Apoptosis and hematopoiesis in murine fetal liver.
Apoptosis and hematopoiesis in murine fetal liver.
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DOI：
10.1182/blood.v81.2.373.bloodjournal812373
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发表时间：
1993-01
期刊：
影响因子：
20.3
通讯作者：
Hong;Brad Bauer;Gretchen;Lipke;Robert L. Phillips;G. Zant
中科院分区：
文献类型：
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作者：
Hong;Brad Bauer;Gretchen;Lipke;Robert L. Phillips;G. Zant
胎鼠肝脏（FL）是一个器官的强烈，但短暂的，造血活动在妊娠中期，与红细胞生成是主要在第11至16天。因此，似乎合理的预期，造血细胞因子，如促红细胞生成素（epo），白细胞介素-3（IL-3），和干细胞因子（SCF），可能在维持红细胞生成和细胞凋亡在个体发育过程中在肝脏的稳态平衡中发挥重要作用。首先，我们通过测量培养的FL的集落形成和血红蛋白合成来确定这些生长因子对造血的影响。其次，我们确定了这些细胞因子所提供的保护凋亡，使用DNA电泳分析和流式细胞术的FL细胞剥夺了epo，IL-3和SCF的培养。促红细胞生成素是必要的，单独足以血红蛋白合成的集落形成单位红细胞集落，但IL-3是一个必要的辅因子，以获得最大的发展爆发形成单位红细胞集落。SCF单独在FL的甲基纤维素培养物中引起很少的集落形成，但当与epo和IL-3组合时，它对集落的数量和它们的大小都有显着的影响。其次，通过测量凋亡细胞中内源性核酸酶活性引起的DNA片段化来确定凋亡指数。来自无细胞因子培养物的肝细胞显示DNA广泛降解为低分子量核小体寡聚体，这是凋亡的特征。EPO对细胞凋亡的保护作用与不同胎龄胎儿的红细胞生成水平直接相关。促红细胞生成素是迄今为止最关键的细胞因子在保存FL细胞凋亡。琼脂糖凝胶分析表明，SCF和IL-3单独在减少片段中的DNA量方面没有明显的效果，当与epo组合时，它们没有比单独epo提供的保护效果更好。然而，使用更灵敏的流式细胞仪测定细胞亚二倍体量的DNA，SCF，IL-3单独具有可测量的保护作用，这是小于由EPO引起的。因此，我们表明，正常的，未转化的造血系统的发展中的细胞不仅需要细胞因子的增殖和分化，但他们有一个初始的和绝对的要求，他们从细胞凋亡的保护。
The fetal mouse liver (FL) is an organ of intense, but transient, hematopoietic activity during mid-gestation, with erythropoiesis being predominant during days 11 through 16. It therefore seemed reasonable to expect that hematopoietic cytokines, such as erythropoietin (epo), interleukin-3 (IL-3), and stem cell factor (SCF), may play important roles in maintaining a homeostatic balance of erythropoiesis and apoptosis in liver during ontogeny. First, we determined the effects of these growth factors on hematopoiesis by measuring colony formation and hemoglobin synthesis of cultured FLs. Secondly, we determined the protection from apoptosis afforded by these cytokines, using electrophoretic analysis of DNA and by flow cytometry of FL cells deprived in culture of epo, IL-3, and SCF. Erythropoietin was necessary and alone sufficient for hemoglobin synthesis in colony-forming units-erythroid colonies, but IL-3 was a required cofactor to obtain maximal development of burst-forming units-erythroid colonies. SCF alone caused little colony formation in methylcellulose cultures of FLs, but when combined with epo and IL-3, it had dramatic effects both on the number of colonies and their size. Secondly, indices of apoptosis were determined by measuring DNA fragmentation caused by endogenous nuclease activity in apoptotic cells. Liver cells from cultures without cytokines showed the extensive degradation of DNA to low molecular weight nucleosomal oligomers, which is characteristic of apoptosis. Protection from apoptosis afforded by epo directly corresponded to the level of erythropoiesis in FLs of different gestational age. Erythropoietin was by far the most critical cytokine in sparing FL cells from apoptosis. Analyses of agarose gels showed that SCF and IL-3 alone had no apparent effect in reducing the amount of DNA in fragments, and when combined with epo they had no more protective effect than that provided by epo alone. However, using the more sensitive flow cytometric determination of cells with subdiploid amounts of DNA, SCF, and IL-3 alone had measurable protective effects that were less than those caused by epo. Thus, we show that normal, untransformed cells of the developing hematopoietic system not only require cytokines for proliferation and differentiation, but they have an initial and absolute requirement of them for protection from apoptosis.