Analysis for a role of dectin-2 in ultraviolet-light-induced immune tolerance

众所周知，紫外线B光(UV)会损害接触性超敏反应(CHS)的诱导期，从而诱导耐受性。虽然这一现象几十年来一直被认为是一种重要的生物学现象，但其背后的免疫学机制尚不完全清楚，但Dectin-2是c型凝集素的新成员，它选择性地表达在表皮朗格汉斯细胞(LC)上。在这里，我们研究了Dectin-2在UVB诱导的CHS免疫耐受中的作用。为此，我们首先产生了一种可溶性的Dectin-2(SDec2)。SDec2是通过重组技术获得的。简而言之，将Dectin-2的胞外区亚克隆到表达载体上，然后转入E.cli并在其中表达。从大肠杆菌裂解产物中回收、纯化可溶性重组蛋白，体外测定其对同源Dectin-2的抑制活性。为了解决最初的问题，在…之前给小鼠注射了sDec2更多的诱导或诱导CHS。然而，无论注射什么，CHS都没有受到影响，这表明Dectin-2介导的通路不参与这一事件，相反，如果Dectin-2被注射到已经以1000J/m^2的紫外线照射连续4天的小鼠身上。然后这些小鼠被半抗原致敏和激发，导致CHS的全部结局，而紫外线显著抑制那些只被致敏和激发但不注射sDec2的CHS。这表明Dectin-2参与了紫外线对CHS的损伤。为了验证它是否参与了紫外线诱导的耐受，紫外线损伤了CHS的小鼠14天不治疗，然后再敏化和再激发，结果与未暴露的动物相比，CHS明显受到抑制，表明诱导了耐受。然而，在第一次致敏前注射sDec2-可恢复CHS，并未导致耐受性。根据耐受性是通过产生T抑制细胞(Ts)来调节的这一事实，上述结果表明Dectin-2介导了半抗原特异性ts的诱导。这进一步证实，sDec2结合但不结合的T细胞在转移到幼稚接受者时会削弱CHS的诱导。为了对sDec2结合的ts进行表型分析，FACS分析表明，紫外线诱导的ts在其表面表达CD4和CD25，类似于目前高亮的调节性T细胞(Ir)。T细胞受体是新命名的T细胞，具有预防自身免疫性疾病的发生的能力，包括I型糖尿病或多发性硬化症小鼠模型。总之，我们的研究揭示了紫外线诱导的耐受性是通过Tr的产生来调节的，而不是

It is known that ultraviolet B light(UV) impairs the induction phase of contact hypersensitivity responses(CHS) and subsequently induces tolerance. Although this phenomenon has been appreciated for decades to be a major biological importance, immunological mechanisms underlying this event are not completely clear yet Dectin-2 is a new member of c-type lectin, which is selectively expressed on epidermal langerhans cells(LC). Here we examined roles of dectin-2 in UVB-induced immune tolerance of CHS. To do so, we first generated a soluble form of dectin-2(sDec2). sDec2 was generated by use of recombinant technology. Briefly, an extracellular domain of dectin-2 cDNA was subcloned to an expression vector, which was then transfected to and expressed in E.cli. Soluble recombinant protein was recovered from lysates of coli, purified and its inhibitory activity to cognate dectin-2 was ascertained in in vitro assays. To address the initial questions, sDec2 was injected to mice before either the … More induction or the elicitation of CHS. However, CHS was not affected regardless of the injection, indicating that dectin-2-mediated pathways are not involved in this event By contrast, if dectin-2 was injected to mice which was already UV exposed at 1,000J/m^2 for 4 consecutive days. Those mice were then hapten sensitized and challenged, leading to full outcome of CHS, while UV significantly suppressed CHS when those were only sensitized and challenged but not sDec2-injected. This indicates that dectin-2 is involved in the UV-mediated impairment of CHS. To check for its involvement in UV-induced tolerance, mice whose CHS were impaired by UV were left untreated for 14 days and then resensitized and rechallenged, resulting that CHS was significantly suppressed as compared to unexposed animals, indicating that tolerance was induced. However, sDec2-injection before the first sensitization restored CHS and did not lead to tolerance. Based on the fact that tolerance is mediated via generation of T suppressor cells(Ts), the above mentioned results indicate that dectin-2 mediates the induction of hapten specific Ts. This was further confirmed as sDec2-bound, but not unbound T cells impaired the induction of CHS upon transfer to naive recipients. To phenotypically characterize sDec2-bound Ts, FACS analysis was conducted, demonstrating that UV-induced Ts express CD4 and CD25 on their surface resembling currently hightlighted regulatory T cells(Ir). Tr are the re-nomenclatured Ts, which possess the ability to prevent the onset of autoimmune disorders, including type I diabetes mellitus or multiple sclerosis of murine models. Together, our study disclosed that UV-induced tolerance is mediated via generation of Tr, Less