Rosendahl Alexander, Speletas Matthaios, Leandersson Karin, Ivars Fredrik, Sideras Paschalis
AstraZeneca R & D Lund, Department of Bio & Molecular Sciences, Scheelevägen 2, 221 87 Lund, Sweden.
Int Immunol. 2003 Dec;15(12):1401-14. doi: 10.1093/intimm/dxg139.
Members of the transforming growth factor (TGF)-beta family play pivotal roles in the control of differentiation, proliferation and tolerance in peripheral T cells. Recently, they have been implicated in thymic selection, but their role is so far not well characterized. In the present study, we demonstrate that specific thymocyte populations are under the influence of either the TGF-beta and/or Activin pathway, and transduce signals into the nucleus via phosphorylated Smad2 (pSmad2). Thymocytes in the medulla and in the subcapsular zone expressed nuclear translocated pSmad2, a hallmark of active TGF-beta/Activin receptor signaling. When analyzed at the cellular level, the pSmad2(+) cells were confined to the double-negative (DN) and single-positive (SP) subpopulations. Moreover, the most immature DN thymocytes (CD44(+)CD25(-) and CD44(+)CD25(+)) expressed higher levels of pSmad2 compared to the more mature DN. In vitro stimulation demonstrated that pure CD44(+)CD25(-), CD44(+)CD25(+) and CD44(+)CD25(+) thymocytes respond to ActivinA, while the mature CD4(+) and CD8(+) SP thymocytes respond to TGF-beta stimulation measured as enhanced phosphorylation of Smad2. Double staining of pSmad2(+) cells with either the Activin type I receptor, ALK4, or the TGF-beta type I receptor, ALK5, demonstrated that pSmad2(+) DN cells exhibited high levels of immunoreactivity to ALK4 and moderate levels of immunoreactivity to the TGF-beta-responsive ALK5 receptor. In sharp contrast, the SP pSmad2(+) cells were predominately ALK5(+). Collectively, our results demonstrate that early and late thymocytes express pSmad2 in the nuclei in vivo. The functional experiments in vitro suggest that members of the TGF-beta family (TGF-beta or Activin) may play important non-redundant roles during different stages of thymopoiesis.
转化生长因子（TGF）-β家族成员在控制外周T细胞的分化、增殖和耐受性方面发挥着关键作用。最近，它们被认为与胸腺选择有关，但到目前为止其作用尚未得到充分表征。在本研究中，我们证明特定的胸腺细胞群体受到TGF-β和/或激活素途径的影响，并通过磷酸化的Smad2（pSmad2）将信号转导至细胞核。髓质和被膜下区的胸腺细胞表达核转位的pSmad2，这是活跃的TGF-β/激活素受体信号传导的标志。在细胞水平分析时，pSmad2(+)细胞局限于双阴性（DN）和单阳性（SP）亚群。此外，与更成熟的DN胸腺细胞相比，最不成熟的DN胸腺细胞（CD44(+)CD25(-)和CD44(+)CD25(+)）表达更高水平的pSmad2。体外刺激表明，纯CD44(+)CD25(-)、CD44(+)CD25(+)和CD44(+)CD25(+)胸腺细胞对激活素A有反应，而成熟的CD4(+)和CD8(+)SP胸腺细胞对TGF-β刺激有反应，表现为Smad2磷酸化增强。用激活素I型受体ALK4或TGF-βI型受体ALK5对pSmad2(+)细胞进行双重染色表明，pSmad2(+)DN细胞对ALK4表现出高水平的免疫反应性，对TGF-β反应性ALK5受体表现出中等水平的免疫反应性。形成鲜明对比的是，SP pSmad2(+)细胞主要是ALK5(+)。总体而言，我们的结果表明早期和晚期胸腺细胞在体内细胞核中表达pSmad2。体外功能实验表明，TGF-β家族成员（TGF-β或激活素）可能在胸腺生成的不同阶段发挥重要的非冗余作用。
