Figure 4

Role of Klf4 in mediating inflammation. Knockdown of Klf4 in BV-2 mouse microglial cells using SiRNA against Klf4 mRNA and subsequent decrease in the expression of pro-inflammatory cytokines. (A) q(RT)-PCR demonstrate a significant decrease in Klf4 mRNA levels in Si+LPS cells compared to LPS alone-treated cells. Cells treated with lipofectamine alone served as controls. The graph represents relative Klf4 mRNA expression values normalized to 18S rRNA internal control. (B) Immunoblot analysis of Klf4 protein isolated from BV-2 cells. Klf4 protein levels were decreased significantly in the Si+LPS group compared to LPS alone and to the Sc+LPS group. The graph represents Klf4 protein levels normalized to β-tubulin. No significant differences were observed in Si-alone and Sc-alone conditions compared to control cells for both mRNA and protein levels. (C-E) Cytokine bead array analysis of pro-inflammatory cytokines upon Klf4 knockdown. There is a more-than-two-fold decrease in TNF-α (C) and MCP-1 (D) levels in Klf4 knockdown samples, and a significant three-fold decrease is noticed in the case of IL-6 (E). *, **, Statistical differences in comparison to control values (* p < 0.05; ** p < 0.01) and #, Statistical differences with respect to LPS treated values, (# p < 0.01).