Department of Physiology and Pharmacology, Western University, London, Ontario, Canada.
Molecular Medicine Research Laboratories, Krembil Centre for Stem Cell Biology, Robarts Research Institute, London, Ontario, Canada.
Stem Cells. 2018 May;36(5):723-736. doi: 10.1002/stem.2790. Epub 2018 Feb 12.
Blood-derived progenitor cell transplantation holds potential for the treatment of severe vascular diseases. Human umbilical cord blood (UCB)-derived hematopoietic progenitor cells purified using high aldehyde dehydrogenase (ALDH ) activity demonstrate pro-angiogenic functions following intramuscular (i.m.) transplantation into immunodeficient mice with hind-limb ischemia. Unfortunately, UCB ALDH cells are rare and prolonged ex vivo expansion leads to loss of high ALDH-activity and diminished vascular regenerative function. ALDH-activity generates retinoic acid, a potent driver of hematopoietic differentiation, creating a paradoxical challenge to expand UCB ALDH cells while limiting differentiation and retaining pro-angiogenic functions. We investigated whether inhibition of ALDH-activity during ex vivo expansion of UCB ALDH cells would prevent differentiation and expand progeny that retained pro-angiogenic functions after transplantation into non-obese diabetic/severe combined immunodeficient mice with femoral artery ligation-induced unilateral hind-limb ischemia. Human UCB ALDH cells were cultured under serum-free conditions for 9 days, with or without the reversible ALDH-inhibitor, diethylaminobenzaldehyde (DEAB). Although total cell numbers were increased >70-fold, the frequency of cells that retained ALDH /CD34+ phenotype was significantly diminished under basal conditions. In contrast, DEAB-inhibition increased total ALDH /CD34+ cell number by ≥10-fold, reduced differentiation marker (CD38) expression, and enhanced the retention of multipotent colony-forming cells in vitro. Proteomic analysis revealed that DEAB-treated cells upregulated anti-apoptotic protein expression and diminished production of proteins implicated with megakaryocyte differentiation. The i.m. transplantation of DEAB-treated cells into mice with hind-limb ischemia stimulated endothelial cell proliferation and augmented recovery of hind-limb perfusion. DEAB-inhibition of ALDH-activity delayed hematopoietic differentiation and expanded multipotent myeloid cells that accelerated vascular regeneration following i.m. transplantation in vivo. Stem Cells 2018;36:723-736.
血液来源的祖细胞移植在治疗严重血管疾病方面具有潜力。使用高醛脱氢酶（ALDH）活性从人脐带血（UCB）中纯化的造血祖细胞，在免疫缺陷小鼠后肢缺血的肌肉内（i.m.）移植后显示出促血管生成功能。不幸的是，UCB ALDH 细胞很少，体外延长扩增会导致高 ALDH 活性丧失和血管再生功能减弱。ALDH-活性产生视黄酸，这是造血分化的有力驱动因素，这给 UCB ALDH 细胞的扩增带来了一个悖论挑战，既要限制分化，又要保留促血管生成功能。我们研究了在 UCB ALDH 细胞的体外扩增过程中抑制 ALDH-活性是否会阻止分化，并在非肥胖型糖尿病/严重联合免疫缺陷小鼠中移植到股动脉结扎诱导的单侧后肢缺血模型后保留具有促血管生成功能的祖细胞。人 UCB ALDH 细胞在无血清条件下培养 9 天，有或没有可逆的 ALDH 抑制剂二乙氨基苯甲醛（DEAB）。尽管细胞总数增加了 70 多倍，但在基础条件下，保留 ALDH/CD34+表型的细胞频率显著降低。相比之下，DEAB 抑制增加了总 ALDH/CD34+细胞数≥10 倍，降低了分化标志物（CD38）的表达，并增强了体外多能集落形成细胞的保留。蛋白质组学分析表明，DEAB 处理的细胞上调了抗凋亡蛋白的表达，并减少了与巨核细胞分化相关的蛋白质的产生。DEAB 处理的细胞肌肉内移植到后肢缺血小鼠中，刺激了内皮细胞的增殖，并增强了后肢灌注的恢复。DEAB 抑制 ALDH-活性延迟了造血分化，并扩增了多能髓样细胞，这些细胞在体内肌肉内移植后加速了血管再生。干细胞 2018;36:723-736.
