Overexpression of Notch3 is associated with metastasis and poor prognosis in osteosarcoma patients

Abstract

Xue-Feng Tang,1 Ya Cao,1 Dong-Bin Peng,1 Guo-Sheng Zhao,2 Ying Zeng,1,3 Zi-Ran Gao,1 Yang-Fan Lv,1 Qiao-Nan Guo1 1Department of Pathology, Xinqiao Hospital, Army Medical University, Chongqing 400037, China; 2Department of Orthopedic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, China; 3Department of Pathology, Daping Hospital, Army Medical University, Chongqing 400037, China Background: Notch signaling abnormalities are associated with the development of various tumors, including hematopoietic and epithelium-derived tumors. However, the role of Notch signaling in tumors originating from mesenchymal cells is unclear. The effect of Notch3 expression on the prognosis of osteosarcoma and its role and mechanism in osteosarcoma cells have never been reported. Materials and methods: In this study, we performed a clinicopathological analysis of 70 cases of osteosarcoma, with primary focus on survival. Osteosarcoma cell lines MTH and U2OS were used. After knockdown of Notch3 by lentiviral transfection and siRNA, the cell cycle, cell viability, and wound healing capacity were assessed. Subsequently, the Transwell assay was performed, and the expression levels of hairy and enhancer of split-1 (Hes1) and matrix metalloproteinase 7 (MMP7) were detected by RT-PCR and Western blot assay. The expression of MMP7 was also detected after knockdown of Hes1. Animal experiments were performed by injecting the cell lines MTH of Notch3 knockdown into mice tail veins and comparing the development of lung metastasis with the control group. Results: Comparison of survival curves showed that Notch3 expression significantly impacts patient survival. Additionally, multivariate analysis revealed that Notch3 is an independent prognostic factor for osteosarcoma. In in vivo experiments, osteosarcoma-associated pulmonary metastasis in nude mice was reduced after Notch3 silencing. The expression of downstream effector molecule, Hes1, and that of the invasion and metastasis-associated proteolytic enzyme, MMP7, were reduced, and MMP7 was further decreased by Hes1 knockdown in in vitro experiments. Conclusion: Notch3 is a prognostic factor for osteosarcoma and might regulate its invasion and metastasis through the downstream target gene Hes1 and effector MMP7. Keywords: osteosarcoma, prognosis, metastasis, Notch3, Hes1, MMP7

Keywords

• osteosarcoma
• prognosis
• metastasis
• Notch3
• Hes1
• MMP7