Characterization of mesenchymal stem cells and fibrochondrocytes in three-dimensional co-culture: analysis of cell shape, matrix production, and mechanical performance.
Characterization of mesenchymal stem cells and fibrochondrocytes in three-dimensional co-culture: analysis of cell shape, matrix production, and mechanical performance.
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DOI：
10.1186/s13287-016-0301-8
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发表时间：
2016-03-12
影响因子：
7.5
通讯作者：
Bonassar LJ
中科院分区：
文献类型：
--
作者：
McCorry MC;Puetzer JL;Bonassar LJ
骨髓间充质干细胞（MSC）对半月板再生和修复显示出积极的治疗作用。初步体外工作表明 MSC 在半月板组织工程中的应用取得了积极的结果；然而，需要更多关于如何指导 MSC 行为的信息。本研究的目的是检查 MSC 与原代半月板纤维软骨细胞 (FCC) 在纤维软骨形成介质中的三维胶原支架中共培养的效果。假设 MSC 和 FCC 共培养可促进 MSC 向 FCC 细胞表型的转变（通过基质分泌和形态测量）。 MSC 和 FCC 分别从牛骨髓和半月板中分离出来。将细胞以 0:100、25:75、50:50、75:25 和 100:0 的 MSC:FCC 比例接种在 20 mg/mL 高密度 I 型胶原凝胶中。将构建体培养长达 2 周，然后分析细胞形态、糖胺聚糖含量、胶原蛋白含量以及 I、II 和 X 型胶原蛋白的产生。细胞在整个支架中均匀混合，细胞之间的直接细胞接触有限。培养 2 周后，MSC 从纺锤状形态转变为圆形表型，而 FCC 在整个培养过程中保持圆形。尽管 MSC 形状随培养而变化，但在整个培养过程中，其总体尺寸明显大于 FCC。虽然 75:25 和 100:0（MSC 单一培养物）培养组比单一培养物中的 FCC 产生显着更多的糖胺聚糖 (GAG)/DNA，但在 50:50 共培养物中 GAG 保留率最高。同样，100:0 和 50:50 共培养物的总模量最高。两周后，所有样品均含有 I 型和 II 型胶原蛋白，X 型胶原蛋白表达仅在 MSC 单一培养物凝胶中明显。在单一培养和共培养中，MSC 都会转变为 FCC 形态。共培养减少了 MSC 的肥大，由 X 型胶原表明。这项研究表明 MSC 表型可以受到三维凝胶中间接均质细胞培养的影响，证明了 MSC 在半月板组织工程应用中的适用性。本文的在线版本 (doi:10.1186/s13287-016-0301-8) 包含补充材料，可供授权用户使用。
Bone marrow mesenchymal stem cells (MSCs) have shown positive therapeutic effects for meniscus regeneration and repair. Preliminary in vitro work has indicated positive results for MSC applications for meniscus tissue engineering; however, more information is needed on how to direct MSC behavior. The objective of this study was to examine the effect of MSC co-culture with primary meniscal fibrochondrocytes (FCCs) in a three-dimensional collagen scaffold in fibrochondrogenic media. Co-culture of MSCs and FCCs was hypothesized to facilitate the transition of MSCs to a FCC cell phenotype as measured by matrix secretion and morphology. MSCs and FCCs were isolated from bovine bone marrow and meniscus, respectively. Cells were seeded in a 20 mg/mL high-density type I collagen gel at MSC:FCC ratios of 0:100, 25:75, 50:50, 75:25, and 100:0. Constructs were cultured for up to 2 weeks and then analyzed for cell morphology, glycosaminoglycan content, collagen content, and production of collagen type I, II, and X. Cells were homogeneously mixed throughout the scaffold and cells had limited direct cell–cell contact. After 2 weeks in culture, MSCs transitioned from a spindle-like morphology toward a rounded phenotype, while FCCs remained rounded throughout culture. Although MSC shape changed with culture, the overall size was significantly larger than FCCs throughout culture. While 75:25 and 100:0 (MSC mono-culture) culture groups produced significantly more glycosaminoglycan (GAG)/DNA than FCCs in mono-culture, GAG retention was highest in 50:50 co-cultures. Similarly, the aggregate modulus was highest in 100:0 and 50:50 co-cultures. All samples contained both collagen types I and II after 2 weeks, and collagen type X expression was evident only in MSC mono-culture gels. MSCs shift to a FCC morphology in both mono- and co-culture. Co-culture reduced hypertrophy by MSCs, indicated by collagen type X. This study shows that MSC phenotype can be influenced by indirect homogeneous cell culture in a three-dimensional gel, demonstrating the applicability of MSCs in meniscus tissue engineering applications. The online version of this article (doi:10.1186/s13287-016-0301-8) contains supplementary material, which is available to authorized users.