漿狀樹突細胞增強類鐸受體調控之B淋巴球活化與分化

Abstract

B淋巴球藉由活化與分化並分泌抗體在適應性免疫反應扮演著重要的角色。而另一方面，漿狀樹突細胞(pDC)可藉由類鐸受體(Toll-like receptor, TLR)辨識病毒，產生大量的第一型干擾素(Type I IFN, IFN-I)在先天性抗病毒免疫反應中扮演至關重要的角色。B 細胞與pDC曾被報導可增強體液免疫反應，然而pDC如何協調B細胞分化的詳細機制仍尚未明確。因此我們使用TLR7配體R848的刺激並建立小鼠脾臟pDC和B細胞體外共培養系統。藉由此系統我們發現pDC可增強TLR7引起的B細胞活化，並增加MHCII，CD86和CD69表現，顯著提升IL-6和CXCL10分泌。此外，B/pDC藉由可溶性因子和細胞接觸依賴性方式促進漿細胞分化，但會降低B細胞存活和生發中心B細胞 (germinal center B cell, GC B）的形成。另外我們發現了一群表現CD19+的pDC，在TLR7刺激下會以依賴IFNαR和STAT1訊息傳遞的方式增生。而B細胞子集部分，在 pDC/R848刺激下，濾泡B細胞（Follicular B cell）比邊緣區B細胞 (Marginal Zone B)的增強反應更敏感。此外，在B/pDC共培養系統中我們發現雖然IFNαR訊息傳遞是必須的，但只是IFN-I並不足以誘導B細胞完全活化。有趣的是，不僅B細胞，pDC的IFNαR信號傳導也有助於增強反應。而B/pDC對於STAT1，一個IFNαR下游分子，訊息傳遞的需求也相似於IFNαR的結果。當B細胞與pDC缺乏IFNαR與STAT1時會嚴重減少細胞激素與細胞趨化因子的分泌，進而影響漿細胞的存活與分化。在活體動物實驗方面，把B細胞送入Rag1基因剔除老鼠中去重建B細胞反應時若去除pDC會減少R848誘導的IgM分泌，這結果意味著pDC在胸腺非依賴性抗體反應扮演重要的角色。總而言之，我們在體外培養與活體實驗均驗證pDC可增強B細胞活化、增生與漿細胞分化並證實在B細胞與pDC的IFN-I訊息傳遞可正向調控體液免疫反應。

B cells function to secret antibodies upon activation and differentiation, which play an important role in adaptive immunity. Plasmocytoid dendritic cells (pDCs), on the other hand, are a rare population known for robust type I IFN (IFN-I) production upon TLR stimulation and are crucial for antivirus response of innate immunity. B cells and pDCs can cooperate to boost the humoral immunity. However, the detailed mechanisms of how pDCs orchestrate B cell differentiation remains elusive. To investigate the mechanisms, we set up an in vitro coculture system of splenic pDCs and B cells and stimulated with R848, a TLR7 agonist. pDCs enhanced TLR7-mediated B cell activation with increased expression of MHCII, CD86 and CD69 and significantly augmented the production of IL-6 and CXCL10. Moreover, increased proliferation and plasma cell differentiation but decreased survival and germinal center B cell (GC B) formation were also found through both soluble factor- and cell-to-cell contact- dependent manner. Interestingly, a distinct subset of CD19+ pDCs proliferated in response to TLR7 stimulation in a IFNαR and STAT1 signaling-dependent manner. In addition to pDC, follicular B cells (FO B) were more sensitive to pDC/R848 stimulation than marginal zone B cells (MZ B). Among soluble factors secreted in the coculture system, IFN-I was found to be critical for the enhanced response. B cells lacking IFNαR showed impaired responses. Moreover, IFNαR signaling was necessary but not sufficient to induce full B cell activation. Interestingly, IFNαR signaling in pDCs also contributed to the enhanced responses. A similar phenotype was also observed for cell-intrinsic requirement of STAT1 signaling for both B cells and pDCs. Lack of IFNαR or STAT1 signaling in pDC and B cell severely impaired the production of proinflammatory cytokines and chemokines, especially IL-6 and CXCL10 which are an impotent for plasma cell survival and differentiation. Depletion of pDCs in Rag1-/- mice that had been adoptively transferred with B cells also reduced IgM production induced by R848, suggesting an essential role of pDC in T-independent antibody response in vivo. Collectively, we have identified that murine splenic pDCs directly enhance B cell activation, proliferation and plasma cell differentiation in vitro and in vivo, and defined a crucial role of IFN-I signaling in both pDCs and B cells to positively modulate the humoral immunity.