Yuanlin Liu1, Jie Li2, Wei Xia2, Chen Chen1, Heng Zhu1, Jide Chen1, Shaohua Li2, Xueting Su2, Xingliang Qin2, Hongmei Ding2, Long Long3, Lili Wang3, Zhanghua Li4, Wen Liao5, Yi Zhang6, Ningsheng Shao7. 1. Department of Cell Biology, Institute of Basic Medical Sciences, Beijing 100850, China. 2. Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, Beijing 100850, China. 3. Laboratory of Computer-aided Drug Design & Discovery, Institute of Pharmacology and Toxicology, Beijing 100850, China. 4. Department of Orthopedics, Renmin Hospital of Wuhan University, Wuhan 430060, China. 5. Baoding First Central Hospital, Hebei Province, Baoding 071000, China. 6. Department of Cell Biology, Institute of Basic Medical Sciences, Beijing 100850, China. Electronic address: zhangyi612@hotmail.com. 7. Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, Beijing 100850, China. Electronic address: shaonsh@hotmail.com.
Abstract
AIMS: The aim of the study was to explore the effect of miR-200b on the development of human peripheral blood monocyte-deriveddendritic-cell (DC) and its mechanisms. MAIN METHODS: Expression levels of miR-200b and its predicted targets were measured by real time-PCR. Protein expression of WASF3 was determined by Western blot and immunohistochemistry. Human peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque density gradient centrifugation from the buffy coat fraction of anticoagulated blood. Monocytes were purified from PBMCs using anti-CD14 microbeads. The immunophenotypes of DCs were tested by flow cytometry. KEY FINDINGS: A strong reduction in miR-200b expression was associated with human peripheral blood monocyte-derivedDC differentiation. The overexpression of miR-200b significantly reduced the numbers of protruding veils in mature DCs (mDCs) that are critical for promoting antigen-specificT-cell activation. Further experiments showed that miR-200b could regulate the function of DCs by targeting WASF3, a protein involved in cell movement and invasion. SIGNIFICANCE: Our results define an important function of miR-200b in the negative regulation of DC development and provide a potential form of miRNA-mediated cell therapy for diseases that range from auto-immunity to graft-versus-host disease.
AIMS: The aim of the study was to explore the effect of miR-200b on the development of human peripheral blood monocyte-deriveddendritic-cell (DC) and its mechanisms. MAIN METHODS: Expression levels of miR-200b and its predicted targets were measured by real time-PCR. Protein expression of WASF3 was determined by Western blot and immunohistochemistry. Human peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque density gradient centrifugation from the buffy coat fraction of anticoagulated blood. Monocytes were purified from PBMCs using anti-CD14 microbeads. The immunophenotypes of DCs were tested by flow cytometry. KEY FINDINGS: A strong reduction in miR-200b expression was associated with human peripheral blood monocyte-derivedDC differentiation. The overexpression of miR-200b significantly reduced the numbers of protruding veils in mature DCs (mDCs) that are critical for promoting antigen-specificT-cell activation. Further experiments showed that miR-200b could regulate the function of DCs by targeting WASF3, a protein involved in cell movement and invasion. SIGNIFICANCE: Our results define an important function of miR-200b in the negative regulation of DC development and provide a potential form of miRNA-mediated cell therapy for diseases that range from auto-immunity to graft-versus-host disease.