期刊
JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH
卷 30, 期 -, 页码 -出版社
BMC
DOI: 10.1186/1756-9966-30-82
关键词
Indoleamine-Pyrrole 2, 3-Dioxygenase; breast neoplasms; immune tolerance; CHO Cells; regulatory T-Lymphocytes
类别
资金
- National Natural Science Foundation of China [30972694, 81072159]
- Tianjin Municipal Education Commission, P. R. China [20090133, 20090217]
Introduction: The inflammatory enzyme indoleamine 2, 3-dioxygenase (IDO) participates in immune tolerance and promotes immune escape of IDO+ tumors. A recent hypothesis suggested that IDO may contribute to the differentiation of new T regulatory cells (Tregs) from naive CD4+ T cells. In this study we investigated the role of IDO in induction of immunosuppression in breast cancer by increasing the apoptosis of T cells and the proportion of Tregs. Methods: An IDO expression plasmid was constructed and Chinese hamster ovary (CHO) cells were stably transfected with human IDO. Purified CD3+ T cells were isolated from the peripheral blood monouclear cells of breast cancer patients. After co-culturing IDO expressing or untransfected (control) CHO cells with T cells, T cells apoptosis were determined by flow cytometry analysis and annexin-V and PI staining. The proportion of the regulatory T cell (Tregs [CD4 + CD25 + CD127-]) subset was measured by flow cytometry analysis. T cells total RNA and cellular protein samples were isolated for detecting Foxp3 gene and protein expression. Results: IDO transgenic CHO cells yielded high levels of IDO enzymatic activity, resulting in complete depletion of tryptophan from the culture medium. We found that apoptosis occurred in 79.07 +/- 8.13% of CD3+ T cells after co-cultured with IDO+ CHO cells for 3 days and the proportion of CD4 + CD25 + CD127- T cells increased from 3.43 +/- 1.07% to 8.98 +/- 1.88% (P < 0.05) as well. The specific inhibitor of IDO, 1-MT efficiently reversed enhancement of T cells apoptosis and amplification of Tregs in vitro. Increased expression of Foxp3, a key molecular marker of Tregs, was confirmed by RT-PCR, real-time RT-PCR and Western blot analysis at the same time. Conclusions: These results suggest that IDO helps to create a tolerogenic milieu in breast tumors by directly inducing T cell apoptosis and enhancing Treg-mediated immunosuppression.
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