Background/Purpose
Autotaxin (ATX) is an enzyme present in biological fluids that is responsible for the production of the lipid mediator, lysophosphatidic acid (LPA). We previously implicated LPA and its receptor, LPA1 in SSc pathogenesis.1 Here we studied the role of ATX in SSc dermal fibrosis using the bleomycin (BLM) mouse model and skin biopsy samples from SSc patients and healthy controls. We evaluated the role of IL-6, a cytokine implicated in SSc, in mediating ATX-induced fibrosis. Additionally, we investigated the therapeutic potential of targeting ATX, by using a novel ATX inhibitor, PAT-048 in this model.
Methods
BLM or saline (PBS) was administered subcutaneously to C57Bl/6 mice daily for 3, 7, 14 and 28 days. 6mm dermal punch biopsies were obtained and ATX levels were measured by qPCR and ELISA. ATX inhibition with PAT-048 (20mg/kg oral gavage daily) was assessed in the model. PAT-048 was administered concurrently with BLM or PBS for 28 days, or initiated at 7 or 14 days after BLM. Dermal thickness was measured using H&E-stained sections. Collagen was visualized by Masson’s trichrome stain, and quantified by hydroxyproline measurement. Skin IL-6 expression was evaluated by immunohistochemistry (IHC). The effect of LPA-induced ATX expression was tested on human dermal fibroblasts transfected with IL-6 siRNA. Additionally, healthy and SSc dermal fibroblasts were stimulated with LPA and IL-6 in vitro, and IL-6 and ATX induction were evaluated by ELISA, respectively. Skin ATX expression was measured in SSc patients and healthy controls by qPCR, and IL-6 expression by IHC.
Results
ATX expression at both the mRNA and protein level was increased at Day 3 after BLM injection (3-fold increase, p=0.05) suggesting a role for ATX early in fibrosis. Treatment with PAT-048 attenuated BLM-induced dermal fibrosis in all treatment groups (50% reduction, Day 28, p=0.01), and reduced IL-6 expression in the dermis. In vitro studies of human dermal fibroblasts showed that LPA-induced ATX expression was attenuated with siRNA knock-down of IL-6 (65% reduction, p<0.05). SSc fibroblasts demonstrated increased LPA-induced IL-6 expression, and increased IL-6-induced ATX expression, compared to healthy fibroblasts. Furthermore, ATX expression was increased in SSc skin (n=7) compared to healthy controls (n=5; 3-fold increase, p=0.006) and IL-6 expression by IHC was increased in SSc skin compared to healthy controls (n=3 per group).
Conclusion
We demonstrate that ATX has an important role in SSc fibrosis. Pharmacologic inhibition of ATX with a novel inhibitor, PAT-048, attenuated dermal fibrosis and IL-6 expression. Knock-down of IL-6 in fibroblasts in vitro abrogated LPA-induced ATX expression, suggesting an autocrine loop for ATX/LPA/IL-6 signaling. Both ATX and IL-6 are increased in SSc skin compared to healthy controls, and LPA-induced IL-6 and IL-6-induced ATX expression are increased in SSc fibroblasts, further supporting an ATX/LPA/IL-6 autocrine loop in SSc. Targeting ATX may thus be an effective new therapeutic strategy for SSc fibrosis.
Reference
Castelino FV et al. Amelioration of dermal fibrosis by genetic deletion or pharmacologic antagonism of LPA1 in a mouse model of scleroderma. Arth Rheum, 2011; 63(5):1405-15.
Disclosure:
F. V. Castelino,
None;
L. M. George,
None;
G. Bain,
PharmaAkea Pharmaceuticals,
3;
L. Goulet,
PharmAkea Pharmaceuticals,
3;
R. Lafyatis,
None;
A. M. Tager,
PharmAkea Pharmaceuticals,
6.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/autotaxin-is-highly-expressed-in-systemic-sclerosis-ssc-skin-mediates-dermal-fibrosis-via-il-6-and-is-a-target-for-ssc-therapy/