Krüppel-like factor 4 (KLF4) is an evolutionarily conserved zinc finger-containing transcription factor. In the current study, RAW264.7 murine macrophages were treated with Escherichia coli lipopolysaccharide (LPS) to determine the expression of KLF4. A full-length cDNA or antisense oligonucleotides of KLF4 was transfected into RAW264.7 macrophages, and the expression and release of IL-10 were analyzed by RT-PCR and ELISA. The transcription and DNA binding activities of KLF4 to the IL-10 promoter were detected by the luciferase reporter and electrophoretic mobility shift assays. The results showed that treatment of RAW264.7 macrophages with LPS resulted in a dose- and time-dependent increase in KLF4 protein levels; KLF4 overexpression increased the expression of IL-10, while KLF4 inhibition decreased the expression of IL-10. The overexpression of KLF4 promoted the transcription and DNA binding activities of KLF4 to the IL-10 promoter. These results indicate that KLF4 plays an important role in regulating the expression of IL-10.