Upregulation of FoxO6 in nucleus pulposus cells promotes DNA damage repair via activation of RAD51.

Abstract

DNA damage is an essential risk for intervertebral disc degeneration (IDD). Here, we attempted to uncover the effect of FoxO6 and RAD51 on the DNA damage repair of nucleus pulposus (NP) cells in IDD. We collected the human NP tissues of different degeneration degrees and tested the collagen II, FoxO6, and RAD51 expression. Besides, the IL-1β induced NP cell model was also used to elucidate the degenerative progress in vitro. We used Chromatin immunoprecipitation (ChIP) and luciferase reporter assay to confirm whether the FoxO6 protein could enhance the RAD51 expression by binding to its promoter. The FoxO6 gene was upregulated in NP cells by vectors transfection. Immunofluorescence staining was used to measure the RAD51 and γH2AX foci formation. Besides, the typical NP cell gene expression was analyzed by RT-PCR. Cell proliferation was determined by CCK-8, and the cell cycle distribution was determined by flow cytometry. Like collagen II, FoxO6 and RAD51 expression were all decreased both in the severe degenerated NP tissue and in the IL-1β treated NP cells. Upregulation of FoxO6 gene in NP cells enhanced the RAD51 expression via activating the promoter region and inhibited the DNA damage marker γH2AX formation. FoxO6 upregulation alleviated the loss of collagen II, aggrecan, SOD1, and CAT, and suppressed the increase of collagen I/X, TNF-α, and IL-1β expression, which was affected by IL-1β. Besides, FoxO6 also helped the proliferation and cell cycle of NP cells with the activation of RAD51. Upregulation of FoxO6 promotes the DNA repair and maintains the typical phenotype of NP cells, via somehow the mediation of RAD51.