Diabetic kidney disease (DKD) is one of the complications of diabetes mellitus, which triggers kidney fibrosis and eventually develops into end-stage renal disease. Nuciferine (NF) is one of the most important functional components in lotus leaves (LL), but its role and mechanism for the treatment of DKD are unclear. A high-fat-diet (HFD)-induced DKD model in KK-AY mice was established in this study. NF treatment significantly improved blood glucose and blood biochemical indices in DKD mice. Furthermore, NF reduced the levels of mALB, UCRE, Scr, and BUN in mice urine. Further, the extent of renal lesions in the mice in this study was at stage IV according to the Mogensen staging method. NF treatment was effective in ameliorating renal injury during this period. Concurrently, the protein levels of FN, N-cadherin, TGFβ, p-Smad3, p-PI3K, p-AKT, p-mTOR, and p62 were decreased. In contrast, the level of expression of Beclin-1 was increased. In the high glucose-exposed HK-2 cell model, the expression of p-PI3K, p-AKT, and p-mTOR was all downregulated, and autophagy proteins were increased after NF intervention. In addition, HK-2 cells were treated with high glucose in combination with Wortmannin and 3-MA, respectively. The results demonstrated that NF inhibited the expression of TGFβ and p-Smad3 by regulating autophagy through the PI3K-AKT-mTOR pathway, thereby ameliorating renal fibrosis at stage IV in mice. Therefore, LL can be used as a dietary component for the prevention of renal fibrosis in DKD patients.