IL6/sIL6R regulates TNFα-inflammatory response in synovial fibroblasts through modulation of transcriptional and post-transcriptional mechanisms

Abstract

Introduction The clinical efficacy of specific interleukin-6 inhibitors has confirmed the central role of IL6 in rheumatoid arthritis (RA). However the local role of IL6, in particular in synovial fibroblasts (SF) as a direct cellular target to IL6/sIL6R signal is not well characterized. The purpose of the study was to characterize the crosstalk between TNFα and IL6/sIL6R signaling to the effector pro-inflammatory response of SF. Methods SF lines were stimulated with either TNFα, IL6/sIL6R, or both together, for the time and dose indicated for each experiment, and where indicated, cells were treated with inhibitors actinomycin D, adalimumab, ruxolitinib and cycloheximide. mRNA expression of cytokines, chemokines and matrix metalloproteases (MMPs) were analyzed by quantitative RT-PCR. Level of IL8/CXCL8 and CCL8 in culture supernatants was measured by ELISA. Mononuclear and polymorphonuclear cells migration assays were assessed by transwell using conditioned medium from SF cultures. Statistical analyses were performed as indicated in the corresponding figure legends and a p-value < 0.05 was considered statistically significant. Results The stimulation of SF with IL6/sIL6R and TNFα, cooperatively promotes the expression of mono- and lymphocytic chemokines such as IL6, CCL8 and CCL2, as well as matrix degrading enzymes such as MMP1, while inhibiting the induction of central neutrophil chemokines such as IL8/CXCL8. These changes in the pattern of chemokines expression resulted in reduced polymorphonuclear (PMN) and increased mononuclear cells (MNC) chemoattraction by SF. Mechanistic analyses of the temporal expression of genes demonstrated that the cooperative regulation mediated by these two factors is mostly induced through de novo transcriptional mechanisms activated by IL6/sIL6R. Furthermore, we also demonstrate that TNFα and IL6/sIL6R cooperation is partially mediated by the expression of secondary factors signaling through JAK/STAT pathways. Conclusions These results point out to a highly orchestrated response to IL6 in TNFα-induced SF and provide additional insights into the role of IL6/sIL6R in the context of RA, highlighting the contribution of IL6/sIL6R to the interplay of SF with other inflammatory cells.

Distinct stromal and immune cell interactions shape the pathogenesis of rheumatoid and psoriatic arthritis

“…Importantly, the combined treatment with TGF-β and IL-1β resulted in a significant (**p=0.0047) increase in ICAM-1 expression compared with IL-1β only treated synovial fibroblasts (figure 8A). Fibroblasts are the main source of IL-6 in RA with pro-inflammatory synovial fibroblast secreting high levels of IL-6 in response to TNF-α 6 37. Similarly to the expression of ICAM-1, treatment of synovial fibroblasts with TGF-β did not lead to increased IL-6 secretion compared with untreated synovial fibroblasts, however the combined treatment with TGF-β and IL-1β resulted in a significant (**p=0.0018) increase in IL-6 compared with IL-1β only treated synovial fibroblasts (figure 8B).…”

The Regulatory Effect of Coaggregation Between Fusobacterium nucleatum and Streptococcus gordonii on the Synergistic Virulence to Human Gingival Epithelial Cells

“…A previous study showed MK2 was involved in regulating the TNF-induced expression of IL-8 by p38 MAPK in human lung microvascular endothelial cells at a post-transcriptional level (Su et al, 2008). Another study showed the stimulation of synovial fibroblasts with IL-6 and TNF-a cooperatively inhibited the induction of IL-8 (Valin et al, 2020). It was speculated that a more complex mechanism in IL-8 secretion existed induced by coaggregation of F. nucleatum subsp.…”

Osteoarticular Involvement-Associated Biomarkers and Pathways in Psoriasis: The Shared Pathway With Ankylosing Spondylitis

“…PsA is a chronic immune-mediated rheumatic disease. Studies have reported that the posttranscriptional regulation of gene expression plays a vital role in rheumatic disease ( 26 , 27 ). However, most studies have focused on microRNA and related pathways, while only a few focused on the mRNA surveillance pathway ( 28 , 29 ).…”