Thrombosis I Laboratory, Hematological Research Institute Mariano R. Castex, National Academy of Medicine, CONICET, Buenos Aires, Argentina.
Transfusion. 2011 Aug;51(8):1784-95. doi: 10.1111/j.1537-2995.2010.03051.x. Epub 2011 Feb 18.
Transplanted hematopoietic progenitor cells (CD34+) have shown great promise in regenerative medicine. However, the therapeutic potential of transplanted cells is limited by their poor viability. It is well known that the microenvironment in which progenitors reside substantially affects their behavior. Because extracellular acidosis is a common feature of injured tissues or the tumor microenvironment and is a critical regulator of cell survival and activation, we evaluated the impact of acidosis on CD34+ cell biology.
Apoptosis was evaluated by fluorescence microscopy and binding of annexin V, hypodiploid cells, Bcl-xL expression, active caspase-3, and mitochondrial membrane potential was determined by flow cytometry. Colony-forming units were studied by clonogenic assays, and cell cycle was evaluated by flow cytometry.
Exposure of CD34+ cells to low pH (7.0-6.5) caused intracellular acidification, decreased cell proliferation, and triggered apoptosis via the mitochondrial pathway. Whereas exposure to thrombopoietin (TPO), stem cell factor (SCF), interleukin (IL)-3 or increases in cyclic adenosine monophosphate (cAMP) levels prevented CD34+ cell death induced by acidic conditions, granulocyte-macrophage-colony-stimulating factor, FMS-like tyrosine kinase 3-ligand, erythropoietin, and vascular endothelial growth factor had no effect. Despite their cytoprotective effect, CD34+ cell expansion triggered by TPO, SCF, or IL-3 was significantly impaired at low pH. However, a cocktail of these three cytokines synergistically supported proliferation, cell cycle progression, and colony formation.
Our findings indicate that an acidic milieu is deleterious for CD34+ cells and that a combination of certain cytokines and cAMP donors may improve cell viability and function. These data may be useful to develop new therapeutic strategies or to optimize protocols for regenerative medicine.
移植的造血祖细胞（CD34+）在再生医学中显示出巨大的潜力。然而，移植细胞的治疗潜力受到其存活率差的限制。众所周知，祖细胞所在的微环境对其行为有很大影响。由于细胞外酸中毒是损伤组织或肿瘤微环境的共同特征，也是细胞存活和激活的关键调节剂，因此我们评估了酸中毒对 CD34+细胞生物学的影响。
通过荧光显微镜和 Annexin V 结合评估细胞凋亡，通过流式细胞术检测低倍体细胞、Bcl-xL 表达、活性 caspase-3 和线粒体膜电位。通过集落形成单位研究克隆形成单位，通过流式细胞术评估细胞周期。
将 CD34+细胞暴露于低 pH（7.0-6.5）会导致细胞内酸化、细胞增殖减少，并通过线粒体途径触发细胞凋亡。而暴露于血小板生成素（TPO）、干细胞因子（SCF）、白细胞介素（IL）-3 或增加环腺苷酸（cAMP）水平可防止 CD34+细胞因酸性条件而死亡，但粒细胞-巨噬细胞集落刺激因子、FMS 样酪氨酸激酶 3 配体、促红细胞生成素和血管内皮生长因子则没有效果。尽管 TPO、SCF 或 IL-3 触发的 CD34+细胞扩张具有细胞保护作用，但在低 pH 条件下，其扩增明显受损。然而，这三种细胞因子的鸡尾酒协同支持增殖、细胞周期进程和集落形成。
我们的研究结果表明，酸性环境对 CD34+细胞有害，而某些细胞因子和 cAMP 供体的组合可能会提高细胞活力和功能。这些数据可能有助于开发新的治疗策略或优化再生医学的方案。
