Role of nuclear factor of activated T-cells in regulating ADAMTS-4 promoter activity in nucleus pulposus cells / 第二军医大学学报

ABSTRACT

Objective: To observe the regulatory effect of nuclear factor of activated T-cells (NFAT)-1, 4, 5 on ADAMTS-4. promoter activity in nucleus pulposus cells, so as to discuss the underlying molecular mechanism of intervertebral disc degeneration. Methods: The ADAMTS-4-pβ-gal-Basic was inserted into PGL3-Basic vector after double digestion by the restriction enzyme Xho I and Hind III. The purified PGL3-ADAMTS-4 plasmid was obtained through screening. Rat nucleus pulposus cells were cultured in vitro and, via Lipofectamine2000 transfection reagent, transfected with NFAT-1, NFAT-4, NFAT-5 or DN-TonEBP expressing plasmids with or without appropriate backbone vector and 175 ng ADAMTS-4 promoter. The normal cells were taken as controls and transfected cells were taken as treatment group. Dual-Luciferase™ reporter assay system was used to detect the effect of NFAT-1, 4, 5(TonEBP) on ADAMTS-4 promoter activity. Results: We successfully constructed PGL3-ADAMTS-4 plasmid and identified two NFAT-Bind elements in the promoter. Compared with the control group, ADAMTS-4 promoter activity was significantly inhibited in NFAT-1 group (P 0. 05). Conclusion: NFAT-1 can regulate ADAMTS-4 expression, which may play a role in modulating aggrecan content in the intervertebral disc physiology and/or intervertebral disc degeneration, providing a new strategy for biological treatment of intervertebral disc degeneration.