Targeting TGF-β1/miR-21 Pathway in Keratinocytes Reveals Protective Effects of Silymarin on Imiquimod-Induced Psoriasis Mouse Model

Epidermal cells integrate multiple signals that activate the signaling pathways involved in skin homeostasis. TGF-β1 signaling pathway upregulates microRNA (miR)-21-5p in keratinocytes and is often deregulated in skin diseases. To identify the bioactive compounds that enable to modulate the TGF-β1/miR-21-5p signaling pathway, we screened a library of medicinal plant extracts using our miR-ON RILES luciferase reporter system placed under the control of the miR-21-5p in keratinocytes treated with TGF-β1. We identified silymarin, a mixture of flavonolignans extracted from Silybum marianum (L.) Gaertn., as the most potent regulator of miR-21-5p expression. Using Argonaute 2 immunoprecipitation and RT-qPCR, we showed that silymarin regulates the expression of miR-21-5p through a noncanonical TGF-β1 signaling pathway, whereas RNA-sequencing analysis revealed three unexpected transcriptomic signatures associated with keratinocyte differentiation, cell cycle, and lipid metabolism. Mechanistically, we demonstrated that SM blocks cell cycle progression, inhibits keratinocyte differentiation through repression of Notch3 expression, stimulates lipid synthesis via activation of PPARγ signaling and inhibits inflammatory responses by suppressing the transcriptional activity of NF-κB. We finally showed that topical application of silymarin alleviates the development of imiquimod-induced psoriasiform lesions in mice by abrogating the altered expression levels of markers involved in inflammation, proliferation, differentiation, and lipid metabolism.