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首页> 美国卫生研究院文献>Biomedical Optics Express >Three-dimensional characterization of collagen remodeling in cell-seeded collagen scaffolds via polarization second harmonic generation
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Three-dimensional characterization of collagen remodeling in cell-seeded collagen scaffolds via polarization second harmonic generation

机译：通过极化第二谐波产生细胞播种胶原支架中胶原重塑的三维表征

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In this study, we use non-linear imaging microscopy to characterize the structural properties of porous collagen-GAG scaffolds (CGS) seeded with human umbilical vein endothelial cells (HUVECs), as well as human mesenchymal stem cells (hMSCs), a co-culture previously reported to form vessel-like structures inside CGS. The evolution of the resulting tissue construct was monitored over 10 days via simultaneous two- and three-photon excited fluorescence microscopy. Time-lapsed 2- and 3-photon excited fluorescence imaging was utilized to monitor the temporal evolution of the vascular-like structures up to 100 µm inside the scaffold up to 10 days post-seeding. 3D polarization-dependent second harmonic generation (PSHG) was utilized to monitor collagen-based scaffold remodeling and determine collagen fibril orientation up to 200 µm inside the scaffold. We demonstrate that polarization-dependent second harmonic generation can provide a novel way to quantify the reorganization of the collagen architecture in CGS simultaneously with key biomechanical interactions between seeded cells and CGS that regulate the formation of vessel-like structures inside 3D tissue constructs. A comparison between samples at different days in vitro revealed that gradually, the scaffolds developed an orthogonal net-like architecture, previously found in real skin.
机译：在该研究中，我们使用非线性成像显微镜表征与人脐静脉内皮细胞（HUVECS）和人间充质干细胞（HMSCs）播种的多孔胶原-GAG支架（CGS）的结构性质。以前报道的培养物在CGS内形成血管样结构。通过同时的两℃和三光子激发荧光显微镜进行10天监测所得组织构建体的进化。利用时间失效的2-和3-光子激发荧光成像，以监测血管状结构的时间演变在接种后10天内高达100μm的血管状结构。 3D偏振依赖性的二次谐波产生（PSHG）用于监测基于胶原基的支架重塑，并确定支架内高达200μm的胶原型原纤维取向。我们证明，偏振依赖性的二次谐波产生可以提供一种新的方法，可以同时通过调节3D组织构建体内的血管样结构的颈部和CGS之间的关键生物力学相互作用来量化CGS中的胶原型架构的重组。在体外不同日子的样品之间的比较显示，逐渐地，支架开发出一个正交的网状架构，以前在真正的皮肤中发现。

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