Interleukin-1 (IL-1) is known to be involved in cartilage degeneration following joint injury or due to osteoarthritis. In the present study, we explored the effects of miR-150 on IL-1-stimulated human chondrogenic cells ATDC5.
ATDC5 cells were transfected with the mimic, inhibitor or negative controls specific for miR-150, and subsequently treated by IL-1. CCK8 assay, PI and FITC-conjugated Annexin V double-staining, Western blot, qRT-PCR and ELISA assay were performed to determine the changes of cell viability, apoptosis, and the release of pro-inflammatory cytokines. Targeting relationship between miR-150 and KLF2 was detected by dual luciferase activity assay.
IL-1 reduced cell viability, induced apoptosis, and enhanced the expression and release of pro-inflammatory cytokines (IL-6, IL-8 and TNF-α) in ATDC5 cells. IL-1 also increased the expression of miR-150. Suppression of miR-150 alleviated IL-1-induced cell damage in ATDC5 cells, while overexpression of miR-150 resulted in an opposite impact. KLF2 was negatively regulated by miR-150, and it was proved as a target gene of miR-150. KLF2 overexpression exhibited protective actions in IL-1-injured ATDC5 cells, even if miR-150 was suppressed in cell. Moreover, IL-1-induced activation of NF-kB and Notch pathways was alleviated by KLF2 overexpression.
Suppression of miR-150 led to up-regulation of KLF2, which in turn protected ATDC5 cells against IL-1-induced injury.