A rapid histological preparation method for observation of morphology and composition distribution of tendon collagen fascicle and endotendinium_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

ZHANG Yi ¹ , ZHOU Li ¹ , TANG Linqiao ¹ , QIN Tingwu ² ,  NING Liangju ²

1. Research Core Facility of West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P.R.China;
2. Laboratory of Stem Cell and Tissue Engineering, West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P.R.China;

Corresponding author：

NING Liangju, Email: nlj1024@163.com

Keywords：

Tendon; frozen section; baking; Masson staining

DOI：

10.7507/1002-1892.201903101

Video：

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Objective To explore a rapid histological preparation method to observe morphology and composition distribution of tendon collagen fascicle and endotendinum. Methods Taking porcine superflexor tendon of foot as an example, tendons were sliced into sections with 6 μm by frozen section technology, after which general observation of the section integrity was carried out. After fixed with 10% neutral buffered formalin and performed with HE staining, the tissue integrity and ice crystal formation were observed under microscope. Sections were then divided into 5 groups by different methods of dyeing. Group A: Priodic acid-Shiff (PAS) staining; group B: Masson staining; group C: reticular fibers staining; group D: immunohistochemical and immunofluorescent staining of type Ⅲ collagen; group E: the sections were baked at 65℃ for 10 minutes and stained with Masson. The composition distribution of tendon collagen fascicle and endotendinum in different groups were observed. Results From general observation, the frozen section of tendon tissue was complete and continuous. Although the tissue integrity in the tendon sections could be seen and no ice crystal was formed, the composition distribution could not be identified by HE staining. The entire tendons in groups A, B, and C were dyed, and the composition distribution of collagen fascicle and endotendinum could not be identified. The endotendinum in group D was stained weakly positive for type Ⅲ collagen alone, and the two components were differentiated dyed but the contrast was not obvious. In group E, the collagen fascicle and endotendinium were differentiated dyed and the two components in tendon tissue were clearly visible. Conclusion The morphology and the composition distribution of tendon collagen fascicle and endotendinum can be characterized rapidly and accurately, using a combination of baking at 65℃ for 10 minutes and Masson staining after porcine superflexor tendons were sliced by frozen section technology.

Citation： ZHANG Yi, ZHOU Li, TANG Linqiao, QIN Tingwu, NING Liangju. A rapid histological preparation method for observation of morphology and composition distribution of tendon collagen fascicle and endotendinium. Chinese Journal of Reparative and Reconstructive Surgery, 2019, 33(9): 1169-1173. doi: 10.7507/1002-1892.201903101 Copy

1.	Liu Y, Ramanath HS, Wang DA. Tendon tissue engineering using scaffold enhancing strategies. Trends Biotechnol, 2008, 26(4): 201-209.
2.	Kannus P. Structure of the tendon connective tissue. Scand J Med Sci Sports, 2000, 10(6): 312-320.
3.	Qin TW, Chen Q, Sun YL, et al. Mechanical characteristics of native tendon slices for tissue engineering scaffold. J Biomed Mater Res B Appl Biomater, 2012, 100B(3): 752-758.
4.	顾怡瑾, 陆新元, 付华辉, 等. Masson 染色在辅助诊断肝纤维化中应用. 临床与实验病理学杂志, 2012, 28(1): 101-102.
5.	杨小荣, 刘芳, 汪芳, 等. Masson 染色法在周围神经损伤中的应用. 诊断病理学杂志, 2012, 19(6): 473-474.
6.	Huang YZ, Wang JJ, Huang YC, et al. Organic composite-mediated surface coating of human acellular bone matrix with strontium. Mater Sci Eng C Mater Biol Appl, 2017, 84: 12-20.
7.	Ni P, Ding Q, Fan M, et al. Injectable thermosensitive PEG-PCL-PEG hydrogel/acellular bone matrix composite for bone regeneration in cranial defects. Biomaterials, 2014, 35(1): 236-248.
8.	Wang ZL, Wu SZ, Li ZF, et al. Comparison of small intestinal submucosa and polypropylene mesh for abdominal wall defect repair. J Biomater Sci Polym Ed, 2018, 29(6): 1-20.
9.	He SK, Guo JH, Wang ZL, et al. Efficacy and safety of small intestinal submucosa in dural defect repair in a canine model. Mater Sci Eng C Mater Biol Appl, 2017, 73: 267-274.
10.	韩天龙, 王敏, 红敏, 等. Masson 三色染色与 HE 染色在豚鼠牙齿组织石蜡切片中的比较研究及应用. 中国畜牧兽医, 2011, 38(3): 55-57.
11.	王珏, 朱礼国, 唐幕湘, 等. 改良 Masson 三色染色法在胶原纤维中的应用. 郧阳医学院学报, 2006, 25(1): 44.
12.	张哲, 王伟, 商建峰, 等. 胶原纤维特殊染色法在心肌纤维化诊断中的应用比较. 诊断病理学杂志, 2016, 23(8): 578-580.
13.	蔡新华, 郭志坤, 霍殿样. 石蜡包埋对兔肝、肾组织块的收缩影响. 解剖科学进展, 2000, 6(4): 380-381.
14.	蔡新华, 郭志坤. 石蜡包埋对心、肺和脾组织的收缩影响. 解剖学杂志, 2002, 25(2): 198-199.
15.	王华, 罗殿中. 术中冰冻切片制片的规范化操作与常见问题原因分析. 2011 年全国西安病理技术学术会议暨全军病理技术学术会议论文集. 西安: 解放军医学科学技术委员会, 2011.
16.	包翠芬, 刘霞, 穆长征, 等. 比较三种防冰晶法对切片保存效果的影响. 中国组织化学与细胞化学杂志, 2007, 16(1): 122-123.
17.	杨秀静, 滕孝静. 冰冻切片技术经验探讨. 临床和实验医学杂志, 2014, 13(23): 2002-2004.
18.	王珏, 朱礼国, 张健, 等. 肾脏 PAS 染色法的探讨. 实用医技杂志, 2006, 13(22): 4097.
19.	钟妮娜, 耿毅, 彭西. PAS 染色技术的改进. 解剖学杂志, 2005, 28(6): 723-724.
20.	胡沛臻, 马福成, 张传山, 等. 微波网状纤维染色法在病理诊断中的应用. 诊断病理学杂志, 2002, 9(4): 249.
21.	李萍萍, 葛青. 软骨寡聚基质蛋白 (COMP) 对 B 淋巴细胞发育及功能的影响.中国细胞生物学学会全国学术大会论文集. 武汉: 中国细胞生物学学会, 2013.
22.	魏壮, 崔树森, 张巨, 等. 预防肌腱粘连的研究与进展. 吉林大学学报, 2001, 27(2): 218-220.
23.	Franchi M, Trirè A, Quaranta M, et al. Collagen structure of tendon relates to function. The Scientific World Journal, 2017, 7: 404-420.
24.	解立武, 常芳, 卢小波. 经典和改良 Gomori 网状纤维染色的比较. 中国药物与临床, 2018, 18(1): 31-32.
25.	蔡新华, 郭志坤, 毛会丽, 等. 大鼠心肌组织中网状纤维组织学构筑及增龄变化. 解剖学报, 2009, 40(1): 127-129.
26.	廖大清, 马玉琼, 陈思秀. 一种显示网状纤维的方法. 四川解剖学杂志, 2001, 9(2): 92-93.
27.	韩庆伟, 张云. 特殊染色网状纤维在病理诊断中的应用. 实用医技杂志, 2007, 14(11): 1420.
28.	田玉旺, 李琳, 朱红艳, 等. 提高网状纤维染色质量的技术探讨. 中国组织化学与细胞学杂志, 2012, 21(3): 316-318.
29.	Wu F, Nerlich M, Docheva D. Tendon injuries: Basic science and repair proposals. EFFORT Open Rev, 2017, 2(7): 332-342.

1. Liu Y, Ramanath HS, Wang DA. Tendon tissue engineering using scaffold enhancing strategies. Trends Biotechnol, 2008, 26(4): 201-209.
2. Kannus P. Structure of the tendon connective tissue. Scand J Med Sci Sports, 2000, 10(6): 312-320.
3. Qin TW, Chen Q, Sun YL, et al. Mechanical characteristics of native tendon slices for tissue engineering scaffold. J Biomed Mater Res B Appl Biomater, 2012, 100B(3): 752-758.
4. 顾怡瑾, 陆新元, 付华辉, 等. Masson 染色在辅助诊断肝纤维化中应用. 临床与实验病理学杂志, 2012, 28(1): 101-102.
5. 杨小荣, 刘芳, 汪芳, 等. Masson 染色法在周围神经损伤中的应用. 诊断病理学杂志, 2012, 19(6): 473-474.
6. Huang YZ, Wang JJ, Huang YC, et al. Organic composite-mediated surface coating of human acellular bone matrix with strontium. Mater Sci Eng C Mater Biol Appl, 2017, 84: 12-20.
7. Ni P, Ding Q, Fan M, et al. Injectable thermosensitive PEG-PCL-PEG hydrogel/acellular bone matrix composite for bone regeneration in cranial defects. Biomaterials, 2014, 35(1): 236-248.
8. Wang ZL, Wu SZ, Li ZF, et al. Comparison of small intestinal submucosa and polypropylene mesh for abdominal wall defect repair. J Biomater Sci Polym Ed, 2018, 29(6): 1-20.
9. He SK, Guo JH, Wang ZL, et al. Efficacy and safety of small intestinal submucosa in dural defect repair in a canine model. Mater Sci Eng C Mater Biol Appl, 2017, 73: 267-274.
10. 韩天龙, 王敏, 红敏, 等. Masson 三色染色与 HE 染色在豚鼠牙齿组织石蜡切片中的比较研究及应用. 中国畜牧兽医, 2011, 38(3): 55-57.
11. 王珏, 朱礼国, 唐幕湘, 等. 改良 Masson 三色染色法在胶原纤维中的应用. 郧阳医学院学报, 2006, 25(1): 44.
12. 张哲, 王伟, 商建峰, 等. 胶原纤维特殊染色法在心肌纤维化诊断中的应用比较. 诊断病理学杂志, 2016, 23(8): 578-580.
13. 蔡新华, 郭志坤, 霍殿样. 石蜡包埋对兔肝、肾组织块的收缩影响. 解剖科学进展, 2000, 6(4): 380-381.
14. 蔡新华, 郭志坤. 石蜡包埋对心、肺和脾组织的收缩影响. 解剖学杂志, 2002, 25(2): 198-199.
15. 王华, 罗殿中. 术中冰冻切片制片的规范化操作与常见问题原因分析. 2011 年全国西安病理技术学术会议暨全军病理技术学术会议论文集. 西安: 解放军医学科学技术委员会, 2011.
16. 包翠芬, 刘霞, 穆长征, 等. 比较三种防冰晶法对切片保存效果的影响. 中国组织化学与细胞化学杂志, 2007, 16(1): 122-123.
17. 杨秀静, 滕孝静. 冰冻切片技术经验探讨. 临床和实验医学杂志, 2014, 13(23): 2002-2004.
18. 王珏, 朱礼国, 张健, 等. 肾脏 PAS 染色法的探讨. 实用医技杂志, 2006, 13(22): 4097.
19. 钟妮娜, 耿毅, 彭西. PAS 染色技术的改进. 解剖学杂志, 2005, 28(6): 723-724.
20. 胡沛臻, 马福成, 张传山, 等. 微波网状纤维染色法在病理诊断中的应用. 诊断病理学杂志, 2002, 9(4): 249.
21. 李萍萍, 葛青. 软骨寡聚基质蛋白 (COMP) 对 B 淋巴细胞发育及功能的影响.中国细胞生物学学会全国学术大会论文集. 武汉: 中国细胞生物学学会, 2013.
22. 魏壮, 崔树森, 张巨, 等. 预防肌腱粘连的研究与进展. 吉林大学学报, 2001, 27(2): 218-220.
23. Franchi M, Trirè A, Quaranta M, et al. Collagen structure of tendon relates to function. The Scientific World Journal, 2017, 7: 404-420.
24. 解立武, 常芳, 卢小波. 经典和改良 Gomori 网状纤维染色的比较. 中国药物与临床, 2018, 18(1): 31-32.
25. 蔡新华, 郭志坤, 毛会丽, 等. 大鼠心肌组织中网状纤维组织学构筑及增龄变化. 解剖学报, 2009, 40(1): 127-129.
26. 廖大清, 马玉琼, 陈思秀. 一种显示网状纤维的方法. 四川解剖学杂志, 2001, 9(2): 92-93.
27. 韩庆伟, 张云. 特殊染色网状纤维在病理诊断中的应用. 实用医技杂志, 2007, 14(11): 1420.
28. 田玉旺, 李琳, 朱红艳, 等. 提高网状纤维染色质量的技术探讨. 中国组织化学与细胞学杂志, 2012, 21(3): 316-318.
29. Wu F, Nerlich M, Docheva D. Tendon injuries: Basic science and repair proposals. EFFORT Open Rev, 2017, 2(7): 332-342.

Chinese Journal of Reparative and Reconstructive Surgery

A rapid histological preparation method for observation of morphology and composition distribution of tendon collagen fascicle and endotendinium

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