DataSheet_2_Tipping the balance between erythroid cell differentiation and induction of anemia in response to the inflammatory pathology associated with chronic trypanosome infections.xlsx

Infection caused by extracellular single-celled trypanosomes triggers a lethal chronic wasting disease in livestock and game animals. Through screening of 10 Trypanosoma evansi field isolates, exhibiting different levels of virulence in mice, the current study identifies an experimental disease model in which infection can last well over 100 days, mimicking the major features of chronic animal trypanosomosis. In this model, despite the well-controlled parasitemia, infection is hallmarked by severe trypanosomosis-associated pathology. An in-depth scRNA-seq analysis of the latter revealed the complexity of the spleen macrophage activation status, highlighting the crucial role of tissue resident macrophages (TRMs) in regulating splenic extramedullary erythropoiesis. These new data show that in the field of experimental trypanosomosis, macrophage activation profiles have so far been oversimplified into a bi-polar paradigm (M1 vs M2). Interestingly, TRMs exert a double-sided effect on erythroid cells. On one hand, these cells express an erythrophagocytosis associated signature. On another hand, TRMs show high levels of Vcam1 expression, known to support their interaction with hematopoietic stem and progenitor cells (HSPCs). During chronic infection, the latter exhibit upregulated expression of Klf1, E2f8, and Gfi1b genes, involved in erythroid differentiation and extramedullary erythropoiesis. This process gives rise to differentiation of stem cells to BFU-e/CFU-e, Pro E, and Baso E subpopulations. However, infection truncates progressing differentiation at the orthochromatic erythrocytes level, as demonstrated by scRNAseq and flow cytometry. As such, these cells are unable to pass to the reticulocyte stage, resulting in reduced number of mature circulating RBCs and the occurrence of chronic anemia. The physiological consequence of these events is the prolonged poor delivery of oxygen to various tissues, triggering lactic acid acidosis and the catabolic breakdown of muscle tissue, reminiscent of the wasting syndrome that is characteristic for the lethal stage of animal trypanosomosis.

胞外单细胞锥虫感染可引发家畜与野生动物罹患致死性慢性消耗性疾病。本研究通过筛选10株在小鼠中表现出不同毒力水平的伊氏锥虫（Trypanosoma evansi）野外分离株，建立了一种感染时长可超过100天的实验疾病模型，该模型可模拟慢性动物锥虫病的主要病理特征。在该模型中，尽管虫血症（parasitemia）水平得到良好控制，但感染仍以严重的锥虫病相关病理损伤为核心特征。研究人员对该病理特征开展深入的单细胞RNA测序（scRNA-seq）分析，揭示了脾脏巨噬细胞活化状态的复杂性，并凸显了组织驻留巨噬细胞（tissue resident macrophages, TRMs）在调控脾脏髓外红细胞生成过程中的关键作用。上述新数据表明，在实验性锥虫病研究领域中，巨噬细胞活化特征此前被过度简化为M1与M2两极化范式。值得注意的是，组织驻留巨噬细胞（TRMs）对红系细胞具有双向调控作用：一方面，这类细胞可表达与红细胞吞噬相关的特征基因谱；另一方面，其可高表达血管细胞黏附分子1（VCAM1），该分子已知可介导其与造血干祖细胞（hematopoietic stem and progenitor cells, HSPCs）的相互作用。在慢性感染过程中，造血干祖细胞（HSPCs）的Klf1、E2f8及Gfi1b基因表达会上调，这些基因参与红系分化与髓外红细胞生成过程。该过程可促使干细胞分化为爆式红系集落形成单位/红细胞集落形成单位（BFU-e/CFU-e）、早幼红细胞（Pro E）及嗜碱性幼红细胞（Baso E）亚群。然而，如单细胞RNA测序（scRNA-seq）与流式细胞术（flow cytometry）结果所示，感染会将红系分化进程阻滞于晚幼红细胞阶段。此类细胞无法进一步分化为网织红细胞，最终导致循环成熟红细胞数量减少，并引发慢性贫血。上述事件引发的生理结局为：各组织长期供氧不足，进而诱发乳酸酸中毒与肌肉组织分解代谢性降解，这与动物锥虫病致死阶段典型的消耗综合征特征相符。