Department of Molecular Craniofacial Embryology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
Department of Maxillofacial Surgery, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
PLoS One. 2023 Feb 2;18(2):e0281345. doi: 10.1371/journal.pone.0281345. eCollection 2023.
Engineered cartilage tissue from differentiated mesenchymal stem cells (MSCs) can generate bone in vivo through endochondral ossification (ECO). This ECO-mediated approach has the potential to circumvent the severe problems associated with conventional MSC-based bone tissue engineering techniques that lack mechanisms to induce angiogenesis. Hyaluronic acid (HA) is a key component in the cartilage extracellular matrix. However, the ECO-supporting properties of HA remain largely unclear. This study aimed to compare the ability of HA and collagen hydrogels to support in vitro differentiation of MSC-based hypertrophic cartilage tissues and to promote endochondral bone formation in vivo. Following the chondrogenic and hypertrophic differentiation in vitro, both HA and collagen constructs accumulated sulfated glycosaminoglycan (sGAG) and type 1, type II, and type X collagen. However, HA hydrogels exhibited a more uniform distribution of sGAG, type 1 collagen, type X collagen, and osteocalcin proteins; in addition, the cells embedded in the hydrogels had more rounded cell morphologies than those in the collagen constructs. At week 5 of in vitro culture, two to three constructs were implanted into a subcutaneous pocket in nude mice and harvested after 4 and 8 weeks. Both HA and collagen constructs promoted endochondral bone formation with vascularization and bone marrow development; however, the HA constructs fused to form integrated bone tissues and the bone marrow developed along the space between the two adhered grafts in all implanted pockets (n = 5). In the collagen constructs, the integration was observed in 40% of the pockets (n = 5). Microcomputer CT analysis revealed that the bone volume of HA constructs was larger than that of collagen constructs. In conclusion, compared to collagen hydrogels, HA hydrogels had superior potential to generate integrated bone with vascularization and bone marrow development. This study provides valuable insights for applying ECO-mediated bone tissue engineering approaches for the repair of critical-sized bone defects.
由分化的间充质干细胞 (MSCs) 制成的工程软骨组织可以通过软骨内骨化 (ECO) 在体内产生骨。这种 ECO 介导的方法有可能避免与缺乏诱导血管生成机制的传统基于 MSC 的骨组织工程技术相关的严重问题。透明质酸 (HA) 是软骨细胞外基质的关键组成部分。然而，HA 的 ECO 支持特性在很大程度上仍不清楚。本研究旨在比较 HA 和胶原水凝胶在体外支持 MSC 衍生的肥大软骨组织分化以及体内促进软骨内骨形成的能力。在体外软骨和成骨分化后，HA 和胶原构建体都积累了硫酸化糖胺聚糖 (sGAG) 和 I 型、II 型和 X 型胶原。然而，HA 水凝胶表现出更均匀的 sGAG、I 型胶原、X 型胶原和骨钙蛋白蛋白的分布；此外，嵌入水凝胶中的细胞比胶原构建体中的细胞具有更圆的细胞形态。在体外培养的第 5 周，将两到三个构建体植入裸鼠的皮下袋中，并在 4 周和 8 周后取出。HA 和胶原构建体均促进了血管化和骨髓发育的软骨内骨形成；然而，HA 构建体融合形成了整合的骨组织，骨髓在所有植入袋中的两个粘附移植物之间的空间中发育（n = 5）。在胶原构建体中，在 40%的口袋中观察到整合（n = 5）。微计算机断层扫描分析显示，HA 构建体的骨体积大于胶原构建体。总之，与胶原水凝胶相比，HA 水凝胶在产生具有血管化和骨髓发育的整合骨方面具有更大的潜力。本研究为应用 ECO 介导的骨组织工程方法修复大尺寸骨缺损提供了有价值的见解。
