Background: To investigate if toll-like receptor (TLR) 4/nuclear factor-kappa B (NF-kappa B) signaling pathways mediated crush injury induced acute kidney injury (AKI) in rats, and if TAK-242 (a specific inhibitor of TLR4) attenuates the injury through inhibiting the signaling pathways. Methods: This study was divided into two parts: (1) Establish the crush injury model: 50 rats were randomly divided into control group and four crush injury groups (n = 10/group). Crush injury groups were given 3kg pressure for eight hours and were sacrificed at the time points of 0h, 6h, 12h, and 24h after relieving pressure. And (2) Select the most obvious injury group (12h group) for drug intervention group. Thirty rats were randomly divided into control group, 12h group, and 12h+TAK-242 group (n = 10/group). Two parts detection were as follows: pathological changes of kidney tissues were observed in Haematoxylin and Eosin (HE) staining. Serum creatinine, blood urea nitrogen (BUN), myoglobin (Mb), and blood potassium were examined by automatic biochemical analysis instrument. Interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) were measured by enzyme-linked immunosorbent assay (ELISA). The TLR4 messenger ribonucleic acid (mRNA), TLR4, and P65 were detected by real-time polymerase chain reaction (PCR), western blot, immunohistochemistry staining. Results: Compared with the control group, kidney tissues were damaged in crush injury groups, and most obvious in the 12h group. The level of serum creatinine, BUN, Mb, blood potassium, IL-6, TNF-alpha, and TLR4mRNA were increased in the crush injury groups and significantly increased in the 12h group (P <.05). The TLR4 and P65 were significantly increased in the 12h group (P <.05). Compared with the 12h group, kidney tissue damage was significantly reduced in the TAK-242 group (P <.05). The level of serum creatinine, BUN, Mb, blood potassium, IL-6, TNF-alpha, TLR4mRNA, TLR4, and P65 in the TAK-242 group were significantly reduced (P <.05). Conclusion: The present findings conclude that TLR4/NF-kappa B signaling pathways mediated crush injury induced AKI in rats, and TAK-242 attenuates the injury through inhibiting the signaling pathways.