在脊椎动物主动脉-性腺-中肾（aorta-gonad-mesonephros，AGM）区域，生血内皮细胞通过内皮-造血转化（endothelial-to-hematopoietic transition，EHT）过程产生造血干细胞（hematopoietic stem cells，HSCs）。EHT过程受到多种因素的精细调控。在本研究中，我们利用斑马鱼模型探索了影响HSC发育的关键因子及其调控机制。首先，我们发现瘦素（leptin）广泛分布于斑马鱼胚胎AGM区血管内皮细胞周围，介导leptin信号传递的瘦素受体（leptin receptor，LepR）编码基因高表达于肌肉细胞和AGM区血管内皮细胞中，提示leptin信号与胚胎期造血发育有关。随后，我们分别构建了leptin及LepR编码基因缺失的斑马鱼突变体，用于检测leptin信号在胚胎期造血发育过程中的作用。功能缺失及细胞特异性回救实验结果显示leptin信号以非细胞自主方式调控HSC产生。为了探究leptin信号影响HSC发育的内在机制，我们进行了转录组学、代谢组学以及相应功能实验。结果表明leptin信号缺失可抑制胚胎中脂肪酸氧化（fatty-acid oxidation，FAO）过程，导致体内能量生成减少，肌肉细胞和血管内皮细胞中脂质累积。激活AMPK信号通路能够降低leptin信号缺失胚胎中脂质水平并回救HSC产生缺陷表型。以上结果说明leptin信号通过AMPK信号通路介导的FAO调控胚胎期HSC产生。

另一方面，我们在急性髓系白血病（acute myeloid leukemia，AML）患者的单细胞转录组数据中鉴定到一类以原始肿瘤细胞为特征的AML亚型，并发现转录因子Erf被高度激活。同时，我们发现该转录因子也高表达于斑马鱼胚胎期HSCs中。通过功能缺失及功能获得实验，我们证实Erf影响斑马鱼早期造血发育过程中HSC产生及分化。

综上，本研究首次揭示了脂质代谢调控因子leptin以及转录因子Erf在HSC发育中的重要调控作用。鉴定体内HSC发育的关键调控因子可为体外产生具有移植重建能力的HSCs提供理论指导，为优化血液疾病治疗策略提供思路。

In the aorta-gonad-mesonephros (AGM) region of vertebrates, hematopoietic stem cells (HSCs) arise from hemogenic endothelium through endothelial-to-hematopoietic transition (EHT), which is finely regulated by various factors. In this study, we investigated the key factors and mechanisms underlying HSC development in zebrafish.

First, we identified that leptin was widely distributed around the vascular endothelial cells in the AGM region of zebrafish embryos. The leptin receptor (LepR) gene was mainly expressed in muscle cells and vascular endothelial cells, indicating that leptin signaling was related to hematopoiesis. Subsequently, we generated lepa and lepr zebrafish mutants to investigate the role of leptin signaling in hematopoiesis. The results of loss-of-function and cell type-specific rescue experiments showed that leptin signaling regulated HSC generation in a non-cell autonomous manner. To investigate the underlying mechanism of leptin signaling in this process, we performed transcriptomics, metabolomics and corresponding functional analysis. The results showed that loss of leptin signaling inhibited fatty acid oxidation (FAO) in the embryos, leading to the disruption of energy generation and lipid accumulation in muscle cells and endothelial cells. Activating AMPK signaling pathway reduced the lipid level and rescued the hematopoietic defects in leptin-deficient embryos. These results indicated that leptin-AMPK-FAO axis regulated HSC generation.

Second, based on the single-cell transcriptome data from acute myeloid leukemia (AML) patients, we identified a subtype of AML characterized by malignant primitive cells and found an activated transcription factor Erf. erf was also highly expressed in HSCs in zebrafish embryos. The results of knockdown and overexpression experiments demonstrated that Erf affected HSC generation and differentiation during hematopoiesis.

In conclusion, our study revealed the important roles of the lipid metabolism regulatory factor leptin and transcription factor Erf in HSC development. Identification of the key regulatory factors for HSC development in vivo could provide theoretical guidance for producing functional HSCs in vitro and optimize the treatment strategies for hematological diseases.