Zhou Hao, Boys Alexander J, Harrod Jordan B, Bonassar Lawrence J, Estroff Lara A
Department of Materials Science and Engineering, Cornell University, Ithaca, New York 14853, United States.
Meinig School of Biomedical Engineering, Cornell University, Ithaca, New York 14853, United States.
Acta Biomater. 2020 Aug;112:274-285. doi: 10.1016/j.actbio.2020.05.032. Epub 2020 May 30.
Interfaces between soft tissue and bone are characterized by transitional gradients in composition and structure that mediate substantial changes in mechanical properties. For interfacial tissue engineering, scaffolds with mineral gradients have shown promise in controlling osteogenic behavior of seeded bone marrow stromal cells (bMSCs). Previously, we have demonstrated a 'top-down' method for creating monolithic bone-derived scaffolds with patterned mineral distributions similar to native tissue. In the present work, we evaluated the ability of these scaffolds to pattern osteogenic behavior in bMSCs in basic, osteogenic, and chondrogenic biochemical environments. Immunohistochemical (IHC) and histological stains were used to characterize cellular behavior as a function of local mineral content. Alkaline phosphatase, an early marker of osteogenesis, and osteocalcin, a late marker of osteogenesis, were positively correlated with mineral content in basic, osteogenic, and chondrogenic media. The difference in bMSC behavior between the mineralized and demineralized regions was most pronounced in an basic biochemical environment. In the mineralized regions of the scaffold, osteogenic markers were clearly present as early as 4 days in culture. In osteogenic media, osteogenic behavior was observed across the entire scaffold, whereas in chondrogenic media, there was an overall reduction in osteogenic biomarkers. Overall, these results indicate local mineral content of the scaffold plays a key role in spatially patterning bMSC behavior. Our results can be utilized for the development of interfacial tissue engineered scaffolds and understanding the role of local environment in determining bMSC behavior. STATEMENT OF SIGNIFICANCE: Soft tissue-to-bone interfaces, such as tendon-bone, ligament-bone, and cartilage-bone, are ubiquitous in mammalian musculoskeletal systems. These interfacial tissues have distinct, hierarchically-structured gradients of cellular, biochemical, and materials components. Given the complexity of the biological structures, interfacial tissues present unique challenges for tissue engineering. Here, we demonstrate that material-derived cues can spatially pattern osteogenic behavior in bone marrow stromal cells (bMSCs). Specifically, we observed that when the bMSCs are cultured on bone-derived scaffolds with mineral gradients, cells in contact with higher mineral content display osteogenic behavior at earlier times than those on the unmineralized substrate. The ability to pattern the cellular complexity found in native interfaces while maintaining biologically relevant structures is a key step towards creating engineered tissue interfaces.
软组织与骨之间的界面具有成分和结构上的过渡梯度，这些梯度介导了力学性能的显著变化。对于界面组织工程而言，具有矿物质梯度的支架在控制接种的骨髓基质细胞（bMSC）的成骨行为方面已显示出前景。此前，我们已经展示了一种“自上而下”的方法，用于创建具有与天然组织相似的图案化矿物质分布的整体骨衍生支架。在当前工作中，我们评估了这些支架在基础、成骨和软骨生成生化环境中对bMSC成骨行为进行图案化的能力。免疫组织化学（IHC）和组织学染色被用于将细胞行为表征为局部矿物质含量的函数。碱性磷酸酶是成骨的早期标志物，骨钙素是成骨的晚期标志物，它们在基础、成骨和软骨生成培养基中与矿物质含量呈正相关。bMSC在矿化区域和脱矿区域之间的行为差异在基础生化环境中最为明显。在支架的矿化区域，早在培养4天时就明显出现了成骨标志物。在成骨培养基中，在整个支架上都观察到了成骨行为，而在软骨生成培养基中，成骨生物标志物总体上有所减少。总体而言，这些结果表明支架的局部矿物质含量在空间上塑造bMSC行为方面起着关键作用。我们的结果可用于开发界面组织工程支架，并理解局部环境在决定bMSC行为中的作用。
软组织与骨的界面，如肌腱 - 骨、韧带 - 骨和软骨 - 骨，在哺乳动物肌肉骨骼系统中无处不在。这些界面组织具有独特的、层次结构的细胞、生化和材料成分梯度。鉴于生物结构的复杂性，界面组织给组织工程带来了独特的挑战。在这里，我们证明材料衍生的线索可以在空间上塑造骨髓基质细胞（bMSC）的成骨行为。具体而言，我们观察到当bMSC在具有矿物质梯度的骨衍生支架上培养时，与较高矿物质含量接触的细胞比在未矿化基质上的细胞更早表现出成骨行为。在保持生物学相关结构的同时，对天然界面中发现的细胞复杂性进行图案化的能力是创建工程化组织界面的关键一步。
