基于小动物荧光成像研究生物材料体内降解的实验方法

刘秀芸; 陈丹藜; 关淇峰; 刘田; 杨丰合; 牛旭锋

doi:10.13801/j.cnki.fhclxb.20201215.009

基于小动物荧光成像研究生物材料体内降解的实验方法

doi: 10.13801/j.cnki.fhclxb.20201215.009

刘秀芸^{1, 2,},
陈丹藜^{1, 2,},
关淇峰^{1, 2,},
刘田^{1, 2,},
杨丰合^{1, 2,},
牛旭锋^{1, 2, ,}

1.
北京航空航天大学　生物与医学工程学院，北京 100083
2.
北京航空航天大学　生物医学工程高精尖创新中心，北京 100083

基金项目: 国家自然科学基金(11872097)；北京市自然科学基金-海淀原始创新联合基金(L182017)

详细信息

通讯作者:
牛旭锋，博士，教授，博士生导师，研究方向为生物材料与再生医学、生物力学与力学生物学　E-mail：nxf@buaa.edu.cn

中图分类号: R318
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- 被引次数: 0
出版历程
- 收稿日期: 2020-04-23
- 录用日期: 2020-06-02
- 网络出版日期: 2020-12-16
- 刊出日期: 2021-02-15

Evaluation method of biomaterial degradation based on in vivo imaging system

LIU Xiuyun^{1, 2
,},
CHEN Danli^{1, 2
,},
GUAN Qifeng^{1, 2
,},
LIU Tian^{1, 2
,},
YANG Fenghe^{1, 2
,},
NIU Xufeng^{1, 2
, ,}

1.
School of Biological Science and Medical Engineering, Beihang University, Beijing 100083, China
2.
Beijing Advanced Innovation Center for Biomedical Engineering, Beihang University, Beijing 100083, China

摘要

摘要: 可降解材料作为生物材料的重要组成部分，其体内降解性能的好坏往往决定着材料植入后的成败。因此，对材料体内降解的评价显得尤为重要。传统的生物材料体内降解评价方法需要在各取样点取出不同批次的降解样品，阻止了对同一个实验样品降解过程的连续测量，并且存在样品需求量大的问题。小动物活体成像系统(in vivo imaging system，IVIS)具有非侵入性、操作性强等特点，为解决上述问题提供了思路。本研究旨在建立一种利用小动物荧光成像系统检测可降解材料降解性能的方法，通过将近红外荧光染料经化学反应标记到可降解材料上，由荧光强度的变化反应材料的降解程度。体内降解实验表明此方法制得的荧光标记材料，荧光稳定性高，材料降解过程中荧光强度变化与质量损失拟合效果良好(R²=0.9994)。综上，该方法解决了测量材料降解样品量大的问题，并且提高了实验过程的连贯性。
- 可降解医用高分子材料 /
- 体内降解 /
- 小动物活体成像系统 /
- 荧光标记 /
- 评价方法
Abstract: Degradable material is an important part of biomaterials, its degradation performance in vivo has great influence on the final success after implantation. Thus, the method of evaluating the degradation of the materials in vivo is crucial to the evaluation of material performance. The traditional method to estimate the degradation in vivo needs to take out different batches of degradation samples at each sampling point, which prevents the continuous measurement of the degradation process and it needs a large number of samples. In vivo imaging system (IVIS) has the characteristics of non-invasive and strong operability, which provides a way to solve the problems above. In this study, we established a method to detect the degradation performance of degradable materials with IVIS. The research method was that the near-infrared fluorescent dye was labeled on the degradable materials by chemical reaction, and then using the change of fluorescence intensity to reflect the degradation degree of the materials. The in vivo degradation experimental results show that the fluorescent labeling materials prepared by this method have high fluorescence stability, and the fitting effect of fluorescence intensity and mass loss in the process of material degradation is good (R² = 0.9994). In conclusion, this method solves the problem of large amount of samples in the traditional method of measuring material degradation, and improves the continuity of the experimental process.
- degradable medical polymer materials /
- in vivo degradation /
- in vivo imaging system /
- fluorescence labeling /
- evaluation method

HTML全文

图 1 聚乳酸-羟基乙酸共聚物(PLGA)、PLGA-叔丁氧羰基(Boc)及PLGA-NH₂的FTIR图谱

Figure 1. FTIR spectra of poly(lactide-co-glycolide) acid (PLGA), PLGA-t-butyloxycarbonyl (Boc) and PLGA-NH₂

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图 2 荧光显微镜拍摄的PLGA-cy5.5多孔膜的明场图像与暗场图像的叠加

Figure 2. Superposition of bright field image and dark field image of PLGA-cy5.5 porous membrane by fluorescence microscope

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图 3 荧光显微镜拍摄的cy5.5标记的胶原蛋白海绵

Figure 3. cy5.5 labeled collagen sponge under the fluorescence microscope

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图 4 IVIS Lumina Ⅲ软件自动圈选感兴趣区域(ROI)

Figure 4. Automatically circle range of interest (ROI) in Living Image software

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图 5 根据植入的PLGA-cy5.5多孔膜的实际大小、位置手动圈选ROI

Figure 5. Manually circle ROI according to actual size and location of implanted PLGA-cy5.5 porous membrane

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图 6 分析cy5.5荧光标记的胶原蛋白海绵降解过程时使用相同大小与相同相对位置的选框圈定ROI

Figure 6. Using ROI of the same size and the same relative location to analyze the degradation process of the cy5.5 labeled collagen sponge

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图 7 根据降解过程中cy5.5荧光标记的胶原蛋白海绵的实际大小和位置圈定ROI

Figure 7. Using ROI defined by the cy5.5 labeled collagen sponge’s actual size and location to analyze the degradation process

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图 8 圈选小鼠无材料位置的皮肤作为背景荧光值(左侧)与植入cy5.5荧光标记的胶原蛋白海绵的荧光值相比较(右侧)

Figure 8. Skin without implanted material was selected as the background (left) and was compared to the fluorescent intensity of the implanted cy5.5 labeled collagen sponge (right)

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图 9 PLGA-cy5.5多孔膜在小鼠体内降解8周的荧光强度变化图

Figure 9. PLGA-cy5.5 porous membrane’s fluorescence intensity loss during 8 weeks degradation in rats

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图 10 PLGA-cy5.5多孔膜在小鼠体内降解8周的质量变化与荧光强度变化对比图

Figure 10. Comparison of the quality loss and fluorescence intensity loss of PLGA-cy5.5 porous membrane during 8 weeks degradation in rats

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