Background Systemic sclerosis (SSc) is an autoimmune disease characterised by fibrosis, vascular dysfunction and abnormal activation of immune cells including monocytes. It was suggested that monocytes along with fibroblasts play an important role in the production of profibrotic factors such as IL-6 and TIMP-1 (tissue-inhibitor of metalloproteinase-1). TIMPs are specific inhibitors of matrix metalloproteinases (MMPs) regulating extracellular matrix (ECM) turnover. Importantly, the balance between TIMPs and MMPs is altered in most pathological stages including SSc and is associated with abnormal ECM formation. However, the exact factors which drive both profibrotic TIMP-1 and IL-6 secretion have not been fully established.
Objectives We aim to test whether circulating monocytes from SSc patients and dermal fibroblasts produce TIMP-1 and IL-6 in response to TLR activation, which contributes to excessive matrix deposition and consequently disease progression.
Methods 23 patients with SSc and 19 HC (healthy control) were included in this study. Peripheral blood monocytes were further separated by CD14+ microbeads. Production of TIMP-1, IL-6 by monocytes and dermal fibroblasts was determined by ELISA, qRT-PCR or functional assay in response to either panel of conventional TLR agonists or Danger Associated Molecular Patterns (DAMPs) such as High-Mobility Group Box-1 protein (HMGB-1) and hyaluronic acid (HA).
Results TIMP-1 produced by monocytes was elevated in SSc patients compared to HC as measured by qRT-PCR (fold increase 3.1, p=0.03) and ELISA (fold increase 3.2, p=0.02). The level of TIMP-1 present in SSc sera patients was also higher than in the HC (fold increase 1.8, p<0.001). In addition, IL-6 and TIMP-1 expression was significantly stronger when SSc and HC monocytes were stimulated with TLR4 (LPS) or TLR8 (ss-RNA) agonists, but the response was more pronounced in SSc monocytes. Agonists against other TLRs were less effective in the induction of TIMP-1 and IL-6. Furthermore, SSc and HC monocytes stimulated with HMGB-1 alone did not induce the secretion of profibrotic factors. However, combination of HMGB-1 and LPS upregulated the expression of both IL-6 and TIMP-1. These results suggest that there is the synergic effect between LPS and HMGB-1 on monocytes and fibroblasts activation. Interestingly, HC and SSc monocytes stimulated with HA alone or in the combination with LPS upregulated TIMP-1 and IL-6 at the same level as HA stimulation only. Furthermore, matrix assay of TLR8 stimulated monocytes also confirmed functional TIMP-1 secretion, as matrix metalloproteinase-1 activity was significantly inhibited.
Conclusions This study indicates a potential link between TLR signaling and excessive TIMP-1 and IL-6 secretion in circulating monocytes from SSc patients and activated dermal fibroblasts, supporting an important role of these cells in production of pro-fibrotic factors.
Disclosure of Interest None Declared
Statistics from Altmetric.com
If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.