The roles of IL-19 and IL-20 in the inflammation of degenerative lumbar spondylolisthesis

Abstract

Background: Degenerative lumbar spondylolisthesis (DLS) is a major cause of spinal canal stenosis and is often related to lower back pain. IL-20 is emerging as a potent angiogenic, chemotactic, and proinflammatory cytokine related to several chronic inflammatory bone disorders likes intervertebral disc herniation, rheumatoid arthritis (RA), osteoporosis, and bone fracture. IL-19 also acts as a proinflammatory cytokine in RA. The aim of the present study was to investigate whether IL-19 and IL-20 are involved in DLS and compare three different tissues including disc, facet joint, and ligamentum flavum of patients with DLS to verify which tissue is affected more by inflammation.

Methods: Disc, facet joint and ligamentum flavum from 13 patients with DLS was retrieved, and the expression pattern of IL-19, IL-20, IL-20R1, IL-20R2, TNF-α, IL-1β, and MCP-1 was evaluated using immunohistochemical staining with specific antibodies. The disc cells were isolated and incubated with IL-19 and IL-20 under CoCl₂-mimicked hypoxic conditions to analyze the proinflammatory cytokine expression pattern using real-time quantitative PCR with specific primers.

Results: IL-19 and IL-20 were positively stained and accompanied by abundant expression of TNF-α, IL-1β, and MCP-1 in facet joints of DLS patients. IL-19 and IL-20's receptors (IL-20R1 and IL-20R2) were expressed on chondrocytes and fibrocytes/fibroblasts in facet joint and ligamentum flavum tissues from patients with DLS. There was a significant correlation between the expression of IL-20 and IL-1β in facet joint. In vitro assay, IL-19 and IL-20 upregulated the expression of IL-1β, IL-6, TNF-α, IL-8, VEGF, and MCP-1 in primary cultured DLS disc cells under CoCl₂-mimicked hypoxic conditions.

Conclusions: IL-19, IL-20, and their receptors as well as proinflammatory cytokines (TNF-α, IL-1β, and MCP-1) were expressed more in facet joints than the other tissues in patients with DLS; therefore, the etiology of inflammation might be more facet-centric. IL-19 and IL-20 induced proinflammatory cytokine expression in disc cells and might play a role in the pathogenesis of DLS.

Keywords: Degenerative lumbar Spondylolisthesis; IL-19; IL-20; Inflammation.

Fig. 1

Expression of IL-19, IL-20, IL-20R1, IL-20R2, IL-1β, TNF-α, and MCP-1 in DLS patients. Micrographs of IHC staining of cytokines and chemokines (IL-19, IL-20, IL-20R1, IL-20R2, IL-1β, TNF-α, and MCP-1) with specific antibodies on (a) Disc, (b) Facet joint, and (c) Ligamentum flavum tissue samples from patients with DLS (Disc, N = 6; Facet joint, N = 12; Ligamentum flavum, N = 13). Chondrocytes were positively stained. The boxed area shows positively stained fibrocytes and fibroblasts (original magnification ×200). Staining with isotype mouse IgG was used as the negative control. All experiments were performed three times with similar results. Data are from a representative experiment.

Fig. 2

Expression of IL-19, IL-20, and their receptors in DLS patients and HIVD patients. Data comparison of positive staining results of IL-19, IL-20, IL-20R1, and IL-20R2 in disc tissues from DLS patients and HIVD patients. Expression of (a) IL-19, (b) IL-20, (c) IL-20R1, (d) IL-20R2 were analyzed by using IHC staining in disc tissues from DLS (n = 6) or HIVD (n = 20) patients (data of HIVD patients came from [22])

Fig. 3

IL-19 and IL-20 induced cytokine and chemokine expression in DLS disc cells. Primary disc cells isolated from DLS patients were incubated with IL-19 or IL-20 in CoCl₂-mimicked hypoxia condition. The expression of cytokines and chemokines was then analyzed using real-time PCR with primers specific for (a) IL-1β, (b) IL-6, (c) IL-8, (d) TNF-α, (e) VEGF, and (f) MCP-1. The relative quantification of PCR products was expressed as 2^−ΔΔCT, normalized using GAPDH expression and relative to the levels of PBS-treated disc cells, data statistics were done by one way ANOVA, * p < 0.05; **: p < 0.01; ***: p < 0.001