Dataset Information

CCN family member 2/connective tissue growth factor (CCN2/CTGF) is regulated by Wnt-β-catenin signaling in nucleus pulposus cells.

ABSTRACT:

Background

The aims of this study were to investigate the gene expression of CCN family members in rat intervertebral disc (IVD) cells and to examine whether Wnt-β-catenin signaling regulates the expression of CCN family 2 (CCN2)/connective tissue growth factor (CTGF) in rat nucleus pulposus (NP) cells.

Methods

The gene expression of CCN family members were assessed in rat IVD cells using real-time reverse transcription polymerase chain reaction (RT-PCR). The expression pattern of CCN2 was also assessed in rat IVD cells using western blot and immunohistochemical analyses. Gain-of-function and loss-of-function experiments were performed to identify the mechanisms by which Wnt-β-catenin signaling influences the activity of the CCN2 promoter. To further determine if the mitogen-activated protein kinase (MAPK) pathway is required for the Wnt-β-catenin signaling-induced regulation of CCN2 expression in the NP cells, CCN2 expression was analyzed by reporter assay, RT-PCR and western blot analysis.

Results

CCN2 messenger RNA (mRNA) and protein were expressed in rat IVDs. Expression of CCN2 was significantly higher than for mRNA of other CCN family members in both rat NP and annulus fibrosus (AF) cells. The relative activity of the CCN2 promoter decreased 24 h after treatment with 6-bromoindirubin-3'-oxime (1.0 μM) (0.773 (95% 0.735, 0.812) P = 0.0077) in NP cells. In addition, treatment with the WT-β-catenin vector (500 ng) significantly decreased CCN2 promoter activity (0.688 (95% 0.535, 0.842) P = 0.0063), whereas β-catenin small interfering RNA (500 ng) significantly increased CCN2 promoter activity (1.775 (95% 1.435, 2.115) P < 0.001). Activation of Wnt-β-catenin signaling decreased the expression of CCN2 mRNA and protein by NP cells. Regulation of CCN2 by Wnt-β-catenin signaling involved the MAPK pathway in rat NP cells.

Conclusions

This study shows that Wnt-β-catenin signaling regulates the expression of CCN2 through the MAPK pathway in NP cells. Understanding the balance between Wnt-β-catenin signaling and CCN2 is necessary for developing therapeutic alternatives for the treatment of IVD degeneration.

SUBMITTER: Hiyama A

PROVIDER: S-EPMC6162946 | biostudies-literature |

REPOSITORIES: biostudies-literature

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