Zhang Xin, Ma Yan-Ni, Zhang Jun-Wu
National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, The Chinese Academy of Medical Sciences, Beijing, Republic of China.
Biol Res. 2007;40(1):41-53. doi: 10.4067/s0716-97602007000100005. Epub 2007 Jul 19.
In vitro suspension culture procedures for erythroid progenitor cells make it possible for us to obtain large cultures of erythrocyte populations for the investigation of globin gene switching. In this study we aimed to establish optimized culture systems for neonatal and adult erythroblasts and to explore the globin expression patterns in these culture systems. To culture CD34+ cells purified from human umbilical cord blood (CB) and adult bone marrow (BM), we respectively replaced the fetal bovine serum (FBS) with human cord serum and human adult serum. These CD34+ cells were then induced to erythroid differentiation. All the globin mRNA (including alpha-, zeta-, beta-, gamma-and epsilon-globin), the hemoglobin (Hb)-producing erythroid cells and the cellular distribution of fetal hemoglobin (Hb F) were identified during the culture process. The results showed that the globin expression pattern during erythroid differentiation in our culture systems closely recapitulated neonatal and adult patterns of globin expression in vivo, suggesting that our specially optimized culture systems not only overcame the higher Hb F levels in the BM-derived CD34+ culture in FBS-containing medium but also eliminated the disadvantages of low cell proliferation rate and low globin mRNA levels in serum-free medium.
红系祖细胞的体外悬浮培养程序使我们能够获得大量红细胞群体培养物,用于研究珠蛋白基因转换。在本研究中,我们旨在建立针对新生儿和成体成红细胞的优化培养系统,并探索这些培养系统中的珠蛋白表达模式。为了培养从人脐带血(CB)和成人骨髓(BM)中纯化的CD34+细胞,我们分别用人脐带血清和成人血清替代胎牛血清(FBS)。然后将这些CD34+细胞诱导进行红系分化。在培养过程中鉴定了所有珠蛋白mRNA(包括α-、ζ-、β-、γ-和ε-珠蛋白)、产生血红蛋白(Hb)的红系细胞以及胎儿血红蛋白(Hb F)的细胞分布。结果表明,我们培养系统中红系分化过程中的珠蛋白表达模式与体内新生儿和成体珠蛋白表达模式密切相似,这表明我们经过特殊优化的培养系统不仅克服了含FBS培养基中BM来源的CD34+培养物中较高的Hb F水平,还消除了无血清培养基中细胞增殖率低和珠蛋白mRNA水平低的缺点。