A comparative study of two routinely used protocols for ex vivo erythroid differentiation

IF 2.1 4区 医学 Q3 HEMATOLOGY Blood Cells Molecules and Diseases Pub Date : 2024-01-23 DOI:10.1016/j.bcmd.2024.102829
Auria Godard , Robert Seute , Alexandra Grimaldi , Thomas Granier , Jacques Chiaroni , Wassim El Nemer , Maria De Grandis
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Abstract

Background

Erythropoiesis is a complex developmental process in which a hematopoietic stem cell undergoes serial divisions and differentiates through well-defined stages to give rise to red blood cells. Over the last decades, several protocols have been developed to perform ex vivo erythroid differentiation, allowing investigation into erythropoiesis and red cell production in health and disease.

Results

In the current study, we compared the two commonly used protocols by assessing the differentiation kinetics, synchronisation, and cellular yield, using molecular and cellular approaches. Peripheral blood CD34+ cells were cultured in a two-phase (2P) or a four-phase (4P) liquid culture (LC) and monitored for 20 days. Both protocols could recapitulate all stages of erythropoiesis and generate reticulocytes, although to different extents. Higher proliferation and viability rates were achieved in the 4P-LC, with a higher degree of terminal differentiation and enucleation, associated with higher levels of the erythroid-specific transcription factors GATA-1, KLF-1, and TAL-1. Although the 2P-LC protocol was less efficient regarding terminal erythroid differentiation and maturation, it showed a higher yield of erythroid progenitors in the erythropoietin (EPO)-free expansion phase.

Conclusions

We provide data supporting the use of one protocol or the other to study the biological processes occurring in the early or late stages of erythroid differentiation, depending on the physiological process or pathological defect under investigation in a given study.

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对两种常规使用的体外红细胞分化方案进行比较研究。
背景:红细胞生成是一个复杂的发育过程,在这一过程中,造血干细胞经过连续分裂和明确阶段的分化生成红细胞。在过去的几十年里,已经开发出了几种进行体外红细胞分化的方案,从而可以研究健康和疾病中的红细胞生成和红细胞生产:在本研究中,我们使用分子和细胞方法评估了两种常用方案的分化动力学、同步性和细胞产量,并对其进行了比较。外周血 CD34+ 细胞在两相(2P)或四相(4P)液体培养液(LC)中培养,并监测 20 天。两种方案都能重现红细胞生成的各个阶段并生成网状细胞,只是程度不同。4P-LC 的增殖率和存活率更高,末期分化和去核程度更高,红细胞特异性转录因子 GATA-1、KLF-1 和 TAL-1 的水平也更高。虽然2P-LC方案在红细胞终末分化和成熟方面效率较低,但它在无促红细胞生成素(EPO)扩增阶段显示出更高的红细胞祖细胞产量:我们提供的数据支持使用一种或另一种方案来研究红细胞分化早期或晚期发生的生物过程,这取决于特定研究中调查的生理过程或病理缺陷。
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来源期刊
CiteScore
4.90
自引率
0.00%
发文量
42
审稿时长
14 days
期刊介绍: Blood Cells, Molecules & Diseases emphasizes not only blood cells, but also covers the molecular basis of hematologic disease and studies of the diseases themselves. This is an invaluable resource to all those interested in the study of hematology, cell biology, immunology, and human genetics.
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