Department of Biomedical Engineering, University of California, Davis, Davis, California 95616, USA.
J Cell Biochem. 2010 May;110(1):87-96. doi: 10.1002/jcb.22514.
Skeletal defects commonly suffer from poor oxygen microenvironments resulting from compromised vascularization associated with injury or disease. Adipose stem cells (ASCs) represent a promising cell population for stimulating skeletal repair by differentiating toward the osteogenic lineage or by secreting trophic factors. However, the osteogenic or trophic response of ASCs to reduced oxygen microenvironments is poorly understood. Moreover, a direct comparison between 2D and 3D response of ASCs to hypoxia is lacking. Thus, we characterized the osteogenic and angiogenic potential of human ASCs under hypoxic (1%), normoxic (5%), and atmospheric (21%) oxygen tensions in both 2D and 3D over 4 weeks in culture. We detected greatest alkaline phosphatase activity and extracellular calcium deposition in cells cultured in both 2D and 3D under 21% oxygen, and reductions in enzyme activity corresponded to reductions in oxygen tension. ASCs cultured in 1% oxygen secreted more vascular endothelial growth factor (VEGF) over the 4-week period than cells cultured in other conditions, with cells cultured in 2D secreting VEGF in a more sustained manner than those in 3D. Expression of osteogenic markers revealed temporal changes under different oxygen conditions with peak expression occurring earlier in 3D. In addition, the increase of most osteogenic markers was significantly higher in 2D compared to 3D cultures at 1% and 5% oxygen. These results suggest that oxygen, in conjunction with dimensionality, affects the timing of the differentiation program in ASCs. These findings offer new insights for the use of ASCs in bone repair while emphasizing the importance of the culture microenvironment.
骨骼缺陷通常由于与损伤或疾病相关的血管化受损而导致的缺氧微环境而受到影响。脂肪干细胞 (ASCs) 通过向成骨谱系分化或分泌营养因子，代表了一种有前途的细胞群体，可刺激骨骼修复。然而，ASCs 对低氧微环境的成骨或营养反应知之甚少。此外，缺乏 ASC 对缺氧的 2D 和 3D 反应的直接比较。因此，我们在 2D 和 3D 培养物中，在 4 周的时间内，在 1%（缺氧）、5%（常氧）和 21%（大气）氧分压下，研究了人 ASC 在低氧（1%）、常氧（5%）和大气（21%）条件下的成骨和成血管潜能。我们发现，在 21%氧分压下，2D 和 3D 培养的细胞中碱性磷酸酶活性和细胞外钙沉积最高，而酶活性的降低与氧分压的降低相对应。与其他条件下培养的细胞相比，在 1%氧分压下培养的 ASC 在 4 周的时间内分泌更多的血管内皮生长因子 (VEGF)，并且 2D 培养的细胞分泌 VEGF 的方式比 3D 培养的细胞更持久。不同氧条件下的成骨标志物表达随时间发生变化，3D 培养物中更早出现峰值表达。此外，在 1%和 5%氧条件下，2D 培养物中的大多数成骨标志物的增加显著高于 3D 培养物。这些结果表明，氧气与维度一起影响 ASC 分化程序的时间。这些发现为 ASC 在骨修复中的应用提供了新的见解，同时强调了培养微环境的重要性。
