The percentage of Treg cells in the tumour tissue was 15·4%, with a standard deviation (s.d.) of 9·9% (range: 7·2–23·6%). There were multiple immune cell populations in the tumour microenvironment. The relationships were evaluated further between Th17 cells and other immune cell subsets, such as IFN-γ+ CD4+ T cells and Treg Talazoparib mouse cells in the same tumours. Flow cytometry analysis revealed that the proportion of Th17 cells was correlated positively with that of IFN-γ+ CD4+ T cells, but correlated inversely with Treg cells in the same tumour microenvironment (Fig. 6a). Several studies suggested

that instillations of IL-2 into the urinary bladder might be effective for treatment of superficial bladder cancer, and recent data also indicated that IL-2 might play a role in regulating the TH17/Treg balance in the tumour microenvironment, so we investigated the potential effects of IL-2 on Th17 and Treg cell differentiation in vitro. A Treg subset from tumour

samples was sorted ex vivo by flow cytometry cell sorting and the purity of the separated cells subset was confirmed to be >97%. Next, we analysed IL-17 production of sorted Treg after stimulation with the autologous irradiated CD3– fraction in the presence of IL-2 for 10 days. As shown in Fig. 6b, Th17 cells were clearly Selleckchem LDK378 detectable in populations from the purified Treg cell fractions. However, no proliferation or IL-17 production was observed after culture of tumour Treg stimulated by the

autologous irradiated CD3– fraction in the absence of IL-2. We also failed to detect any significant proliferation or IL-17 production when the purified tumour Treg cells were cultured with IL-2 alone. To characterize further the tumour Treg after in vitro expansion, we assessed IL-17 production and FoxP3 expression simultaneously by these cells stimulated by the autologous irradiated CD3– fraction in the presence of IL-2. As shown in Fig. 6c, the sorted Treg gradually expressed IL-17 and lost FoxP3 expression. The proportion of Treg co-expressing FoxP3 and IL-17 was increased gradually in the early days, but decreased as culture time went on. Co-culture with responder CD4+CD25– cells and Treg was used to evaluate the function change of tumour Treg after conversion. As shown in Fig. 6d, compared with the tumour Treg before stimulation, the tumour 4-Aminobutyrate aminotransferase Treg after conversion exhibited hampered inhibition of responder CD4+CD25– cell proliferation, which may be associated with down-regulated FoxP3 expression. Little IFN-γ production was found in the Treg cultures (Fig. 6e). Studies have shown that tumour is potentially immunogenic and that the host immune response influences survival [27]. It has been shown that tumour-infiltrating effector T cells correlates with improved prognoses of several types of cancer, whereas tumour-infiltrating Treg cells are associated negatively with patient outcome [28,29].