TRANSITIONAL DENDRITIC CELLS ARE DISTINCT FROM CONVENTIONAL DC2 PRECURSORS AND MEDIATE PRO-INFLAMMATORY ANTIVIRAL RESPONSES

High-dimensional approaches revealed emerging heterogeneity within dendritic cells (DC), including a population of transitional DC (tDC) present in mouse and human. However, tDC origin and relationship to other DC subsets are not fully understood. Here, we show that tDC are distinct from other well-characterized DC and conventional DC precursors (pre-cDC). We demonstrate that tDC originate from bone marrow progenitors shared with plasmacytoid DC (pDC). In the periphery, tDC contribute to the pool of ESAM+ type 2 DC (DC2), and these DC2 harbor pDC-related developmental features. Different from pre-cDC, tDC have lower turnover, capture antigen, respond to stimuli, and activate antigen-specific naive T cells, all characteristics of differentiated DC. Different from pDC, viral sensing by tDC results in IL-1b secretion and fatal immune pathology in a murine coronavirus model. Our findings suggest that tDC are a distinct pDC-related subset with a DC2 differentiation potential and unique pro-inflammatory function during viral infections. Examination of immune cells in murine bone marrow and spleen. Cells were CD135 enriched and sorted for live cells. Cells were barcoded, pooled and run on 2 lanes. Sorted splenocytes were mixed 1:1 from 2 gates: CD3-/CD19-/NK1.1-/Ly6G- and CD3-/CD19-/NK1.1-/Ly6G-/Xcr1-/CD11blo to enrich for tDC. BM cells were gated as CD3-/CD19-/NK1.1-/Ly6G-. After sort, cells were counted and resuspended at a concentration of 600 cells per uL. Cells suspensions were processed for 10X Genomics.高维分析技术揭示了树突状细胞（dendritic cells, DC）内存在的新兴细胞异质性，其中包括一类存在于小鼠和人类体内的过渡型树突状细胞（transitional DC, tDC）亚群。然而，目前对于tDC的起源及其与其他DC亚群的关系尚未完全阐明。本研究证实，tDC与其他已被充分表征的DC以及常规树突状细胞前体（conventional DC precursors, pre-cDC）均存在显著差异。我们证明，tDC起源于与浆细胞样树突状细胞（plasmacytoid DC, pDC）共享的骨髓祖细胞。在外周组织中，tDC可补充ESAM阳性2型树突状细胞（ESAM+ type 2 DC, DC2）的细胞库，且这类DC2携带pDC相关的发育特征。与pre-cDC不同，tDC具有更低的细胞更新率、抗原捕获能力、刺激响应性以及激活抗原特异性初始T细胞的能力，上述均为成熟DC的典型特征。与pDC不同的是，在小鼠冠状病毒模型中，tDC的病毒感知会诱导IL-1b分泌并引发致命性免疫病理损伤。我们的研究结果表明，tDC是一类独特的pDC相关亚群，具备DC2分化潜能，并在病毒感染过程中发挥独特的促炎功能。本研究对小鼠骨髓与脾脏中的免疫细胞开展了分析：首先对细胞进行CD135富集，并分选活细胞；随后对细胞进行条码标记、混合后在两个测序泳道上上机检测。将分选得到的脾细胞按1:1的比例，从两个门群中取样混合：CD3-/CD19-/NK1.1-/Ly6G-以及CD3-/CD19-/NK1.1-/Ly6G-/Xcr1-/CD11blo，以实现tDC的富集。骨髓细胞的门控设为CD3-/CD19-/NK1.1-/Ly6G-。分选完成后，对细胞进行计数并重悬至每微升600个细胞的浓度，随后将细胞悬液使用10X Genomics平台进行处理。