2495: IL-6/STAT3 Signaling is Central for Chemokine Production in E2-Induced Dermal Inflammation

Background/Purpose: Scleroderma (SSc) is an autoimmune disease characterized by dermal inflammation and fibrosis. Patients with SSc have high systemic levels of estradiol (E2), a form of estrogen, compared to age-matched controls, and worse mortality. E2 also contributes to dermal fibrosis in SSc, but we are unsure of its role in dermal inflammation. Using bulk RNA sequencing (RNA seq), we investigated inflammatory pathways in E2-treated dermal tissue.

Methods: We isolated total RNA from human skin tissue treated with ethanol (ETOH, vehicle) or E2 for 48 hours for bulk RNA seq. Using bioinformatics software, we identified statistically significant inflammatory genes and pathways. We confirmed transcript and protein levels using qPCR and ELISA, respectively. Primary human skin dermal fibroblasts from healthy donors treated with ETOH or E2, and dermal fibroblasts from patients with SSc were used for mechanistic studies. Statistical significance was defined as p< 0.05.

Results: We detected and confirmed that CXCL2, CXCL3, CXCL5, CXCL8, and CCL20 chemokines are significantly increased in E2 treated skin tissue compared to ETOH. Additionally, these chemokines are induced by E2 in primary human dermal fibroblasts, with E2-induced CXCL5 and CCL20 transcript and secreted protein levels dependent on the MAPK pathway. We detected an association between E2-induced IL-6 and CXCL8 and CCL20 ex vivo transcripts. Using inhibitors to prevent IL-6 and STAT3 signaling prior to E2 stimulation prevented E2-induced CXCL2, CXCL3, CXCL8, and CCL20 transcripts ex vivo. While SSc dermal fibroblasts treated with anastrozole to prevent E2 generation produced significantly less CXCL3, CXCL5 and CXCL8, but not CXCL2 or CCL20 transcripts, treatment with bazedoxifene, a selective estrogen receptor modulator and IL-6/STAT3 inhibitor, produced less CXCL5 and CXCL8 secreted protein.

Conclusion: E2 increases CXCL2, CXCL3, CXCL5, CXCL8, and CCL20 chemokines in dermal tissue and fibroblasts. Although the MAPK pathway is important in their E2-induced expression, IL-6/STAT3 signaling is paramount to E2-induced chemokine expression. Our data suggests E2 production and estrogen receptor signaling through IL-6/STAT3 signaling is responsible, in part, for increased CXCL5 and CXCL8 in SSc. Therefore, E2 augments highly active inflammatory pathways in SSc.