Background/Aims: Although recent reports suggest that Toll-like receptor (TLR) 9 is associated with the pathogenesis of polymicrobial septic acute kidney injury (AKI), it is still unclear whether and how renal TLR9 is involved in the development of polymicrobial septic AKI. This study aimed to determine whether the expression of TLR9 in mouse renal cells is related to the development of polymicrobial septic AKI. Methods: The efficacy of small interfering RNA (siRNA) targeting TLR9 was tested in a cultured murine macrophage cell line (RAW264.7 cells). The most potent siRNA was transfected into mice using the hydrodynamic method prior to the induction of polymicrobial septic AKI being induced by cecal ligation and puncture (CLP). TLR9 knockdown was determined by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting in RAW264.7 cells and kidney tissues. The levels of serum creatinine and blood urea nitrogen (BUN) and the renal histopathology assessment were determined at 6-, 12-, and 24-hour time points after CLP, and renal cell apoptosis was studied at 24 h. The 4- and 7-day survival rates of mice were also observed. Results: We found that mice developed AKI in our model of polymicrobial sepsis, despite fluid and antibiotic resuscitation, which resembles human sepsis. siRNA to TLR9 successfully silenced the induction of renal TLR9 gene and protein expression following CLP. Effective silencing of renal TLR9 expression decreased renal cell apoptosis, mitigated the severity of AKI, and increased the survival of mice. Conclusions: Our data demonstrates the induction of TLR9 expression in mouse kidney tissue following CLP. Renal cell apoptosis and AKI in our model of polymicrobial sepsis are dependent on TLR9. Thus, TLR9 may play a critical role in the pathophysiology of polymicrobial septic AKI. Copyright (C) 2013 S. Karger AG, Basel