KLF5 and nuclear factor kappa B (NF-kappa B) regulate cell proliferation and inflammation. Vitamin D signaling through vitamin D receptor (VDR) exerts anti-proliferative and anti-inflammatory actions. However, an actual relationship between KLF5, NF-kappa B and VDR in the inflammation and proliferation of macrophages is still unclear. Here, we showed that LPS and proinflammatory cytokines stimulate KLF5 gene expression in macrophages, and that 1, 25(OH)(2)D-3 suppresses LPS-induced KLF5 expression and cell proliferation via upregulation of VDR expression. Mechanistic studies suggested that KLF5 interacts with p50 subunit of NF-kappa B to cooperatively induce the expressions of positive cell cycle regulators cyclin B1 and Cdk1/Cdc2 in LPS-treated macrophages. Further studies revealed that 1, 25(OH)(2)D-3-induced interaction of VDR with p50 decreases LPS-induced interaction of KLF5 with p50. Collectively, we identify a novel regulatory pathway in which 1, 25(OH)(2)D-3 induces VDR expression and promotes VDR interaction with p50 subunit of NF-kappa B, which in turn attenuates the association of KLF5 with p50 subunit of NF-kappa B and thus exerts anti-inflammatory and anti-proliferative effects on macrophages. (C) 2016 Elsevier Inc. All rights reserved.