*Purpose: One of the possible steps in the development of post-transplant interstitial fibrosis is epithelial to mesenchymal transition (EMT) of proximal renal tubular cells with concomitant release of pro-fibrotic factors. One of the hallmarks of EMT is de novo expression of vimentin, an intermediate filament protein that forms the basis of the mesenchyme cytoskeleton. Withaferin A (WFA), a steroidal lactone, has been shown to inhibit vimentin filament assembly. Therefore, we aimed to determine whether WFA could ameliorate the development of fibrosis after renal injury.

*Methods: Withaferin A was administered via intraperitoneal injection to 129svs mice following unilateral ischemia/reperfusion injury (30 minutes ischemia). Kidneys, and sera were then obtained for analysis via western blotting and qPCR analyses. In addition, kidney samples were analyzed with both Masson’s Trichrome and Pico Sirius staining to determine collagen deposition. Additionally, Withaferin A was added to cultured HK-2 cells following EMT stimulation by addition of TGF-β. Assembled, and soluble vimentin concentrations were determined following detergent extraction. Finally, time lapse phase contrast microscopy was used to visualize wound healing following TGF-β induction. Quantification of cellular migration was carried out by measuring the changes in the wound area after 24 hours.

*Results: Immunofluorescence of mouse kidneys following treatment with WFA showed decreased vimentin expression 3 weeks after injury. Pico Sirius Red staining showed decreased collagen deposition in mice treated with WFA. The addition of WFA to HK-2 cells in wound healing assays under TGF-β exposure results in very little motility when compared to controls.

*Conclusions: We conclude that WFA inhibits vimentin filament assembly, thus inhibiting EMT and slowing the development of IFTA. Future experiments will look at the mechanism behind WFA mediated inhibition of vimentin assembly as well as the difference in functional outputs such as GFR and creatinine in mice treated with WFA.

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