Steroid treatment can inhibit nuclear localization of members of the NF-κB pathway in human disc cells stimulated with TNF-α.

Abstract

Steroid applications are able to repress inflammatory activity in various conditions, including herniation of the nucleus pulposus (HNP), by inhibiting tumour necrosis factor (TNF)-α, but the effects of long-term use are unknown. Here, we investigated the effect of dexamethasone (DEXA) on TNF-α-stimulated intervertebral disc cells by monitoring the expression and localization of NF-κB in the cytoplasm and nucleus. Cultured human intervertebral disc cells were left untreated or treated with only TNF-α, only DEXA, or with TNF-α and DEXA simultaneously. Cytoplasmic and nuclear proteins were extracted and Western blotted after 10 min, 1 or 2 h, to evaluate the expression of p50, p65, p52, and p100 (components of NF-κB). Immunofluorescence analysis was used to determine the subcellular localization of the proteins at 1 h. DEXA had limited effects on NF-κB expression in TNF-α-stimulated disc cells within the first 10 min. At 1 h, DEXA prevented the TNF-α-stimulated translocation of p50, p52, and p65. After 2 h, DEXA reduced the nuclear expression of p50, p65, and p52. Thus, DEXA resulted in delayed expression of NF-κB components and inhibited the translocation of p50, p52, and p65 to the nucleus, which would prevent expression of the corresponding genes. Therefore, following stimulation with TNF-α, transcriptional regulation of NF-κB in disc cells is mainly mediated via the classical pathway, but also to some extent via the alternative pathway. Hence, blockade of sub-acute inflammatory changes in HNP can be achieved by early injection of steroids, whereas long-term injection of a steroid may initiate NF-κB autophosphorylation.