Determination of Anti-Fibrotic Effects of Possible Scar-Collagen Antagonists on TGF-β1 Treated Dermal Fibroblasts

High levels of collagen being produced by cells in response to wounding is scarring. Scarring, or fibrosis, is a major medical science problem because of the lack of functionality of the affected organ. To reenact collagen levels during scarring, primary dermal fibroblasts were treated with TGF-β1 before the addition of test compounds. Transforming growth factor beta I (TGF-β1), a cytokine, is a known inducer of scar collagen production. Anti-fibrotic effects of test compounds were screened and evaluated in vitro by measuring amounts of collagen produced at the protein level. To measure secreted collagen as part of the extra-cellular matrix, monoclonal antibodies against collagen I and III were used. Collagen I is the most abundant type in the human body, type III is present along with type I in connective tissue, blood vessels, and skin. Anti-fibrotic effects were measured in a fluorescent microplate assay for secreted collagen I and III, and confirmed by confocal microscopy for collagen I. MTT assays confirmed anti-collagen effects from the test compounds were not due to decreased cell viability. From the results of these tests epigallocatechin gallate (EGCG), and nintedanib show promising decreases in collagen production in comparison to the control, illustrating the usefulness of the assay for screening scar antagonists