Upon Hsp70 engagement, Tlr4 initiates a pro-inflammatory response via Myd88- and Trif-dependent signaling cascades. A) Expression analysis of Tlr4 signaling cascade genes in Hsp70 (5 μg/ml) and PBS-treated glial cells demonstrated increased expression of Myd88 in wild type astrocytes (**P < 0.01, n = 6; WTa and WTm referred to WT astrocytes and WT microglia; Tlr4KOa and Tlr4KOm referred to Tlr4KO astrocytes and Tlr4KO microglia). B) A comparative analysis of Tlr4, Myd88, Trif, Ripk1 gene expression in wild type and Tlr4KO glial cells suggests the preferential activation of Myd88 signaling cascade. C) The amount of Tnf in culture media from wild type, Tlr4KO, Myd88KO and TrifKO glial cells treated with Hsp70 and PBS (control) (**P < 0.01, n = 6). D) Two main downstream branches of Tlr4 signaling. Hsp70, heat shock protein 70; PBS, phosphate buffering saline; WT, wild type.