Figure 6

Effects of lubiprostone (Lu), ML-7, TAP, and TAK-242 on scratching behavior (A, B, C, G), epidermis, and dermis thickness of ear tissue (D, E, F, J), respectively, in the TMA + IMO-treated group. Lu was administered at two different doses: 100 μg/kg in the TMA + IMO + Lu-h group and 20 μg/kg in the TMA + IMO + Lu-l group. White arrows indicate the time points of inhibitor treatments in A, B, C, and D. Scratching bouts (H) and serum levels of LPS (K) and corticosterone (L) were assessed in TLR4 KO male mice on a C57BL/6 background with TMA or TMA + IMO treatment. The time points of IMO stress (a total of 5 instances) are denoted by the black arrows in E. In the small box in H, scratching behaviors of TMA-and TMA + IMO-treated naïve mice of the C57BL/6 background are presented as a control. TLR4 protein (yellow-stained) in ear tissues was immunostained with PGP9.5 (red-stained, a marker for peripheral nerve fibers) in the TMA + IMO-treated group on day 8 (I). White arrows indicate representative immunostained spots of TLR4 proteins in the TMA- and TMA + IMO-treated groups. The border line between the epidermis and dermis layers is indicated by the yellow lines in F. ^*p < 0·05, ^**p < 0·01, ^***p < 0·001 versus TMA group (or TMA_KO); ^$$$p < 0·001 versus TMA + IMO-treated group. TMA, trimellitic anhydride; NOR, normal; KO, knockout; TLR4, Toll□like receptor 4; IMO, immobilization.