Huaqiang Zhu1, Xian Jiang2, Xu Zhou1, Xuesong Dong2, Kai Xie1, Chuncheng Yang1, Hongchi Jiang2, Xueying Sun2, Jun Lu1. 1. Department of Hepatobiliary Surgery, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, China. 2. Key Laboratory of Hepatosplenic Surgery, Department of General Surgery, the First Affiliated Hospital of Harbin Medical University, Harbin, China.
Abstract
BACKGROUND & AIMS: Neuropilin-1 (NRP-1) activates signalling pathways as multifunctional co-receptors in cancer cells. However, its role and how it is regulated by miRNAs in cholangiocarcinoma (CCA) have not yet been investigated. METHODS: The expression of NRP-1, miR-320 and key molecules involved in cell proliferation, migration and related signalling pathways were detected by immunohistochemistry, immunoblotting and qRT-PCR. Stable transfectants depleted of NRP-1 were generated. The regulatory effect of miR-320 on NRP-1 was evaluated by luciferase reporter assays. Cell proliferation, cell cycle distribution and migration were examined. Xenograft tumour models were established to assess tumourigenesis, tumour growth and lung metastasis. RESULTS: Cholangiocarcinoma tissues expressed higher levels of NRP-1 than adjacent normal biliary tissues, and its expression negatively correlated with miR-320. NRP-1 depletion inhibited cell proliferation and induced cell cycle arrest in the G1/S phase by upregulating p27, and downregulating cyclin E and cyclin-dependent kinase 2; and reduced cell migration by inhibiting the phosphorylation of focal adhesion kinase. NRP-1 depletion suppressed tumourigenesis, tumour growth and lung metastasis by inhibiting cell proliferation and tumour angiogenesis in experimental animals. Depletion of NRP-1 inhibited the activation of VEGF/VEGFR2, EGF/EGFR and HGF/c-Met pathways stimulated by respective ligands. MiR-320 negatively regulated the expression of NRP-1 by binding to the 3'-UTR of NRP-1 promoter, and miR-320 mimics inhibited cell proliferation and migration, and the growth of established tumours in animals by downregulating NRP-1. CONCLUSIONS: The present results indicate that NRP-1 is negatively regulated by miR-320, and both of them may be potentially therapeutic targets for CCA.
BACKGROUND & AIMS:Neuropilin-1 (NRP-1) activates signalling pathways as multifunctional co-receptors in cancer cells. However, its role and how it is regulated by miRNAs in cholangiocarcinoma (CCA) have not yet been investigated. METHODS: The expression of NRP-1, miR-320 and key molecules involved in cell proliferation, migration and related signalling pathways were detected by immunohistochemistry, immunoblotting and qRT-PCR. Stable transfectants depleted of NRP-1 were generated. The regulatory effect of miR-320 on NRP-1 was evaluated by luciferase reporter assays. Cell proliferation, cell cycle distribution and migration were examined. Xenograft tumour models were established to assess tumourigenesis, tumour growth and lung metastasis. RESULTS:Cholangiocarcinoma tissues expressed higher levels of NRP-1 than adjacent normal biliary tissues, and its expression negatively correlated with miR-320. NRP-1 depletion inhibited cell proliferation and induced cell cycle arrest in the G1/S phase by upregulating p27, and downregulating cyclin E and cyclin-dependent kinase 2; and reduced cell migration by inhibiting the phosphorylation of focal adhesion kinase. NRP-1 depletion suppressed tumourigenesis, tumour growth and lung metastasis by inhibiting cell proliferation and tumour angiogenesis in experimental animals. Depletion of NRP-1 inhibited the activation of VEGF/VEGFR2, EGF/EGFR and HGF/c-Met pathways stimulated by respective ligands. MiR-320 negatively regulated the expression of NRP-1 by binding to the 3'-UTR of NRP-1 promoter, and miR-320 mimics inhibited cell proliferation and migration, and the growth of established tumours in animals by downregulating NRP-1. CONCLUSIONS: The present results indicate that NRP-1 is negatively regulated by miR-320, and both of them may be potentially therapeutic targets for CCA.
Authors: Raphael Mohr; Burcin Özdirik; Jana Knorr; Alexander Wree; Münevver Demir; Frank Tacke; Christoph Roderburg Journal: Int J Mol Sci Date: 2020-07-15 Impact factor: 5.923