Developmental and Stem Cell Biology Program, Hospital for Sick Children, Toronto, ON, Canada.
The Institute of Medical Sciences, The University of Toronto, Toronto, ON, Canada.
Front Immunol. 2021 Oct 28;12:748519. doi: 10.3389/fimmu.2021.748519. eCollection 2021.
Inherited defects that abrogate the function of the adenosine deaminase (ADA) enzyme and consequently lead to the accumulation of toxic purine metabolites cause profound lymphopenia and severe combined immune deficiency. Additionally, neutropenia and impaired neutrophil function have been reported among ADA-deficient patients. However, due to the rarity of the disorder, the neutrophil developmental abnormalities and the mechanisms contributing to them have not been characterized. Induced pluripotent stem cells (iPSC) generated from two unrelated ADA-deficient patients and from healthy controls were differentiated through embryoid bodies into neutrophils. ADA deficiency led to a significant reduction in the number of all early multipotent hematopoietic progenitors. At later stages of differentiation, ADA deficiency impeded the formation of granulocyte colonies in methylcellulose cultures, leading to a significant decrease in the number of neutrophils generated from ADA-deficient iPSCs. The viability and apoptosis of ADA-deficient neutrophils isolated from methylcellulose cultures were unaffected, suggesting that the abnormal purine homeostasis in this condition interferes with differentiation or proliferation. Additionally, there was a significant increase in the percentage of hyperlobular ADA-deficient neutrophils, and these neutrophils demonstrated significantly reduced ability to phagocytize fluorescent microspheres. Supplementing iPSCs and methylcellulose cultures with exogenous ADA, which can correct adenosine metabolism, reversed all abnormalities, cementing the critical role of ADA in neutrophil development. Moreover, chemical inhibition of the ribonucleotide reductase (RNR) enzyme, using hydroxyurea or a combination of nicotinamide and trichostatin A in iPSCs from healthy controls, led to abnormal neutrophil differentiation similar to that observed in ADA deficiency, implicating RNR inhibition as a potential mechanism for the neutrophil abnormalities. In conclusion, the findings presented here demonstrate the important role of ADA in the development and function of neutrophils while clarifying the mechanisms responsible for the neutrophil abnormalities in ADA-deficient patients.
腺苷脱氨酶 (ADA) 酶功能缺失的遗传缺陷会导致有毒嘌呤代谢物的积累，从而导致严重的联合免疫缺陷和深刻的淋巴细胞减少症。此外，ADA 缺乏症患者还存在中性粒细胞减少和中性粒细胞功能受损的情况。然而，由于这种疾病的罕见性，中性粒细胞发育异常及其相关机制尚未得到明确。从两名无关的 ADA 缺乏症患者和健康对照者中生成诱导多能干细胞 (iPSC)，并通过胚状体分化为中性粒细胞。ADA 缺乏导致所有早期多能造血祖细胞数量显著减少。在分化的后期阶段，ADA 缺乏会阻碍甲基纤维素培养物中粒细胞集落的形成，从而导致从 ADA 缺乏的 iPSC 生成的中性粒细胞数量显著减少。从甲基纤维素培养物中分离出的 ADA 缺乏中性粒细胞的活力和凋亡不受影响，这表明该条件下异常的嘌呤稳态会干扰分化或增殖。此外，ADA 缺乏的中性粒细胞中出现显著增多的巨球形中性粒细胞，并且这些中性粒细胞吞噬荧光微球的能力明显降低。向 iPSC 和甲基纤维素培养物补充外源性 ADA（可以纠正腺苷代谢）可逆转所有异常，这进一步证实了 ADA 在中性粒细胞发育中的关键作用。此外，用羟基脲或烟酰胺和曲古抑菌素 A 的组合在健康对照者的 iPSC 中化学抑制核昔酸还原酶 (RNR) 酶，会导致类似于 ADA 缺乏症中观察到的异常中性粒细胞分化，这表明 RNR 抑制可能是中性粒细胞异常的潜在机制。总之，本研究结果表明 ADA 在中性粒细胞发育和功能中的重要作用，同时阐明了 ADA 缺乏症患者中性粒细胞异常的相关机制。
