

Indoleamine 2,3-dioxygenase (IDO), a tryptophan-catabolizing enzyme, is an intracellular enzyme possessing various immunosuppressive properties. Here, we report the possible use of this enzyme to suppress proliferation of immune cells cocultured with IDO-expressing fibroblasts of an allogenic skin substitute. Fetal skin fibroblasts embedded within bovine collagen were treated with cytokine interferon-gamma (IFN-gamma) to induce expression of IDO mRNA and protein. Expression of IDO mRNA was evaluated by Northern analysis. IDO enzyme activity was evaluated by measurement of kynurenine and tryptophan levels in the IFN-gamma untreated and treated fibroblasts. The results of Northern analysis showed a dose-dependent increase in expression of IDO mRNA in response to various concentrations of IFN-gamma used. The levels of kynurenine and tryptophan measured, as the bioactivity of IDO, were significantly different in the IFN-gamma treated fibroblasts, compared to those of controls (P < 0.001). In a lasting effect experiment, the expression of IDO mRNA was gradually reduced to an undetectable level within 32 h of IFN-gamma removal. The results of Western blot analysis, however, revealed a significantly longer (192 h) lasting effect of IFN-gamma on IDO protein level, relative to that of mRNA expression. To demonstrate immunosuppressive effects of IDO on proliferation of immune cells, IDO-expressing fibroblasts were cocultured with peripheral blood mononuclear cells (PBMC) for a period of 5 days. The results of (3)H-thymidine incorporation showed a significant reduction in proliferation of PBMC when cocultured with IDO-expressing fibroblasts, compared to those cocultured with non-IDO-expressing fibroblasts (P < 0.001). Furthermore, addition of IDO-inhibitor (1-methyl-d-tryptophan) reversed the suppressive effects of IDO on PBMC proliferation in a dose-dependant fashion. To test the viability of immune cells cocultured with IDO-expressing fibroblasts, FACS analysis of the PI stained PBMC was conducted and no significant difference was found between these cells and the controls. In another set of experiments, we showed that migration rate and subsequent proliferation of IDO-expressing fibroblasts are also the same as those of control cells. In conclusion, IDO-expressing allogenic fibroblasts embedded within collagen gel suppress the proliferation of allogenic immune cells, while they still remain viable in this IDO-induced tryptophan-deficient culture environment.

Ability of IDO To Attenuate Liver Injury in α-Galactosylceramide–Induced Hepatitis Model

“…It was previously reported that IDO induces inhibition of immune cell (e.g., NK cells and T cells) proliferation (25,26). In our data, intrahepatic MNCs from IDO-KO mice treated with a-GalCer were apoptosis resistant with those from WT mice.…”

Interferon-γ-modified dendritic cells suppress B cell function and ameliorate the development of experimental autoimmune myasthenia gravis

“…would interact with other immune cells, first in the regional lymph nodes and then in the spleen, within the first week after administration. Indoleamine 2,3-dioxygenase (IDO) is an intracellular tryptophan-catabolizing enzyme that possesses various immunosup- pressive properties [19]. Antigen-presenting cells have been shown to suppress T cell proliferation via a rapid and selective IDO-mediated degradation of tryptophan [20].…”

Cutting Edge: Human Eosinophils Regulate T Cell Subset Selection through Indoleamine 2,3-Dioxygenase

“…Rabbit anti-human IDO Ab was generated in-house (11). rIL-2, IL-3, IL-5, and GM-CSF were obtained from BD Biosciences (BD Pharmingen, Mississauga, Ontario, Canada).…”