Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands.
Tissue Eng Part A. 2012 Oct;18(19-20):1957-66. doi: 10.1089/ten.TEA.2011.0657. Epub 2012 Jun 25.
Within the field of bone tissue engineering, the endochondral approach to forming bone substitutes represents a novel concept, where cartilage will undergo hypertrophic differentiation before its conversion into bone. For this purpose, clinically relevant multipotent stromal cells (MSCs), MSCs, can be differentiated into the chondrogenic lineage before stimulating hypertrophy. Controversy exists in literature on the oxygen tensions naturally present during this transition in, for example, the growth plate. Therefore, the present study focused on the effects of different oxygen tensions on the progression of the hypertrophic differentiation of MSCs. Bone marrow-derived MSCs of four human donors were expanded, and differentiation was induced in aggregate cultures. Normoxic (20% oxygen) and hypoxic (5%) conditions were imposed on the cultures in chondrogenic or hypertrophic differentiation media. After 4 weeks, the cultures were histologically examined and by real-time polymerase chain reaction. Morphological assessment showed the chondrogenic differentiation of cultures from all donors under normoxic chondrogenic conditions. In addition, hypertrophic differentiation was observed in cultures derived from all but one donor. The deposition of collagen type X was evidenced in both chondrogenically and hypertrophically stimulated cultures. However, mineralization was exclusively observed in hypertrophically stimulated, normoxic cultures. Overall, the progression of hypertrophy was delayed in hypoxic compared with normoxic groups. The observed delay was supported by the gene expression patterns, especially showing the up-regulation of the late hypertrophic markers osteopontin and osteocalcin under normoxic hypertrophic conditions. Concluding, normoxic conditions are more beneficial for hypertrophic differentiation of MSCs than are hypoxic conditions, as long as the MSCs possess hypertrophic potential. This finding has implications for cartilage tissue engineering as well as for endochondral bone tissue engineering, as these approaches deal with, respectively, the inhibition or enhancement of hypertrophic chondrogenesis.
在骨组织工程领域，形成骨替代物的软骨内骨生成方法代表了一种新的概念，其中软骨在转化为骨之前会经历肥大分化。为此，可以将临床相关的多能基质细胞 (MSCs) 分化为软骨细胞谱系，然后再刺激肥大。在生长板等部位，文献中对这种转变过程中自然存在的氧张力存在争议。因此，本研究重点研究了不同氧张力对 MSCs 肥大分化进展的影响。从四名人类供体的骨髓中扩增 MSCs，并在聚集培养物中诱导分化。在软骨生成或肥大分化培养基中，对培养物施加常氧（20%氧气）和低氧（5%）条件。4 周后，对培养物进行组织学检查和实时聚合酶链反应。形态学评估显示，所有供体的培养物在常氧软骨生成条件下均进行了软骨分化。此外，除了一名供体之外，所有供体的培养物都观察到了肥大分化。在软骨生成和肥大刺激的培养物中都观察到了胶原 X 型的沉积。然而，仅在肥大刺激的常氧培养物中观察到了矿化。总的来说，与常氧组相比，低氧组的肥大进展延迟。基因表达模式支持了这种观察结果，特别是在常氧肥大条件下，晚期肥大标志物骨桥蛋白和骨钙素的上调。总之，只要 MSCs 具有肥大潜力，常氧条件比低氧条件更有利于 MSCs 的肥大分化。这一发现对软骨组织工程以及软骨内骨组织工程具有重要意义，因为这些方法分别涉及抑制或增强肥大性软骨生成。
