TGF-β–mediated epithelial–mesenchymal transition of keratinocytes promotes fibrosis in secondary lymphedema

Secondary lymphedema is characterized by fibrosis and impaired lymphatic function. Although TGF-β is a key regulator of fibrosis in this disease, the cellular mechanisms regulating this process remain unknown. Epithelial-mesenchymal transition(EMT), a mechanism by which TGF-β induces fibrosis in other skin diseases, is characterized by loss of epithelial cell markers and cellular polarity, upregulation of fibrotic gene expression, and gain of migratory capacity. Using clinical lymphedema biopsy specimens and animal models, we show that keratinocytes in the basal layer of epidermis undergo EMT in lymphedematous skin, migrate into the dermis, and contribute to dermal fibrosis. In vitro studies using cultured primary human keratinocytes treated with lymphatic fluid from the affected limbs of patients with secondary lymphedema resulted in a TGF-β-mediated increased expression of EMT markers. We show for the first time that EMT is activated by TGF-β in secondary lymphedema and that this process plays an important role in regulating skin fibrosis in this disease. RNAseq profiling of four pairs of frozen clinical lymphedema biopsy specimens. The ribodepletion method was used for RNAseq. mRNA expression was standardized and analyzed. Standardized expression for each molecule was assessed and data are presented as Z-scores.