中国组织工程研究

中国组织工程研究 ›› 2024, Vol. 28 ›› Issue (22): 3451-3456.doi: 10.12307/2024.473

• 材料力学及表面改性 material mechanics and surface modification • 上一篇    下一篇

弹性蛋白对前交叉韧带力学响应影响的弹性蛋白酶定量分析

张闻天1，邓羽平2，3，4，刘晓云2，4，李彩娟5，王  勉2，6，梁泽宇2，熊  亮3，黄  刚3，陈广新7，李子涛8，9，黄文华1，2，3，6，10   

  1. 1福建医科大学基础医学院，福建省福州市  350122；2南方医科大学基础医学院人体解剖学国家重点学科，广东省医学生物力学重点实验室，广东省医学3D打印应用转化工程技术研究中心，广东省广州市  510515；3南方医科大学中西医结合医院骨伤科，广东省广州市  510315；4生物医学工程研究所深圳湾实验室，广东省深圳市  518132；牡丹江医学院附属红旗医院，5超声科，9骨科，黑龙江省牡丹江市  157011；6深圳市坪山区人民医院(南方医科大学坪山总医院)，广东省深圳市  518118；7牡丹江医学院医学影像学院，黑龙江省牡丹江市  157011；8牡丹江市北药资源开发与应用协同创新中心，黑龙江省牡丹江市  157011；10南方医科大学第三附属医院，广东省医学3D打印应用转化创新平台，广东省广州市  510000
  • 收稿日期:2023-08-05 接受日期:2023-08-26 出版日期:2024-08-08 发布日期:2024-01-19
  • 通讯作者: 黄文华，博士，教授，博士生导师，福建医科大学基础医学院，福建省福州市 350122；南方医科大学基础医学院人体解剖学国家重点学科，广东省医学生物力学重点实验室，广东省医学3D打印应用转化工程技术研究中心，广东省广州市 510515；南方医科大学中西医结合医院骨伤科，广东省广州市510315；深圳市坪山区人民医院(南方医科大学坪山总医院)，广东省深圳市 518118；南方医科大学第三附属医院，广东省医学3D打印应用转化创新平台，广东省广州市 510000 李子涛，硕士，教授，硕士生导师，牡丹江市北药资源开发与应用协同创新中心，黑龙江省牡丹江市 157011；牡丹江医学院附属红旗医院骨科，黑龙江省牡丹江市 157011
  • 作者简介:张闻天，男，1997年生，福建省泉州市人，汉族，福建医科大学在读硕士，主要从事软组织生物力学方面的研究。
  • 基金资助:
    国家重点研发计划(2022YFB4600600)，项目负责人：黄文华；国家自然科学基金(31972915，32271181)，项目负责人：黄文华；国家自然科学基金(82171966)，项目负责人：李彩娟；广东省基础与应用基础研究基金(2020B1515120001)，项目负责人：黄文华；中国博士后科学基金(2022M711533)，项目负责人：邓羽平；广东省优秀青年人才国际培养计划博士后项目，项目负责人：邓羽平；广东省医学科学技术研究基金项目(A2023129)，项目负责人：熊亮；黑龙江省自然科学基金项目(SS2023H004)，项目负责人：李子涛；牡丹江医学院科学基金火炬计划(2022-MYHJ-007)，项目负责人：李彩娟

Elastase quantitative analysis of elastin effect on mechanical response of anterior cruciate ligament

Zhang Wentian1, Deng Yuping2, 3, 4, Liu Xiaoyun2, 4, Li Caijuan5, Wang Mian2, 6, Liang Zeyu2, Xiong Liang3, Huang Gang3, Chen Guangxin7, Li Zitao8, 9, Huang Wenhua1, 2, 3, 6, 10   

  1. 1School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350122, Fujian Province, China; 2Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangdong Provincial Key Laboratory of Medical Biomechanics, National Key Discipline of Human Anatomy, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, Guangdong Province, China; 3Department of Orthopedics and Traumatology, Integrated Hospital of Traditional Chinese and Western Medicine, Southern Medical University, Guangzhou 510315, Guangdong Province, China; 4Shenzhen Bay Laboratory, Institute of Biomedical Engineering, Shenzhen 518132, Guangdong Province, China; 5Ultrasonography Lab, 9Department of Orthopedics, Hongqi Hospital Affiliated to Mudanjiang Medical University, Mudanjiang 157011, Heilongjiang Province, China; 6Pingshan District People’s Hospital of Shenzhen, Pingshan General Hospital of Southern Medical University, Shenzhen 518118, Guangdong Province, China; 7College of Medical Imaging, Mudanjiang Medical University, Mudanjiang 157011, Heilongjiang Province, China; 8Mudanjiang Beiyao Resources Development and Application Collaborative Innovation Center, Mudanjiang 157011, Heilongjiang Province, China; 10Guangdong Medical Innovation Platform for Translation of 3D Printing Application, Third Affiliated Hospital of Southern Medical University, Guangzhou 510000, Guangdong Province, China
  • Received:2023-08-05 Accepted:2023-08-26 Online:2024-08-08 Published:2024-01-19
  • Contact: Huang Wenhua, MD, Professor, Doctoral supervisor, School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350122, Fujian Province, China; Guangdong Engineering Research Center for Translation of Medical 3D Printing Application, Guangdong Provincial Key Laboratory of Medical Biomechanics, National Key Discipline of Human Anatomy, School of Basic Medical Sciences, Southern Medical University, Guangzhou 510515, Guangdong Province, China; Department of Orthopedics and Traumatology, Integrated Hospital of Traditional Chinese and Western Medicine, Southern Medical University, Guangzhou 510315, Guangdong Province, China; Pingshan District People’s Hospital of Shenzhen, Pingshan General Hospital of Southern Medical University, Shenzhen 518118, Guangdong Province, China; Guangdong Medical Innovation Platform for Translation of 3D Printing Application, Third Affiliated Hospital of Southern Medical University, Guangzhou 510000, Guangdong Province, China Li Zitao, Master, Professor, Master’s supervisor, Mudanjiang Beiyao Resources Development and Application Collaborative Innovation Center, Mudanjiang 157011, Heilongjiang Province, China; Department of Orthopedics, Hongqi Hospital Affiliated to Mudanjiang Medical University, Mudanjiang 157011, Heilongjiang Province, China
  • About author:Zhang Wentian, Master candidate, School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350122, Fujian Province, China
  • Supported by:
    National Key Research & Development Plan, No. 2022YFB4600600 (to HWH); National Natural Science Foundation of China, No. 31972915, 32271181 (to HWH); National Natural Science Foundation of China, No. 82171966 (to LCJ); Guangdong Basic and Applied Basic Research Fund, No. 2020B1515120001 (to HWH); China Postdoctoral Science Foundation, No. 2022M711533 (to DYP); Postdoctoral Project of the Guangdong Provincial Excellent Youth Talent International Training Program (to DYP); Guangdong Medical Science and Technology Research Fund Project, No. A2023129 (to XL); Heilongjiang Natural Science Foundation Project, No. SS2023H004 (to LZT); Science Foundation Torch Program of Mudanjiang Medical University, No. 2022-MYHJ-007 (to LCJ)

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摘要：


文题释义：

弹性模量：对弹性体施加一个外界作用力，弹性体会发生形变，其应力和应变呈正比例关系，该比例系数称为弹性模量。弹性模量可视为衡量材料产生弹性变形难易程度的指标，该指标的值越大，使材料发生一定弹性变形的应力也越大，即材料刚度越大。
黏弹性：生物组织的力学特性首先表现在它既有弹性又有一定的黏性，其应力应变不是线弹性关系，也不是弹塑性的非线性关系。生物组织的材料特性被认为是介于黏性和弹性之间的黏弹性体，它在受力时会被拉伸，解除外力也不会立即返回，而是缓慢恢复到原始状态。


背景：前交叉韧带在复杂的生理加载环境下具有其独特的非线性力学特性。弹性蛋白作为前交叉韧带力学特性的重要贡献者，其在轴向拉伸下对前交叉韧带力学响应特性尚不明确。

目的：定量分析弹性蛋白对前交叉韧带拉伸力学响应的影响。
方法：制备弹性蛋白酶溶液及对照缓冲液。将猪前交叉韧带样品制备成小规格样品，随机分别浸泡于 0，0.1，1.0，2.0，5.0，10.0 U/mL的弹性蛋白酶溶液中6 h，另取相同规格小样品分别浸泡于2 U/mL的弹性蛋白酶溶液中 0，1，3，6，9，12 h，以确定适合弹性蛋白靶向酶的浸泡条件并进行消化效果验证，使用组织学染色比较酶处理对组织结构与成分的影响。将韧带样品随机分为酶处理组和PBS组，分别浸泡在2 U/mL弹性蛋白酶溶液和PBS中6 h，于浸泡前、后均进行力学拉伸测试。

结果与结论：①生化结果表明，2 U/mL弹性蛋白酶溶液浸泡6 h能将弹性蛋白含量降低31.1%，且没有显著影响组织内其他力学相关成分；②组织学结果显示，弹性蛋白酶可渗透入组织内部，浸泡后的组织松散程度增加；③在浸泡前、后的力学结果中，PBS组的多项力学性能均显著下降，酶处理组仅有低张弹性模量显著升高及初长度显著增加；④组间比较结果显示，浸泡前PBS组与酶处理组差异均无显著意义，而浸泡后的酶处理组的低张弹性模量、初始斜率、饱和斜率和初长度比PBS组显著增加；⑤以上结果表明，弹性蛋白降解显著影响了前交叉韧带的生物力学属性，进一步补充了对前交叉韧带结构-功能关系之间的理解。  

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料；口腔生物材料；纳米材料；缓释材料；材料相容性；组织工程

关键词: 前交叉韧带, 生物力学, 弹性蛋白, EVG染色, 准静态拉伸, 黏弹性

Abstract: BACKGROUND: The anterior cruciate ligament has unique nonlinear mechanical properties under a complex physiological loading environment. Elastin is an important contributor to the mechanical properties of the anterior cruciate ligament, but its mechanical response to the anterior cruciate ligament under axial tension is not clear.
OBJECTIVE: To quantitatively analyze the effect of elastin on the tensile mechanical response of the anterior cruciate ligament.
METHODS: Elastase solution and control buffer were prepared. The porcine anterior cruciate ligament samples were prepared into small-size samples and randomly soaked in 0, 0.1, 1.0, 2.0, 5.0, and 10.0 U/mL elastase solution for 6 hours, and other small samples of the same size were soaked in 2 U/mL elastase solution for 0, 1, 3, 6, 9, and 12 hours. To determine suitable soaking conditions for elastin-targeted enzymes and verify the digestive effect, histological staining was used to compare the effects of enzyme treatment on tissue structure and composition. The ligament samples were randomly divided into elastase-treated group and PBS group, and immersed in 2 U/mL elastase solution and PBS buffer for 6 hours, respectively. A mechanical tensile test was performed before and after immersion.
RESULTS AND CONCLUSION: (1) The biochemical results showed that being treated in 2 U/mL elastase solution for 6 hours could reduce the elastin content by 31.1%, and had no significant effect on other mechanical-related components in the tissue. (2) The histological results showed that elastase was able to penetrate the tissue, and the loose degree of tissue increased after treatment. (3) In the mechanical results before and after treatment, the mechanical properties of the PBS group decreased significantly, only the low-tension elastic modulus increased significantly and the initial length increased significantly in the elastase-treated group. (4) The intergroup comparison results showed that there was no significant difference between the two groups in pre-treatment, but the low-tension elastic modulus, initial slopes, saturated slopes, and initial length of the elastase-treated group after treatment were significantly higher than those in the PBS group. (5) These results suggest that elastin degradation significantly affects the biomechanical properties of the anterior cruciate ligament and further complements our understanding of the structure-function relationship of the anterior cruciate ligament. 

Key words: anterior cruciate ligament, biomechanics, elastin, EVG staining, quasi-static tensile, viscoelasticity

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