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. 2019 Mar 29;10:285. doi: 10.3389/fphys.2019.00285

FIGURE 6.

FIGURE 6

Roles of MAPKs/NF-κB signaling pathway in the synergistic effect of CSE on the cold temperature-induced production of inflammatory cytokines in 16HBE cells. (A) Protein expression of ERK, P38, JNK, and IκBα was analyzed by western blotting in 16HBE cells exposed to medium, CSE, both CSE and 18°C with or without transfection of TRPM8 shRNA or scramble shRNA, respectively. (B) NF-KB activity was measured with luciferase assay in 16HBE cells exposed to medium, CSE, both CSE and 18°C with or without transfection of TRPM8 shRNA or scramble shRNA, respectively. (C) mRNA and protein expression of IL-6, IL-8, and TNF-α were separately measured with RT-RCR and ELISA in 16HBE cells exposed to both CSE and 18°C with pretreatment with an ERK inhibitor (U0126), a P38 inhibitor (SB203580), or a NF-κB inhibitor (BAY 11-7085). p- and t- represent phospho- and total-, respectively. Data in each group are mean ± SD; n = 3. ∗p < 0.05 vs. control, #p < 0.05 vs. cold alone, Δp < 0.05 vs. CSE alone.