Renal interstitial fibrosis is the main factor determining chronic kidney disease (CKD) progression, and renal tubular epithelial cells are the key drivers of this pathological process. Herein, we revealed significantly increased ubiquitin-specific peptidase 10 (USP10) expression in the kidney tissues of both patients with CKD and mice induced by unilateral ureteral obstruction, as well as in transforming growth factor-beta 1 (TGFβ1)-induced renal tubular epithelial cells. In vivo, treatment with the USP10 small molecule inhibitor Spautin-1, which inhibits its deubiquitinating activity, weakened renal interstitial fibrosis progression and alleviated the subsequent inflammatory response and oxidative stress in male mice. In vitro, knocking down USP10 or inhibiting its deubiquitinating activity through Spautin-1 significantly reduced fibronectin expression and ameliorated TGFβ1-induced renal tubular epithelial cell dedifferentiation. Additionally, our results revealed that USP10 directly binds to P53 and removes the K48-linked polyubiquitin chains from P53, thereby affecting its ubiquitination, stability, and nuclear translocation, which subsequently leads to the upregulation of P21 and promotes fibrotic gene expression in injured renal tubular epithelial cells, ultimately exacerbating renal interstitial fibrosis. In conclusion, USP10 is inhibited through the P53 signaling pathway to alleviate the progression of renal interstitial fibrosis and serve as a potential target for treating CKD.