Laboratory Research
Mechanism of recombinant human S100 calcium binding protein A4 inducing vascular endothelial growth factors in rheumatoid arthritis fibroblast-like synoviocytes
Dingsheng Zha, Wuyan Xu, Hao Wu, Hongsheng Lin, Zhengang Zha, Hua Yang, Yuhao Yang, Jie Ren
Published 2018-05-15
Cite as Chin J Orthop Trauma, 2018, 20(5): 430-438. DOI: 10.3760/cma.j.issn.1671-7600.2018.05.011
Abstract
ObjectiveTo investigate the mechanism of inducing production of vascular endothelial growth factors (VEGF) by recombinant human S100 calcium binding protein A4 (rhS100A4) in rheumatoid arthritis fibroblast-like synoviocytes (RAFLSs).
MethodsSynovial tissue was sampled from the patients with rheumatoid arthritis (RA) undergoing knee arthroplasty for in vitro culture of RAFLSs. CCK-8 assay was conducted to detect the effect of rhS100A4 and the effect of its interaction with Rapamycin (Rap), an inhibitor of mammalian rapamycin target 1 (mTORC1) signaling pathway, on the proliferation of RAFLSs. The effects of rhS100A4 and its interaction with Rap on the expression of VEGF in RAFLSs were detected by immunofluorescence. After rhS100A4 and its cooperation with Rap stimulated the conditioned medium (CM) produced by RAFLSs, the effect of CM on formation of lumen in human umbilical vein endothelial cells (HUVECs) in vitro was observed to detect the angiogenic ability of rhS100A4. Western blot was used to detect the effect of rhS100A4 on the phosphorylation of downstream ribosomal protein S6 (S6) in the mTORC1 signaling pathway in RAFLSs and to analyze the effects of rhS100A4 and Rap on phosphorylation of S6 protein and expression of VEGF protein in RAFLSs.
ResultsrhS100A4 promoted cell proliferation and expression of VEGF protein in RAFLSs, and the CM formed by rhS100A4 promoted HUVECs to form blood vessels in vitro. Rap inhibited the above biological effects of rhS100A4. rhS100A4 activated the downstream protein S6 in the mTORC1 signaling pathway in RAFLSs cells to increase their phosphorylation levels. The effects of rhS100A4 on the phosphorylation of S6 protein and on the expression of VEGF protein in RAFLSs were inhibited by Rap.
ConclusionrhS100A4 promotes production of VEGF in RAFLSs by activating the mTORC1 signaling pathway.
Key words:
Arthritis, rheumatoid; Calcium binding protein A4; Vascular endothelial growth factors; mTORC1 signaling pathway
Contributor Information
Dingsheng Zha
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Wuyan Xu
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Hao Wu
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Hongsheng Lin
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Zhengang Zha
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Hua Yang
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Yuhao Yang
Department of Orthopaedics, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
Jie Ren
Department of Rheumatology, The First Affiliated Hospital to Jinan University, Guangzhou 510630, China
