中国组织工程研究 ›› 2024, Vol. 28 ›› Issue (16): 2542-2549.doi: 10.12307/2024.304

• 肌肉肌腱韧带组织构建 tissue construction of the muscle, tendon and ligament • 上一篇    下一篇

补充重组人源胶原蛋白对离心运动后小鼠骨骼肌细胞外基质重塑的影响

赵莎莎1,贺  庆2,李  佳2,吴  迎1   

  1. 1北京体育大学运动人体科学学院,北京市  100084;2艾兰得健康控股有限公司,上海市  200120
  • 收稿日期:2023-03-08 接受日期:2023-04-20 出版日期:2024-06-08 发布日期:2023-07-29
  • 通讯作者: 吴迎,硕士生导师,副教授,北京体育大学运动人体科学学院,北京市 100084
  • 作者简介:赵莎莎,女,1998年生,河南省新乡市人,汉族,北京体育大学在读硕士,主要从事运动与营养相关研究。
  • 基金资助:
    中央高校基本科研业务专项资金资助项目(2020054),项目负责人:吴迎;横向联合项目(20210056),项目负责人:吴迎

Effects of recombinant human collagen supplementation on extracellular matrix remodeling in mouse skeletal muscle after eccentric exercise

Zhao Shasha1, He Qing2, Li Jia2, Wu Ying1   

  1. 1College of Sports Science, Beijing Sport University, Beijing 100084, China; 2Aland Health Holding Ltd., Shanghai 200120, China
  • Received:2023-03-08 Accepted:2023-04-20 Online:2024-06-08 Published:2023-07-29
  • Contact: Wu Ying, Chief physician, Master’s supervisor, Associate professor, College of Sports Science, Beijing Sport University, Beijing 100084, China
  • About author:Zhao Shasha, Master candidate, College of Sports Science, Beijing Sport University, Beijing 100084, China
  • Supported by:
    the Fundamental Research Fund for the Central Universities, No. 2020054 (to WY); Horizontal Joint Project, No. 20210056 (to WY)

摘要:


文题释义:

胶原蛋白:为三螺旋结构,是含量最为丰富的胞外基质成分,属于不完全蛋白质。目前市售胶原蛋白主要为动物来源,存在提取工艺复杂、活性表达较低及存在污染风险等问题,而利用生物工程技术生产的重组人源胶原蛋白具有分子质量确定、可加工性强、水溶性强、排异反应低、无病毒隐患等优势。
细胞外基质:由细胞合成并分泌到胞外的物质,是一种普遍存在于组织器官中的非细胞成分。骨骼肌细胞外基质所构成的膜性网状结构具有支持、连接、营养、保护和修复肌组织的作用,对肌纤维、肌束和肌肉整体活动也起着重要的调节作用。


背景:胶原蛋白是含量最为丰富的胞外基质成分,与骨骼肌细胞外基质的结构、功能密切相关,但补充由生物工程技术生产的重组人源胶原蛋白(recombinant human collagen,rhC)对骨骼肌细胞外基质的影响及机制尚不清楚。

目的:探讨补充rhC对离心运动后骨骼肌细胞外基质重塑的影响及作用机制。
方法:将104只8周龄健康雄性C57小鼠随机分为对照组(生理盐水)、rhC低剂量组(0.2 g/kg)、中剂量组(1.0 g/kg)和高剂量组(2.0 g/kg),连续7 d灌胃干预后每组选取2只小鼠,解剖各脏器进行苏木精-伊红染色判断炎性浸润情况,其余小鼠于第8天进行离心运动,分别在离心运动后即刻(0),24,48,96 h观察细胞外基质结构变化,检测小鼠抓握力、血清肌酸激酶活性、骨骼肌组织基质金属蛋白酶2,9,14和基质金属蛋白酶抑制剂2的蛋白水平。

结果与结论:①苏木精-伊红染色观察短期补充rhC对心脏、肝脏、脾、肾无显著影响;②一次性离心运动后,rhC中、高剂量组小鼠抓握力恢复率显著升高(P < 0.01);③各组肌酸激酶变化趋势一致,组间无显著性差异;④rhC低剂量组细胞外基质恢复进程快于对照组,rhC中、高剂量组肌束膜较为完整;⑤rhC高剂量组基质金属蛋白酶9蛋白水平显著降低(P < 0.05);⑥rhC中、高剂量组基质金属蛋白酶14蛋白水平显著降低(P < 0.05);⑦rhC中、高剂量组基质金属蛋白酶2蛋白水平显著降低(P < 0.05);⑧rhC中、高剂量组基质金属蛋白酶抑制剂2蛋白水平显著升高(P < 0.05);⑨rhC各剂量组基质金属蛋白酶2/基质金属蛋白酶抑制剂2比值显著降低(P < 0.05)。该研究发现连续7 d预补充1.0,2.0 g/kg rhC可通过调节基质金属蛋白酶和基质金属蛋白酶抑制剂来抑制运动后骨骼肌细胞外基质降解,从而促进小鼠骨骼肌力量恢复。

https://orcid.org/0000-0002-0276-2311(赵莎莎)

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程

关键词: 重组人源胶原蛋白, 离心运动, 骨骼肌细胞外基质, 基质金属蛋白酶, 基质金属蛋白酶抑制剂

Abstract: BACKGROUND: Collagen is the most abundant extracellular matrix component, which is closely related to the structure and function of the extracellular matrix of skeletal muscle, but the effect and mechanism of recombinant human collagen (rhC) produced by bioengineering technology on the extracellular matrix of skeletal muscle are unclear.
OBJECTIVE: To investigate the effect of rhC supplementation on the remodeling of skeletal muscle extracellular matrix after eccentric exercise, thereby revealing the possible mechanism by which rhC improves the injury of skeletal muscle extracellular matrix and promote the recovery after exercise.
METHODS: A total of 104 healthy male C57 mice aged 8 weeks old were randomly divided into control group (normal saline), low-dose rhC group (0.2 g/kg), medium-dose rhC group (1.0 g/kg), and high-dose rhC group (2.0 g/kg). Two mice in each group were selected after continuous 7 days of intragastric intervention, and organs were dissected for hematoxylin-eosin staining to determine inflammatory infiltrates. On the 8th day, the remaining mice were subjected to eccentric exercise. The structural changes of the skeletal muscle extracellular matrix were observed under scanning electron microscopy immediately (0), 24, 48, and 96 hours  after eccentric exercise. Meanwhile, grip strength, creatine kinase activity, and protein levels of matrix metalloproteinases 2, 9, 14 and tissue inhibitor of metalloproteinase-2 in skeletal muscle were detected by western blot assay.
RESULTS AND CONCLUSION: Hematoxylin-eosin staining results indicated that short-term rhC supplementation showed no significant effects on the morphology of the heart, liver, spleen and kidney. After one-time eccentric exercise, the recovery rate of grip strength in the medium- and high-dose rhC groups was significantly increased (P < 0.01). The trend of creatine kinase changes was consistent in all groups and there was no significant difference between groups. The recovery process of the extracellular matrix in the low-dose rhC group was faster than that in the control group, and the muscle tract membrane in the medium- and high-dose rhC groups was more complete. The protein level of matrix metalloproteinase 9 in the high-dose rhC group was significantly decreased (P < 0.05). The protein levels of matrix metalloproteinase 14 in the medium- and high-dose rhC groups were significantly decreased (P < 0.05). The protein levels of matrix metalloproteinase 2 in the medium- and high-dose rhC groups were significantly decreased (P < 0.05). Tissue inhibitor of metalloproteinase-2 protein levels in the medium- and high-dose rhC groups were significantly increased (P < 0.05). The ratio of matrix metalloproteinase 2 to tissue inhibitor of metalloproteinase-2 in each rhC group was significantly decreased (P < 0.05). To conclude, pre-supplementation of 1.0 and 2.0 g/kg rhC for 7 days can inhibit extracellular matrix degradation in skeletal muscle after exercise by modulating matrix metalloproteinases and matrix metalloproteinase inhibitors, thereby promoting recovery of skeletal muscle strength in mice.

Key words: recombinant human collagen, eccentric exercise, skeletal muscle extracellular matrix, matrix metalloproteinase, tissue inhibitor of metalloproteinases

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