Acute kidney injury (AKI) is a common and serious clinical complication with high incidence. Polysaccharides extracted from Flammulina velutipes (FVPs) have been proven to possess anti-inflammatory and antioxidant properties. The present study aimed to investigate the ameliorative effect and mechanism of FVPs on cisplatin (CDPP)-induced AKI. The results of our study revealed that FVPs improved CDPP-induced AKI in mice as indicated by decreasing serum creatinine and urea levels and down-regulating the mRNA expression of IL-6 and TNF-α. Moreover, FVPs modified the composition of gut microorganisms and increased the content of short-chain fatty acids (SCFAs). Additionally, kidney metabolomics analysis demonstrated enrichment of the ferroptosis metabolic pathway. Furthermore, FVPs suppressed ferroptosis as shown by increasing levels of GSH, GPX4, and SLC7A11, while reducing the arachidonic acid level. In conclusion, FVPs were confirmed to ameliorate CDPP-induced AKI in the present study. FVPs can modify the composition of the gut microbiota to promote the production of SCFAs, as well as modulate renal metabolism and inhibit ferroptosis.