Chinese Journal of Tissue Engineering Research ›› 2021, Vol. 25 ›› Issue (26): 4168-4174.doi: 10.12307/2021.116

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Extracellular matrix promotes fibroblast proliferation and post-operation knee arthrofibrosis through ERK signaling pathway

Hou Jingzhao1, Zhang Zhen2   

  1. 1Jingjiang People’s Hospital, Taizhou 214500, Jiangsu Province, China; 2Dalian Medical University, Dalian 116044, Liaoning Province, China
  • Received:2020-07-27 Revised:2020-07-29 Accepted:2020-08-19 Online:2021-09-18 Published:2021-05-10
  • Contact: Zhang Zhen, Physician, Dalian Medical University, Dalian 116044, Liaoning Province, China E-mail:1395637072@qq.com
  • About author:Hou Jingzhao, Master, Associate chief physician, Jingjiang People’s Hospital, Taizhou 214500, Jiangsu Province, China

Abstract:

BACKGROUND: Osteoporosis and fracture type are two important reasons for the failure of internal fixation of proximal femoral nail antirotation. Type AO31-A3.3 interArthrofibrosis is one of the most serious complications. It is currently considered that extracellular matrix plays an important role in the occurrence of fibrosis; however, the mechanism of extracellular matrix is still unclear. 

OBJECTIVE: To explore the effect of extracellular matrix on post-operation arthrofibrosis and fibroblast proliferation in vitro.  
METHODS: (1) In vivo test: twelve New Zealand rabbit models of lower-lime knee arthrofibrosis were established; and the influence of the extracellular matrix on post-operation arthrofibrosis was studied by hematoxylin-eosin staining, Masson staining, Sirius red staining and proliferating cell nuclear antigen immunohistochemical staining at 2 and 4 weeks after surgery. (2) In vitro test: Fibroblast derived extracellular matrix and decellularized extracellular matrix were constructed, and the thickness of extracellular matrix was measured by confocal three-dimensional imaging. Fibroblasts were cultured in three-dimensional and two-dimensional extracellular matrix. The function of extracellular matrix was evaluated by cell membrane staining and rapid cell attachment test. Cell count, EdU test, cell cycle and western blot assay were used to evaluate the effect of extracellular matrix on cell proliferation, and the expression of ERK signaling pathway protein was detected.
RESULTS AND CONCLUSION: In vivo test: hematoxylin-eosin staining, Masson trichrome staining and Sirius red staining results showed that the degree of fibrosis was serious at 4 weeks; and the collagen synthesis of extracellular matrix in arthrofibrosis tissue increased obviously. Sirius red staining results showed that the collagen was mainly composited of collagen type I and III. Immunohistochemical staining showed that number of fibroblasts and the level of PCNA protein in fibrotic tissue increased 4 weeks after operation. In vitro test: The thickness of the outer matrix was more than 10 μm. Cell membrane staining showed that fibroblasts in three-dimensional culture were spindle shaped, while those in two-dimensional culture were irregular. rapid cell attachment test demonstrated that the number of cell adhesion in three-dimensional culture was 6 times that in two-dimensional culture (P < 0.05). Cell count, EdU test, and cell cycle analysis showed that the proliferation of fibroblasts in three-dimensional culture was faster than that in two-dimensional culture (P < 0.05). Western blot assay showed that the expression of PCNA, Cyclin D1, p-MEK and p-ERK in three-dimensional culture was higher than that in two-dimensional culture            (P < 0.05). In summary, extracellular matrix promotes the occurrence of post-operation arthrofibrosis and fibroblast proliferation possibly through ERK signaling pathway.

Key words: bone, material, extracellular matrix, fibrosis, extracellular matrix, fibroblast, proliferation, signaling pathway, protein

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