Phosphatidylinositide 3-Kinase Contributes to the Anti-Inflammatory Effect of Abutilon crispum L. Medik Methanol Extract

Figure 3

Effect of Ac-ME on the upstream signaling of the NF-κB pathway. Immunoblotting was performed to determine the target of Ac-ME in modulating the inflammatory response. ((a) left panel) Phosphoprotein and total protein levels of IκBα, IKKα/β, AKT, and p85/PI3K were determined by immunoblotting analysis with specific antibodies. ((a) right panel) Relative intensity was values of ratio calculated using densitometric scanning values of p-IκBα, p-IKKα/β, p-AKT, or p-p85 and densitometric scanning values of their total proteins (IκBα, IKKα/β, AKT, or p85) by the DNR Bio-imaging system (a Gelquant software Ver. 2.7). (b, c) HEK293 cells were transfected with Myc-Syk or HA-Src and, at 36 h, Ac-ME (100 – 200 μg/ml) was treated to the transfected cells for further 12 h. The levels of phospho-p85, β-actin, Myc, or HA were identified by immunoblotting analysis. Relative intensity was values of ratio calculated using densitometric scanning value of p-p85 and densitometric scanning value of β-actin by the DNR Bio-imaging system (a Gelquant software Ver. 2.7). (d) The NO inhibitory levels of Syk and Src inhibitors (PP2 and Piceatannol) were measured with LPS-treated RAW264.7 cells. P < 0.05 and P < 0.01 compared to the control.

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