FIG 6.

Importance of TLR-MyD88-IRAK1-TRAF6 axis in GRA-mediated antileishmanial response. (A) Cells were transiently transfected with the indicated constructs, infected with L. donovani, and stimulated with GRA for 4 h. Cells were lysed and processed for p38 expression and phosphorylation by Western blotting. (B to D) To determine the effect of TLR inhibition, macrophages were transfected (24 h) with either TLR2 siRNA (B) or TLR4 siRNA (C), and TLR expression was determined by Western blotting. Cells were transiently transfected with TLR2 or TLR4 siRNA, respectively, infected with L. donovani, stimulated with GRA for 4 h, and processed for determination of NF-κB luciferase activity (D). The bands were analyzed densitometrically, and fold changes are indicated below each blot. All experiments were repeated at least three times each, and one set of representative data is shown. Error bars represent means ± SD; n = 3. ***, P < 0.001; Student's t test.