Figure 4

Anti-inflammatory effects of docosahexaenoic acid (DHA) are AKT-independent and ERK-independent. Ai. Treatment of rHypoE-7 cells with DHA (100 μM, 1 hr) caused an increase in phospho-AKT (pAKT)/AKT levels relative to DMSO alone and was abolished upon pre-treatment with 1 μM Wortmannin (Wort) (ii) (DHA; white bar: 3.60 ± 0.13 and DHA + Wort; gray bar: 1.13 ± 0.03 pAKT/AKT). B. Pretreatment with DMSO (-), Wort, DHA, or Wort with DHA (W + D) for 1 hr prior to exposure to TNFα (10 ng/mL, 2 hr) and mRNA levels of IκBα (i) (DHA; white striped bar: 0.09 ± 0.01 and Wort + DHA (W + D); gray stripped bar: 0.12 ± 0.01 IκBα/histone mRNA) and TNFα (ii) (DHA: 0.83 ± 0.14 and W + D: 0.92 ± 0.08 TNFα/histone mRNA) were assessed. Basal levels of IκBα and TNFα mRNA without TNFα treatment are shown (0 hr). Ci. Treatment with 100 μM DHA enhanced phospho-ERK (pERK)/ERK levels relative which was abolished upon 1 μM Staurosporine aglycone (Stauro) pretreatment (ii) (DHA: 2.61 ± 0.29 and DHA + Stauro: 1.36 ± 0.28 pERK/total ERK). D. Stauro pretreatment abolished or reduced the effect of IκBα (i) (#; DMSO (-); white bar: 2.46 ± 0.10 and Stauro; gray bar: 0.89 ± 0.07) or TNFα (ii) (#; DMSO (-); white bar: 1.66 ± 0.18 and Stauro; gray bar: 0.33 ± 0.04) respectively.. Stauro pretreatment did not impair the ability of DHA to reduce IκBα mRNA in response to TNFα exposure (DHA; white stripped bar: 0.38 ± 0.17 and Stauro + DHA (S + D); gray stripped bar: 0.44 ± 0.08 IκBα/histone mRNA) but further reduced TNFα mRNA levels (DHA; white stripped bar: 1.19 ± 0.07 and S + D; gray stripped bar: 0.48 ± 0.03 TNFα/histone mRNA). Data are shown as mean ± SEM; **, p < 0.01; *** p < 0.01; non-significant (NS)).