Data_Sheet_3_The Axial Organ and the Pharynx Are Sites of Hematopoiesis in the Sea Urchin.docx

Background: The location of coelomocyte proliferation in adult sea urchins is unknown and speculations since the early 1800s have been based on microanatomy and tracer uptake studies. In adult sea urchins (Strongylocentrotus purpuratus) with down-regulated immune systems, coelomocyte numbers increase in response to immune challenge, and whether some or all of these cells are newly proliferated is not known. The gene regulatory network that encodes transcription factors that control hematopoiesis in embryonic and larval sea urchins has not been investigated in adults. Hence, to identify the hematopoietic tissue in adult sea urchins, cell proliferation, expression of phagocyte specific genes, and expression of genes encoding transcription factors that function in the conserved regulatory network that controls hematopoiesis in embryonic and larval sea urchins were investigated for several tissues.
Results: Cell proliferation was induced in adult sea urchins either by immune challenge through injection of heat-killed Vibrio diazotrophicus or by cell depletion through aspiration of coelomic fluid. In response to either of these stimuli, newly proliferated coelomocytes constitute only about 10% of the cells in the coelomic fluid. In tissues, newly proliferated cells and cells that express SpTransformer proteins (formerly Sp185/333) that are markers for phagocytes are present in the axial organ, gonad, pharynx, esophagus, and gut with no differences among tissues. The expression level of genes encoding transcription factors that regulate hematopoiesis show that both the axial organ and the pharynx have elevated expression compared to coelomocytes, esophagus, gut, and gonad. Similarly, an RNAseq dataset shows similar results for the axial organ and pharynx, but also suggests that the axial organ may be a site for removal and recycling of cells in the coelomic cavity.
Conclusions: Results presented here are consistent with previous speculations that the axial organ may be a site of coelomocyte proliferation and that it may also be a center for cellular removal and recycling. A second site, the pharynx, may also have hematopoietic activity, a tissue that has been assumed to function only as part of the intestinal tract.

背景：成年紫海胆（Strongylocentrotus purpuratus）的体腔细胞（coelomocyte）增殖位点至今尚不明确，自19世纪早期以来的相关推测均基于显微解剖学（microanatomy）与示踪摄取研究（tracer uptake studies）。针对免疫系统下调的成年紫海胆，其体腔细胞数量会在受到免疫刺激（immune challenge）后上升，但目前尚不清楚这些细胞中有多少是新增殖产生的。调控胚胎及幼体海胆造血（hematopoiesis）过程的转录因子（transcription factor）编码基因调控网络（gene regulatory network），尚未在成体海胆中开展研究。因此，为明确成体海胆的造血组织，本研究对多个组织的细胞增殖情况、吞噬细胞（phagocyte）特异性基因的表达，以及调控胚胎与幼体海胆造血过程的保守调控网络中相关转录因子编码基因的表达进行了检测。
结果：通过注射热灭活的重氮营养弧菌（Vibrio diazotrophicus）进行免疫刺激，或通过抽吸体腔液（coelomic fluid）造成细胞耗竭，均可诱导成年海胆发生细胞增殖。在这两种刺激下，新增殖的体腔细胞仅占体腔液中细胞总数的约10%。在各组织中，新增殖的细胞以及表达SpTransformer蛋白（SpTransformer，原名为Sp185/333，吞噬细胞标志物）的细胞可在轴器（axial organ）、性腺、咽、食道及肠道中被检测到，且各组织间无显著差异。调控造血过程的转录因子编码基因的表达水平显示，轴器与咽部的基因表达量均高于体腔细胞、食道、肠道及性腺。类似地，RNA测序（RNAseq）数据集的结果与上述发现一致，但同时提示轴器可能也是体腔腔内细胞清除与回收的场所。
结论：本研究结果与此前的推测一致，即轴器可能是体腔细胞增殖的场所，同时也可作为细胞清除与回收的中枢。此外，咽部作为此前仅被认为仅发挥肠道附属功能的组织，同样具备造血活性，构成第二个造血位点。