Regulation of T cell Differentiation

项目1：以前的研究表明Notch 1和TGF-β信号通路是相互加强的。 鉴于最近的证据表明调节性T细胞（Treg）效应子功能由TGF-信号传导介导，我们研究了Notch 1信号传导是否也参与Treg效应子功能。 最初的研究表明，Notch 1配体，特别是Jagged 1，存在于Treg上，并且确实，用抗Jagged 1或阻断性抗Notch 1抗体阻断Notch 1信号传导抑制Treg抑制剂功能。 然后，我们发现Notch 1激活（NICD）产生的信号组分与TGF-β信号（pSmad 3）产生的信号组分发生物理相互作用。此外，这种相互作用具有功能性下游效应，因为NICD的过表达促进pSmad 3易位到细胞核，并增强与荧光素酶报告基因连接的Smad敏感启动子的pSmad 3转录活性。 最后，我们发现阻断TGF-β信号传导和Notch信号传导对Treg抑制功能没有累加抑制作用。 这些结果与Notch 1信号传导增强和促进TGF-β介导的TGF 1 α效应功能的结论一致。 项目2：最近的研究表明，TGF-β与IL-6一起诱导产生IL-17的T细胞（Th 17）T细胞的分化。因此，我们检查了CD 4 + CD 25 + Foxp 3+调节性T细胞（TcR），即，先前显示产生TGF-β的细胞充当Th 17诱导剂。 我们发现，在活化时，纯化的CD 25 + T细胞（或从Foxp 3-GFP敲入小鼠获得的分选的GFP+ T细胞）产生大量的可溶性TGF-β，并且当在IL-6存在下与CD 4 + CD 25-Foxp 3- T细胞一起培养时，诱导后者分化成Th 17细胞。也许更重要的是，在活化后，CD 4 + CD 25 + Foxp 3+（GFP+）T细胞本身在IL-6存在下（并且在不存在外源性TGF-β的情况下）分化成Th 17细胞。这些结果表明，CD 4 + CD 25 + Foxp 3+调节性T细胞可以作为Th 17细胞的诱导物起作用，并且可以分化成Th 17细胞。因此，它们对我们理解调节性T细胞功能及其可能的治疗用途具有重要意义。

Project 1:Previous studies have shown that the Notch1 and TGF-β signaling pathways are mutually re-enforcing. Given recent evident that regulatory T cell (Treg) effector function is mediated by TGF-β signaling, we investigated if Notch1 signaling also participated in Treg effector function. Initial studies showed that Notch1 ligands, particularly Jagged1, is present on Tregs and that, indeed, blockade of Notch1 signaling with a anti-Jagged1 or a blocking anti-Notch1 antibody inhibits Treg suppressor function in vitro. We then showed that a signaling component generated by Notch1 activation (NICD) physically interacts with a signaling component generated by TGF-β signaling (pSmad3). Furthermore, this interaction has functional downstream effects since over-expression of NICD facilitates pSmad3 translocation to the nucleus and enhances pSmad3 transcriptional activity of a Smad sensitive promoter linked to a luciferase reporter. Finally, we showed that blockade of TGF-β signaling and Notch signaling did not have additive inhibitory effects on Treg suppressor function. These results are consistent with the conclusion that Notch1 signaling enhances and facilitates TGF-β-mediated effector function of Tregs. Project 2: Recent studies have shown that TGF-β together with IL-6 induce the differentiation of IL-17-producing T cells (Th17) T cells. We therefore examined if CD4+CD25+Foxp3+ regulatory T cells (Tregs), i.e., cells previously shown to produce TGF-β, serve as Th17 inducers. We found that upon activation purified CD25+ T cells (or sorted GFP+ T cells obtained from Foxp3-GFP knock-in mice) produce high amount of soluble TGF-β and when cultured with CD4+CD25-Foxp3- T cells in the presence of IL-6 induce the latter to differentiate into Th17 cells. Perhaps more importantly, upon activation, CD4+CD25+Foxp3+(GFP+) T cells themselves differentiate into Th17 cells in the presence of IL-6 (and in the absence of exogenous TGF-β). These results indicate that CD4+CD25+Foxp3+ regulatory T cells can function as inducers of Th17 cells and can differentiate into Th17 cells. They thus have important implications to our understanding of regulatory T cell function and their possible therapeutic use.