A Positive Feedback Loop Exists between Estradiol and IL-6 and Contributes to Dermal Fibrosis.

Systemic sclerosis (SSc) is characterized by dermal fibrosis with a female predominance, suggesting a hormonal influence. Patients with SSc have elevated interleukin (IL)-6 levels, and post-menopausal women and older men also have high estradiol (E2) levels. In the skin, IL-6 increases the enzymatic activity of aromatase, thereby amplifying the conversion of testosterone to E2. Therefore, we hypothesized that an interplay between E2 and IL-6 contributes to dermal fibrosis. We used primary dermal fibroblasts from healthy donors and patients with diffuse cutaneous (dc)SSc, and healthy donor skin tissues stimulated with recombinant IL-6 and its soluble receptor (sIL-6R) or E2. Primary human dermal fibroblasts and tissues from healthy donors stimulated with IL-6+sIL-6R produced E2, while E2-stimulated dermal tissues and fibroblasts produced IL-6. Primary dermal fibroblasts from healthy donors treated with IL-6+sIL-6R and the aromatase inhibitor anastrozole (ANA) and dcSSc fibroblasts treated with ANA produced less fibronectin (FN), type III collagen A1 (Col IIIA1), and type V collagen A1 (Col VA1). Finally, dcSSc dermal fibroblasts treated with the estrogen receptor inhibitor fulvestrant also generated less FN, Col IIIA1, and Col VA1. Our data show that IL-6 exerts its pro-fibrotic influence in human skin in part through E2 and establish a positive feedback loop between E2 and IL-6.

The Relationship between Time, Race, and Estrogen Receptor Alpha in Estradiol-Induced Dermal Fibrosis.

In the skin, estradiol (E2) promotes profibrotic and proinflammatory cytokines, contributing to extracellular matrix (ECM) deposition. However, the magnitude of the response differs. Using the human skin organ culture model, we evaluated donor characteristics and correlations that contribute to E2-induced interleukin-6 (IL-6), transforming growth factor beta 1 and 2 (TGFB1 and TGFB2), collagen IA2 (Col IA2), collagen IIIA1 (Col IIIA1), and fibronectin (FN) expressions. In vehicle- and E2-treated dermal skin tissue transcripts, we confirm differences in the magnitude; however, there were positive correlations between profibrotic mediators and ECM components 48 h after E2 treatment. Also, positive correlations exist between baseline and E2-induced TGFB1, IL-6, Col IIIA1, and FN transcripts. Since estrogen receptor alpha (ERA) can propagate E2's signal, we measured and detected differences in its baseline and fold change transcript levels, with a significant decline in baseline levels 48 h after incubation and an increase 48 h after E2 treatment. There was a trend to higher transcript levels in African American donors 24 h earlier. Finally, E2-induced ERA transcript levels negatively correlated with its own baseline levels and positively correlated with FN, TGFB1, and Col IA2 transcript levels. Therefore, our data suggest ERA, E2 exposure time, and race/ethnicity contribute to E2-induced dermal fibrosis.

Elucidating the cellular mechanism for E2-induced dermal fibrosis.

BACKGROUND: Both TGFß and estradiol (E2), a form of estrogen, are pro-fibrotic in the skin. In the connective tissue disease, systemic sclerosis (SSc), both TGFß and E2 are likely pathogenic. Yet the regulation of TGFß in E2-induced dermal fibrosis remains ill-defined. Elucidating those regulatory mechanisms will improve the understanding of fibrotic disease pathogenesis and set the stage for developing potential therapeutics. Using E2-stimulated primary human dermal fibroblasts in vitro and human skin tissue ex vivo, we identified the important regulatory proteins for TGFß and investigated the extracellular matrix (ECM) components that are directly stimulated by E2-induced TGFß signaling. METHODS: We used primary human dermal fibroblasts in vitro and human skin tissue ex vivo stimulated with E2 or vehicle (ethanol) to measure TGFß1 and TGFß2 levels using quantitative PCR (qPCR). To identify the necessary cell signaling proteins in E2-induced TGFß1 and TGFß2 transcription, human dermal fibroblasts were pre-treated with an inhibitor of the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway, U0126. Finally, human skin tissue ex vivo was pre-treated with SB-431542, a TGFß receptor inhibitor, and ICI 182,780, an estrogen receptor α (ERα) inhibitor, to establish the effects of TGFß and ERα signaling on E2-induced collagen 22A1 (Col22A1) transcription. RESULTS: We found that expression of TGFß1, TGFß2, and Col22A1, a TGFß-responsive gene, is induced in response to E2 stimulation. Mechanistically, Col22A1 induction was blocked by SB-431542 and ICI 182,780 despite E2 stimulation. Additionally, inhibiting E2-induced ERK/MAPK activation and early growth response 1 (EGR1) transcription prevents the E2-induced increase in TGFß1 and TGFß2 transcription and translation. CONCLUSIONS: We conclude that E2-induced dermal fibrosis occurs in part through induction of TGFß1, 2, and Col22A1, which is regulated through EGR1 and the MAPK pathway. Thus, blocking estrogen signaling and/or production may be a novel therapeutic option in pro-fibrotic diseases.