Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, Philadelphia, PA.
Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine and Children's Hospital of Philadelphia, Philadelphia, PA.
Blood Adv. 2024 Mar 26;8(6):1449-1463. doi: 10.1182/bloodadvances.2023011708.
During development, erythroid cells are produced through at least 2 distinct hematopoietic waves (primitive and definitive), generating erythroblasts with different functional characteristics. Human induced pluripotent stem cells (iPSCs) can be used as a model platform to study the development of red blood cells (RBCs) with many of the differentiation protocols after the primitive wave of hematopoiesis. Recent advances have established that definitive hematopoietic progenitors can be generated from iPSCs, creating a unique situation for comparing primitive and definitive erythrocytes derived from cell sources of identical genetic background. We generated iPSCs from healthy fetal liver (FL) cells and produced isogenic primitive or definitive RBCs which were compared directly to the FL-derived RBCs. Functional assays confirmed differences between the 2 programs, with primitive RBCs showing a reduced proliferation potential, larger cell size, lack of Duffy RBC antigen expression, and higher expression of embryonic globins. Transcriptome profiling by scRNA-seq demonstrated high similarity between FL- and iPSC-derived definitive RBCs along with very different gene expression and regulatory network patterns for primitive RBCs. In addition, iPSC lines harboring a known pathogenic mutation in the erythroid master regulator KLF1 demonstrated phenotypic changes specific to definitive RBCs. Our studies provide new insights into differences between primitive and definitive erythropoiesis and highlight the importance of ontology when using iPSCs to model genetic hematologic diseases. Beyond disease modeling, the similarity between FL- and iPSC-derived definitive RBCs expands potential applications of definitive RBCs for diagnostic and transfusion products.
在发育过程中，红细胞通过至少 2 个不同的造血波（原始和定型）产生，产生具有不同功能特征的成红细胞。人类诱导多能干细胞（iPSC）可作为研究红细胞（RBC）发育的模型平台，许多分化方案都在造血原始波之后进行。最近的进展确立了可以从 iPSC 中产生定型造血祖细胞，为比较来自相同遗传背景的细胞来源的原始和定型红细胞创造了独特的情况。我们从健康胎儿肝脏（FL）细胞中生成 iPSC，并产生同基因的原始或定型 RBC，直接与 FL 衍生的 RBC 进行比较。功能测定证实了这两个方案之间的差异，原始 RBC 表现出增殖潜力降低、细胞体积增大、缺乏 Duffy RBC 抗原表达以及胚胎球蛋白表达升高。单细胞 RNA-seq 的转录组分析表明，FL 和 iPSC 衍生的定型 RBC 之间具有高度相似性，而原始 RBC 的基因表达和调控网络模式则非常不同。此外，携带红细胞主调控因子 KLF1 已知致病性突变的 iPSC 系表现出特定于定型 RBC 的表型变化。我们的研究提供了关于原始和定型红细胞生成之间差异的新见解，并强调了在使用 iPSC 对遗传血液疾病进行建模时本体论的重要性。除了疾病建模之外，FL 和 iPSC 衍生的定型 RBC 之间的相似性扩展了定型 RBC 在诊断和输血产品中的潜在应用。
