Session Information
Session Type: Abstract Submissions (ACR)
Background/Purpose: IL6 is a key mediator recently implicated in activation of extracellular matrix (ECM) proteins in scleroderma (SSc) fibroblasts. CCL2 is a proinflammatory chemokine that is overexpressed in diffuse cutaneous systemic sclerosis (dcSSc). We explored interaction between these two major mediators and their role in the recruitment of inflammatory cells and fibroblast ECM production.
Methods: Dermal fibroblasts were cultured from skin biopsies from healthy controls (n=4) and early stage dcSSc (n=4). Peripheral blood mononuclear cells (PBMCs) were isolated from blood samples of the latter group. Induction of CCL2 in dermal fibroblasts by IL6 trans-signalling (via sIL-6R) and the ability of SSc fibroblast derived CCL2 to regulate migration of PBMCs across an endothelial layer in vitro was studied using Transwell migration assays in a co-culture system. The effect of PBMCs-fibroblast cross-talk on induction of ECM proteins: α-smooth muscle actin (αSMA) and Collagen type-I (Col-I) was assessed using neutralising antibodies against CCL2 and/or IL6 ligand-receptor axis and targeted ectodomain shedding inhibition using TNF-α processing inhibitor-1(TAPI-1).
Results: IL6 trans-signalling significantly increased CCL2 expression (mean ± SEM % basal expression) (33± 2.7% p<0.03 and 45±5.6% p<0.04) in control and SSc fibroblasts respectively. CCL2 expression was reduced in the presence of anti-IL6R in control fibroblasts (63±6.4%, p<0.04), but the reduction is not significant in the presence of SSc fibroblast IL6 trans-signalling increased migration of PBMCs (n=4) by 2.1 fold (p<0.05) and 4.5 fold (p<0.03) in the presence of control fibroblasts and SSc fibroblasts respectively. The migration of PBMCs was significantly reduced by the addition of neutralising antibodies against CCL2 and IL6R (44± 5.1%, p<0.05 and 62 ± 5.4%, p=0.04) respectively and both antibodies combined (44± 7.3.2%, p<0.05) in the presence of SSc fibroblasts.In response to IL-6 trans-signalling, there was increased expression of ECM proteins: αSMA (53 ± 5.9%, p<0.04) and Col-I (70± 2.6%, p<0.03) at 24-hour in the presence control fibroblasts and αSMA (37± 5.9%, p<0.03) and Col-I (47± 3.6%, p<0.04) in the presence of SSc fibroblasts. Activation of ECM was significantly abrogated by anti-IL6R by αSMA (46± 4.9%, p<0.05) and (58 ± 5.9%, p<0.04) in the presence of control and SSc fibroblasts respectively. TAPI-1 reduced PBMC migration in a concentration dependent manner with maximal effect at 50μM by (55± 4.1 % p<0.04) and TAPI-1 reduced synthesis of αSMA (37± 4.8 %, p<0.05) and Col-I (41± 3.6%, p<0.03) respectively. Together these data support a key role for IL-6 trans-signalling in regulating cell migration in SSc and confirm the potential importance of IL-6 and CCL2 in matrix overproduction
Conclusion: Our data suggests that fibroblast-derived CCL2 expression is in part dependent on IL-6 trans-signalling . The IL-6/CCL2 interplay regulates recruitment of PBMCs and IL-6 trans-signalling with intramembrane shedding of IL-6R mediates the fibrotic response. Thus,CCL2/IL6 interplay may be important in SSc pathogenesis and could be targeted therapeutically.
Disclosure:
R. Alade,
None;
K. Khan,
None;
X. Shiwen,
None;
C. P. Denton,
Roche-Genentech.,
2;
V. Ong,
Roche-Genentech.,
2.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/il6-and-ccl2-co-regulate-fibroblast-dependent-trans-endothelial-migration-of-mononuclear-cells-and-fibrotic-response-in-scleroderma/