Dataset Information

CCN3 Facilitates Runx2 and Osterix Expression by Inhibiting miR-608 through PI3K/Akt Signaling in Osteoblasts.

ABSTRACT: CCN3, otherwise known as the nephroblastoma overexpressed (NOV) protein, is a cysteine-rich protein that belongs to the CCN family and regulates several cellular functions. Osteoblasts are major bone-forming cells that undergo proliferation, mineralization, renewal, and repair during the bone formation process. We have previously reported that CCN3 increases bone morphogenetic protein 4 (BMP-4) production and bone mineralization in osteoblasts, although the role of CCN3 remains unclear with regard to osteogenic transcription factors (runt-related transcription factor 2 (Runx2) and osterix). Here, we used alizarin red-S and alkaline phosphatase staining to show that CCN3 enhances osteoblast differentiation. Stimulation of osteoblasts with CCN3 increases expression of osteogenic factors such as BMPs, Runx2, and osterix. Moreover, we found that the inhibition of miR-608 expression is involved in the effects of CCN3 and that incubation of osteoblasts with CCN3 promotes focal adhesion kinase (FAK) and Akt phosphorylation. Our results indicate that CCN3 promotes the expression of Runx2 and osterix in osteoblasts by inhibiting miR-608 expression via the FAK and Akt signaling pathways.

SUBMITTER: Chen PC

PROVIDER: S-EPMC6651805 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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CCN3 Facilitates Runx2 and Osterix Expression by Inhibiting miR-608 through PI3K/Akt Signaling in Osteoblasts.

Chen Po-Chun PC Liu Ju-Fang JF Fong Yi-Chin YC Huang Yuan-Lin YL Chao Chia-Chia CC Tang Chih-Hsin CH

International journal of molecular sciences 20190705 13

CCN3, otherwise known as the nephroblastoma overexpressed (NOV) protein, is a cysteine-rich protein that belongs to the CCN family and regulates several cellular functions. Osteoblasts are major bone-forming cells that undergo proliferation, mineralization, renewal, and repair during the bone formation process. We have previously reported that CCN3 increases bone morphogenetic protein 4 (BMP-4) production and bone mineralization in osteoblasts, although the role of CCN3 remains unclear with rega ...[more]

PMID: 31284378

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Project description:Keloids are a skin fibroproliferative condition characterized by the hyperproliferation of fibroblasts and the excessive deposition of extracellular matrix (ECM) components. Previous studies have determined that Caveolin‑1 controlled hyperresponsiveness to mechanical stimuli through Runt‑related transcription factor 2 (Runx2) activation in keloids. However, the molecular mechanism of Runx2 regulating the pathological progression of keloids has not been elucidated. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that most of the differentially expressed genes (DEGs), including Runx2, were significantly enriched in the biological processes 'Positive regulation of cell proliferation', in the cellular components 'Extracellular matrix', in the molecular functions 'Extracellular matrix structural constituents' and in the KEGG 'PI3K‑Akt signaling pathway'. The aim of the present study was to investigate the expression levels of the Runx2 in human keloid tissues and primary human keloid fibroblasts (HKFs), and to determine the underlying molecular mechanisms involved in the fibrotic roles of Runx2 in keloid formation. Runx2 expression levels were analyzed in patient keloid tissues and HKFs using western blotting, reverse transcription‑quantitative PCR (RT‑qPCR) and immunofluorescence microscopy. Primary HKFs were transfected with a small interfering RNA (si) specifically targeting Runx2 (si‑Runx2). Subsequently, Cell Counting Kit‑8, wound healing and Transwell assays, flow cytometry, RT‑qPCR and western blotting were applied to evaluate the proliferation, migration, apoptosis, ECM deposition and PI3K/AKT signaling pathway of HKFs, respectively. In addition, western blotting was also used to determine the expression levels of phosphorylated AKT and PI3K in HKFs. The results revealed that Runx2 expression levels were upregulated in keloid tissues and primary HKFs compared with the normal skin tissues and human normal fibroblasts. Following the transfection with si‑Runx2, the proliferative and migratory abilities of HKFs were significantly reduced and the apoptotic rate was increased. The expression levels of type I, type III collagen, fibronectin, and α‑smooth muscle actin were downregulated in si‑Runx2‑transfected cells, which was hypothesized to occur through following the downregulation of the phosphorylation levels of PI3K and AKT. In conclusion, the findings of the present study indicated that Runx2 silencing in HKFs might significantly inhibit the cell proliferation, migration and the expression levels of ECM‑related proteins, and promote apoptosis via suppressing the PI3K/AKT signaling pathway. Thus, Runx2 siRNA treatment may reverse the pathological phenotype of keloids through the inhibition of PI3K/AKT signaling in patients.

Dataset Information

CCN3 Facilitates Runx2 and Osterix Expression by Inhibiting miR-608 through PI3K/Akt Signaling in Osteoblasts.

Publications

CCN3 Facilitates Runx2 and Osterix Expression by Inhibiting miR-608 through PI3K/Akt Signaling in Osteoblasts.

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